AU2002352976B2

AU2002352976B2 - Nucleic acid corresponding protein entitled 24P4C12 useful in treatment and detection of cancer

Info

Publication number: AU2002352976B2
Application number: AU2002352976A
Authority: AU
Inventors: Pia M. Challita-Eid; Wangmao Ge; Aya Jakobovits; Karen Jane Meyrick Morrison; Arthur B. Raitano
Original assignee: Agensys Inc
Current assignee: Agensys Inc
Priority date: 2002-11-27
Filing date: 2002-11-27
Publication date: 2007-11-08
Anticipated expiration: 2022-11-27
Also published as: AU2008200628B2; IL167892A; WO2004050828A3; AU2009208065A1; AU2008200628A1; AU2002352976A1; JP2006508163A; WO2004050828A2; EP1565200A4; AU2009208065B2; CA2503346A1; CA2503346C; EP1565200A2

Description

WO 2004/050828 PCT/US2002/038264 NUCLEIC ACID AND CORRESPONDING PROTEIN ENTITLED 24P4C12 USEFUL IN TREATMENT AND DETECTION OF CANCER STATEMENT OF RIGHTS TO INVENTIONS MADE UNDER FEDERALLY SPONSORED RESEARCH Not applicable.

FIELD OF THE INVENTION The invention described herein relates to a gene and its encoded protein, termed 24P4C12, expressed in certain cancers, and to diagnostic and therapeutic methods and compositions useful in the management of cancers that express 24P4C1 2 BACKGROUND OF THE INVENTION Cancer is the second leading cause of human death next to coronary disease. Worldwide, millions of people die from cancer every year. In the United States alone, as reported by the American Cancer Society, cancer causes the death of well over a half-million people annually, with over 1.2 million new cases diagnosed per year. While deaths from heart disease have been declining significantly, those resulting from cancer generally are on the rise. In the early part of the next century, cancer is predicted to become the leading cause of death.

Worldwide, several cancers stand out as the leading killers. In particular, carcinomas of the lung, prostate, breast, colon, pancreas, and ovary represent the primary causesof cancer death. These and virtually all other carcinomas share a common lethal feature. With very few exceptions, metastatic disease from a carcinoma is fatal. Moreover, even for those cancer patients who initially survive their primary cancers, common experience has shown that their lives are dramatically altered. Many cancer patients experience strong anxieties driven by the awareness of the potential for recurrence or treatment failure. Many cancer patients experience physical debilitations following treatment. Furthermore, many cancer patients experience a recurrence.

Worldwide, prostate cancer is the fourth most prevalent cancer in men. In North America and Northern Europe, it is by far the most common cancer in males and is the second leading cause of cancer death in men. In the United States alone, well over 30,000 men die annually of this disease second only to lung cancer. Despite the magnitude of these figures, there is still no effective treatment for metastatic prostate cancer. Surgical prostatectomy, radiation therapy, hormone ablation therapy, surgical castration and chemotherapy continue to be the main treatment modalities.

Unfortunately, these treatments are ineffective for many and are often associated with undesirable consequences.

On the diagnostic front, the lack of a prostate tumor marker that can accurately detect early-stage, localized tumors remains a significant limitation in the diagnosis and management of this disease. Although the serum prostate specific antigen (PSA) assay has been a very useful tool, however its specificity and general utility is widely regarded as lacking in several important respects.

Progress in identifying additional specific markers for prostate cancer has been improved by the generation of prostate cancer xenografts that can recapitulate different stages of the disease in mice. The LAPC (Los Angeles Prostate Cancer) xenografts are prostate cancer xenografts that have survived passage in severe combined immune deficient (SCID) mice and have exhibited the capacity to mimic the transition from androgen dependence to androgen independence (Klein et al., 1997, Nat Med. 3:402). More recently identified prostate cancer markers include PCTA-1 (Su et al., 1996, Proc. Natl.

Acad. Sci. USA 93:7252), prostate-specific membrane (PSM) antigen (Pinto et al., Clin Cancer Res 1996 Sep 2 1445- 51), STEAP (Hubert, et al, Proc Nati Acad Sci US A. 1999 Dec 7; 96(25): 14523-8) and prostate stem cell antigen (PSCA) (Reiteret 1998, Proc. Natl. Acad. Sci. USA 95: 1735).

WO 2004/050828 PCT/US2002/038264 While previously identified markers such as PSA, PSM, PCTA and PSCA have facilitated efforts to diagnose and treat prostate cancer, there is need for the identification of additional markers and therapeutic targets for prostate and related cancers in order to further improve diagnosis and therapy.

Renal cell carcinoma (RCC) accounts for approximately 3 percent of adult malignancies. Once adenomas reach a diameter of 2 to 3 cm, malignant potential exists. In the adult, the two principal malignant renal tumors are renal cell adenocarcinoma and transitional cell carcinoma of the renal pelvis or ureter. The incidence of renal cell adenocarcinoma is estimated at more than 29,000 cases in the United States, and more than 11,600 patients died of this disease in 1998. Transitional cell carcinoma is less frequent, with an incidence of approximately 500 cases per year in the United States.

Surgery has been the primary therapy for renal cell adenocarcinoma for many decades. Until recently, metastatic disease has been refractory to any systemic therapy. With recent developments in systemic therapies, particularly immunotherapies, metastatic renal cell carcinoma may be approached aggressively in appropriate patients with a possibility of durable responses. Nevertheless, there is a remaining need for effective therapies for these patients.

Of all new cases of cancer in the United States, bladder cancer represents approximately 5 percent in men (fifth most common neoplasm) and 3 percent in women (eighth most common neoplasm). The incidence is increasing slowly, concurrent with an increasing older population. In 1998, there was an estimated 54,500 cases, including 39,500 in men and 15,000 in women. The age-adjusted incidence in the United States is 32 per 100,000 for men and eight per 100,000 in women. The historic male/female ratio of 3:1 may be decreasing related to smoking patterns in women. There were an estimated 11,000 deaths from bladder cancer in 1998 (7,800 in men and 3,900 in women). Bladder cancer incidence and mortality strongly increase with age and will be an increasing problem as the population becomes more elderly.

Most bladder cancers recur in the bladder. Bladder cancer is managed with a combination of transurethral resection of the bladder (TUR) and intravesical chemotherapy or immunotherapy. The multifocal and recurrent nature of bladder cancer points out the limitations of TUR. Most muscle-invasive cancers are not cured by TUR alone. Radical cystectomy and urinary diversion is the most effective means to eliminate the cancer but carry an undeniable impact on urinary and sexual function. There continues to be a significant need for treatment modalities that are beneficial for bladder cancer patients.

An estimated 130,200 cases of colorectal cancer occurred in 2000 in the United States, including 93,800 cases of colon cancer and 36,400 of rectal cancer. Coloreotal cancers are the third most common cancers in men and women.

Incidence rates declined significantly during 1992-1996 per year). Research suggests that these declines have been due to increased screening and polyp removal, preventing progression of polyps to invasive cancers. There were an estimated 56,300 deaths (47,700 from colon cancer, 8,600 from rectal cancer) in 2000, accounting for about 11% of all U.S.

cancer deaths.

At present, surgery is the most common form of therapy for colorectal cancer, and for cancers that have not spread, it is frequently curative. Chemotherapy, or chemotherapy plus radiation, is given before or after surgery to most patients whose cancer has deeply perforated the bowel wall or has spread to the lymph nodes. A permanent colostomy (creation of an abdominal opening for elimination of body wastes) is occasionally needed for colon cancer and is infrequently required for rectal cancer. There continues to be a need for effective diagnostic and treatment modalities for colorectal cancer.

There were an estimated 164,100 new cases of lung and bronchial cancer in 2000, accounting for 14% of all U.S.

cancer diagnoses. The incidence rate of lung and bronchial cancer is declining significantly in men, from a high of 86.5 per 100,000 in 1984 to 70.0 in 1996. In the 1990s, the rate of increase among women began to slow. In 1996, the incidence rate in women was 42.3 per 100,000.

WO 2004/050828 PCT/US2002/038264 Lung and bronchial cancer caused an estimated 156,900 deaths in 2000, accounting for 28% of all cancer deaths.

During 1992-1996, mortality from lung cancer declined significantly among men per year) while rates for women were still significantly increasing per year). Since 1987, more women have died each year of lung cancer than breast cancer, which, for over 40 years, was the major cause of cancer death in women. Decreasing lung cancer incidence and mortality rates most likely resulted from decreased smoking rates over the previous 30 years; however, decreasing smoking patterns among women lag behind those of men. Of concern, although the declines in adult tobacco use have slowed, tobacco use in youth is increasing again.

Treatment options for lung and bronchial cancer are determined by the type and stage of the cancer and include surgery, radiation therapy, and chemotherapy. For many localized cancers, surgery is usually the treatment of choice.

Because the disease has usually spread by the time it is discovered, radiation therapy and chemotherapy are often needed in combination with surgery. Chemotherapy alone or combined with radiation is the treatment of choice for small cell lung cancer; on this regimen, a large percentage of patients experience remission, which in some cases is long lasting. There is however, an ongoing need for effective treatment and diagnostic approaches for lung and bronchial cancers.

An estimated 182,800 new invasive cases of breast cancer were expected to occur among women in the United States during 2000. Additionally, about 1,400 new cases of breast cancer were expected to be diagnosed in men in 2000.

After increasing about 4% per year in the 1980s, breast cancer incidence rates in women have leveled off in the 1990s to about 110.6 cases per 100,000.

In the U.S. alone, there were an estimated 41,200 deaths (40,800 women, 400 men) in 2000 due to breast cancer.

Breast cancer ranks second among cancer deaths in women. According to the most recent data, mortality rates declined significantly during 1992-1996 with the largest decreases in younger women, both white and black. These decreases were probably the result of earlier detection and improved treatment Taking into account the medical circumstances and the patient's preferences, treatment of breast cancer may involve lumpectomy (local removal of the tumor) and removal of the lymph nodes under the arm; mastectomy (surgical removal of the breast) and removal of the lymph nodes under the arm; radiation therapy; chemotherapy; or hormone therapy.

Often, two or more methods are used in combination. Numerous studies have shown that, for early stage disease, long-term survival rates after lumpectomy plus radiotherapy are similar to survival rates after modified radical mastectomy. Significant advances in reconstruction techniques provide several options for breast reconstruction after mastectomy. Recently, such reconstruction has been done at the same time as the mastectomy.

Local excision of ductal carcinoma in situ (DCIS) with adequate amounts of surrounding normal breast tissue may prevent the local recurrence of the DCIS. Radiation to the breast and/or tamoxilen may reduce the chance of DCIS occurring in the remaining breast tissue. This is important because OCIS, if left untreated, may develop into invasive breast cancer. Nevertheless, there are serious side effects or sequelae to these treatments. There is, therefore, a need for efficacious breast cancer treatments.

There were an estimated 23,100 new cases of ovarian cancer in the United States in 2000. It accounts for 4% of all cancers among women and ranks second among gynecologic cancers. During 1992-1996, ovarian cancer incidence rates were significantly declining. Consequent to ovarian cancer, there were an estimated 14,000 deaths in 2000. Ovarian cancer causes more deaths than any other cancer of the female reproductive system.

Surgery, radiation therapy, and chemotherapy are treatment options for ovarian cancer. Surgery usually includes the removal of one or both ovaries, the fallopian tubes (salpingo-oophorectomy), and the uterus (hysterectomy). In some very early tumors, only the involved ovary will be removed, especially in young women who wish to have children. In advanced disease, an attempt is made to remove all intra-abdominal disease to enhance the effect of chemotherapy. There continues to be an important need for effective treatment options for ovarian cancer.

WO 2004/050828 PCT/US2002/038264 There were an estimated 28,300 new cases of pancreatic cancer in the United States in 2000. Over the past years, rates of pancreatic cancer have declined in men. Rates among women have remained approximately constant but may be beginning to decline. Pancreatic cancer caused an estimated 28,200 deaths in 2000 in the United States. Over the past 20 years, there has been a slight but significant decrease in mortality rates among men (about per year) while rates have increased slightly among women.

Surgery, radiation therapy, and chemotherapy are treatment options for pancreatic cancer. These treatment options can extend survival and/or relieve symptoms in many patients but are not likely to produce a cure for most. There is a significant need for additional therapeutic and diagnostic options for pancreatic cancer.

SUMMARY OF THE INVENTION The present invention relates to a gene, designated 24P4C12, that has now been found to be over-expressed in the cancer(s) listed in Table I. Northern blot expression analysis of 24P4C12 gene expression in normal tissues shows a restricted expression pattern in adult tissues. The nucleotide (Figure 2) and amino acid (Figure 2, and Figure 3) sequences of 24P4C12 are provided. The tissue-related profile of 24P4C12 in normal adult tissues, combined with the over-expression observed in the tissues listed in Table I, shows that 24P4C12 is aberrantly over-expressed in at least some cancers, and thus serves as a useful diagnostic, prophylactic, prognostic, and/or therapeutic target for cancers of the tissue(s) such as those listed in Table I.

The invention provides polynucleotides corresponding or complementary to all or part of the 24P4C12 genes, mRNAs, andlor coding sequences, preferably in isolated form, including polynucleotides encoding 24P4C12-related proteins and fragments of 4, 5, 6, 7, 8, 9, 10, 11, 12, 13,14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, or more than 25 contiguous amino adds; at least 30, 35, 40, 45, 50, 55, 60, 65, 70, 80, 85, 90, 95, 100 or more than 100 contiguous amino acids of a 24P4C12-related protein, as well as the peptides/proteins themselves; DNA, RNA, DNA/RNA hybrids, and related molecules, polynuceotides or oligonucleotides complementary or having at least a 90% homology to the 24P4C12 genes or mRNA sequences or parts thereof, and polynucleotides or oligonucleotides that hybridize to the 24P4C12 genes, mRNAs, or to 24P4C12-encoding polynucleotides. Also provided are means for isolating cDNAs and the genes encocing 24P4C12.

Recombinant DNA molecules containing 24P4C12 polynudeotides, cells transformed or transduced with such molecules, and host-vector systems for the expression of 24P4C12 gene products are also provided. The invention further provides antibodies that bind to 24P4C12 proteins and polypeptide fragments thereof, including polyclonal and monodonal antibodies, murine and other mammalian antibodies, chimeric antibodies, humanized and fully human antibodies, and antibodies labeled with a detectable marker or therapeutic agent. In certain embodiments, there is a proviso that the entire nucleic acid sequence of Figure 2 is not encoded and/or the entire amino acid sequence of Figure 2 is not prepared. In certain embodiments, the entire nucleic acid sequence of Figure 2 is encoded and/or the entire amino acid sequence of Figure 2 is prepared, either of which are in respective human unit dose forms.

The invention further provides methods for detecting the presence and status of 24P4C12 polynudeotides and proteins in various biological samples, as well as methods for identifying cells that express 24P4C12. A typical embodiment of this invention provides methods for monitoring 24P4C12 gene products in a tissue or hematology sample having or suspected of having some form of growth dysregulation such as cancer.

The invention further provides various immunogenic or therapeutic compositions and strategies for treating cancers that express 24P4C12 such as cancers of tissues listed in Table I, including therapies aimed at inhibiting the transcription, translation, processing or function of 24P4C12 as well as cancer vaccines. In one aspect, the invention provides compositions, and methods comprising them, for treating a cancer that expresses 24P4C12 in a human subject wherein the composition comprises a carrier suitable for human use and a human unit dose of one or more than one agent that inhibits WO 2004/050828 PCT/US2002/038264 the production or function of 24P4C12. Preferably, the carrier is a uniquely human carrier. In another aspect of the invention, the agent is a moiety that is immunoreactive with 24P4C12 protein. Non-limiting examples of such moieties include, but are not limited to, antibodies (such as single chain, monoclonal, polyclonal, humanized, chimeric, or human antibodies), functional equivalents thereof (whether naturally occurring or synthetic), and combinations thereof. The antibodies can be conjugated to a diagnostic or therapeutic moiety. In another aspect, the agent is a small molecule as defined herein.

In another aspect, the agent comprises one or more than one peptide which comprises a cytotoxic T lymphocyte (CTL) epitope that binds an HLA class I molecule in a human to elicit a CTL response to 24P4C12 and/or one or more than one peptide which comprises a helper T lymphocyte (HTL) epitope which binds an HLA class II molecule in a human to elicit an HTL response. The peptides of the invention may be on the same or on one or more separate polypeptide molecules. In a further aspect of the invention, the agent comprises one or more than one nucleic acid mclecule that expresses one or more than one of the CTL or HTL response stimulating peptides as described above. In yet another aspect of the invention, the one or more than one nucleic acid molecule may express a moiety that is immunologically reactive with 24P4C12 as described above. The one or more than one nucleic acid molecule may also be, or encodes, a molecule that inhibits production of 24P4C12. Non-limiting examples of such molecules include, but are not limited to, those complementary to a nucleotide sequence essential for production of 24P4C12 antisense sequences or molecules that form a triple helix with a nucleotide double helix essential for 24P4C12 production) or a ribozyme effective to lyse 24P4C12 mRNA.

Note that to determine the starting position of any peptide set forth in Tables VIII-XXI and XXII to XLIX (collectively HLA Peptide Tables) respective to its parental protein, variant 1, variant 2, etc., reference is made to three factors: the particular variant the length of the peptide in an HLA Peptide Table, and the Search Peptides in Table VII. Generally, a unique Search Peptide is used to obtain HLA peptides of a particular for a particular variant. The position of each Search Peptide relative to its respective parent molecule is listed in Table VII. Accordingly, if a Search Peptide begins at position one must add the value "X 1" to each position in Tables VIII-XXI and XXII to XLIX to obtain the actual position of the HLA peptides in their parental molecule. For example, if a particular Search Peptide begins at position 150 of its parental molecule, one must add 150 1, 149 to each HLA peptide amino acid position to calculate the position of that amino acid in the parent molecule.

One embodiment of the invention comprises an HLA peptide, that occurs at least twice in Tables VIII-XXI and XXII to XLIX collectively, or an oligonucleotide that encodes the HLA peptide. Another embodiment of the invention comprises an HLA peptide that occurs at least once in Tables VIII-XXI and at least once in tables XXII to XLIX, or an oligonudeotide that encodes the HLA peptide.

Another embodiment of the invention is antibody epitopes, which comprise a peptide regions, or an oligonucleotide encoding the peptide region, that has one two, three, four, or five of the following characteristics: i) a peptide region of at least 5 amino acids of a particular peptide of Figure 3, in any whole number increment up to the full length of that protein in Figure 3, that includes an amino acid position having a value equal to or greater than 0.6, 0.7, 0.8, 0.9, or having a value equal to 1.0, in the Hydrophilicity profile of Figure ii) a peptide region of at least 5 amino acids of a particular peptide of Figure 3, in any whole number increment up to the full length of that protein in Figure 3, that includes an amino acid position having a value equal to or less than 0.5, 0.4, 0.3, 0.2, 0.1, or having a value equal to 0.0, in the Hydropathicity profile of Figure 6; iii) a peptide region of at least 5 amino acids of a particular peptide of Figure 3, in any whole number increment up to the full length of that protein in Figure 3, that includes an amino acid position having a value equal to or greater than 0.6, 0.7,0.8, 0.9, or having a value equal to 1.0, In the Percent Accessible Residues profile of Figure 7; WO 2004/050828 PCT/US2002/038264 iv) a peptide region of at least 5 amino acids of a particular peptide of Figure 3, in any whole number increment up to the full length of that protein in Figure 3, that includes an amino acid position having a value equal to or greater than 0.6, 0.7, 0.8, 0.9, or having a value equal to 1.0, in the Average Flexibility profile of Figure 8; or v) a peptide region of at least 5 amino acids of a particular peptide of Figure 3, in any whole number increment up to the full length of that protein in Figure 3, that includes an amino acid position having a value equal to or greater than 0.6, 0.7, 0.8, 0.9, or having a value equal to 1.0, in the Beta-turn profile of Figure 9.

BRIEF DESCRIPTION OF THE FIGURES Figure 1. The 24P4C12 SSH sequence of 160 nucleotides.

Figure 2. A) The cDNA and amino acid sequence of 24P4C12 variant 1 (also called "24P4C12 v.1" or "24P4C12 variant is shown in Figure 2A. The start methionine is underlined. The open reading frame extends from nucleic acid 6- 2138 including the stop codon.

B) The cDNA and amino acid sequence of 24P4C12 variant 2 (also called "24P4C12 is shown in Figure 2B.

The codon for the start methionine is underlined. The open reading frame extends from nucleic acid 6-2138 including the stop codon.

C) The cDNA and amino acid sequence of 24P4C12 variant 3 (also called '24P4C12 is shown in Figure 2C.

D) The cDNA and amino acid sequence of 24P4C12 variant 4 (also called "24P4C12 is shown in Figure 2D.

The coder for the start methionine is underlined. The open reading frame extends from nucleic acid 6-2138 including the stop codon.

E) The cDNA and amino acid sequence of 24P4C12 variant 5 (also called "24P4C12 is shown in Figure 2E.

F) The cDNA and amino acd sequence of 24P4C12 variant 6 (also called "24P4C12 is shown in Figure 2F.

G) The cDNA and amino acid sequence of 24P4C12 variant 7 (also called "24P4C12 is shown in Figure 2G.

The codon for the start methionine is underlined. The open reading frame extends from nucleic acid 6-1802 including the stop codon.

H) The cDNA and amino acid sequence of 24P4C12 variant 8 (also called "24P4C12 is shown in Figure 2H.

The codon for the start methionine is underlined. The open reading frame extends from nucleic acid 6-2174 including the stop codon.

I) The cDNA and amino acid sequence of 24P4C12 variant 9 (also called "24P4C12 v.9) is shown in Figure 21.

The codon for the start methionine is underlined. The open reading frame extends from nucleic acid 6-2144 including the stop codon.

Figure 3.

A) Amino acid sequence of 24P4C12 v.1 is shown in Figure 3A; it has 710 amino acids.

B) The amino acid sequence of 24P4C12 v.3 is shown in Figure 3B; it has 710 amino acids.

C) The amino acid sequence of 24P4C12 v.5 is shown in Figure 3C; it has 710 amino acids.

D) The amino acid sequence of 24P4C12 v.6 is shown in Figure 3D; it has 710 amino acids.

WO 2004/050828 PCT/US2002/038264 E) The amino acid sequence of 24P4C12 v.7 is shown in Figure 3E; it has 598 amino acids.

F) The amino acid sequence of 24P4C12 v.8 is shown in Figure 3F; it has 722 amino acids.

G) The amino acid sequence of 24P4C12 v.9 is shown in Figure 3G; it has 712 amino adds. As used herein, a reference to 24P4C12 includes all variants thereof, including those shown in Figures 2, 3, 10, and 11, unless the context clearly indicates otherwise.

Figure 4. Alignment or 24P4C12 with human choline transporter-like protein 4 (CTL4) (gi114249468).

.Figure 5. Hydrophilicity amino acid profile of 24P4C12 determined by computer algorithm sequence analysis using the method of Hopp and Woods (Hopp Woods 1981. Proc. Natl. Acad. Sci. U.S.A. 78:3824-3828) accessed on the Protscale website located on the World Wide Web at (.expasy.ch/cgi-binlprotscale.pl) through the ExPasy molecular biology server.

Figure 6. Hydropathicity amino acid profile of 24P4C12 determined by computer algorithm sequence analysis using the method of Kyte and Doolittle (Kyte Doolittle 1982. J. Mol. Biol. 157:105-132) accessed on the ProtScale website located on the World Wide Web at (.expasy.ch/cgi-binlprotscale.pl) through the ExPasy molecular biology server.

Figure7. Percent accessible residues amino acid profile of 24P4C12 determined by computer algorithm sequence analysis using the method of Janin (Janin 1979 Nature 277:491-492) accessed on the ProtScale website located on the World Wide Web at (.expasy.ch/cgi-bin/protscale.pl) through the ExPasy molecular biology server.

Figure Average flexibility amino acid profile of 24P4C12 determined by computer algorithm sequence analysis using the method of Bhaskaran and Ponnuswamy (Bhaskaran and Ponnuswamy 1988. Int. J. Pept. Protein Res.

32:242-255) accessed on the ProtScale website located on the World Wide Web at (.expasy.ch/cgi-bin/protscale.pl) through the ExPasy molecular biology server.

Figure 9. Beta-turn amino acid profile of 24P4C12 determined by computer algorithm sequence analysis using the method of Deleage and Roux (Deleage, Roux B. 1987 Protein Engineering 1:289-294) accessed on the ProtScale website located on the World Wide Web at (.expasy.chlcgi-bin/protscalo.pl) through the ExPasy molecular biology server.

Figure 10. Schematic alignment of SNP variants of 24P4C12. Variants 24P4C12 v.2 through v.6 are variants with single nucleotide differences. Though these SNP variants are shown separately, they could also occur in any combinations and in any transcript variants that contains the base pairs. Numbers correspond to those of 24P4C12 v.1. Black box shows the same sequence as 24P4C12 v.1. SNPs are indicated above the box.

Figure 11. Schematic alignment of protein variants of 24P4C12. Protein variants correspond to nucleotide variants. Nucleotide variants 24P4C12 v.2, v.4 in Figure 10 code for the same protein as 24P4C12 v.1. Nucleotide variants 24P4C12 v.7, v.8 and v.9 are splice variants of v.1, as shown in Figure 12. Single amino acid differences were indicated above the boxes. Black boxes represent the same sequence as 24P4C12 v.1. Numbers underneath the box correspond to 24P4C12 v.1.

Figure 12. Exon compositions of transcript variants of 24P4C12. Variant 24P4C12 v.7, v.8 and v.9 are transcript variants of 24P4C12 v.1. Variant 24P4C12 v.7 does not have exons 10 and 11 of variant 24P4C12 v.1. Variant 24P4C12 v.8 extended 36 bp at the 3' end of exon 20 of variant 24P4C12 v.1. Variant 24P4C12 v,9 had a longer exon 12 and shorter exon 13 as compared to variant 24P4C12 v.1. Numbers in underneath the boxes correspond to those of 24P4C12 v.1.

Lengths of introns and exons are not proportional.

Figure 13. Secondary structure and transmembrane domains prediction for 24P4C12 protein variant 1 (SEQ ID NO: 112). A: The secondary structure of 24P4C12 protein variant 1 was predicted using the HNN Hierarchical Neural Network method (Guermeur, 1997, http:llpbil.ibcp.frlcgi-bin/npsa.automat.pl?page=npsa_nn.html), accessed from the ExPasy molecular biology server (http:llwww.expasy.chitoolsl). This method predicts the presence and location of alpha helices, extended strands, and random coils from the primary protein sequence. The percent of the protein in a given WO 2004/050828 PCT/US2002/038264 secondary structure is also listed. B: Schematic representation of the probability of existence of transmembrane regions and orientation of 24P4C12 variant 1 based on the TMpred algorithm of Hofmann and Stoffel which utilizes TMBASE (K.

Hofmann, W. Stoffel. TMBASE A database of membrane spanning protein segments Biol. Chem. Hoppe-Seyler 374:166, 1993). C: Schematic representation of the probability of the existence of transmembrane regions and the extracellular and intracellular orientation of 24P4C12 variant 1 based on the TMHMM algorithm of Sonnhammer, von Heijne, and Krogh (Erik L.L. Sonnhammer, Gunnar von Heijne, and Anders Krogh: A hidden Markov model for predicting transmembrane helices in protein sequences. In Proc. of Sixth Int. Conf. on Intelligent Systems for Molecular Biology, p 175-182 Ed J. Glasgow, T.

Littlejohn, F. Major, R. Lathrop, D. Sankoff, and C. Sensen Menlo Park, CA: AAAI Press, 1998). The TMpred and TMHMM algorithms are accessed from the ExPasy molecular biology server (http://www.expasy.ch/toolsl).

Figure 14. 24P4C12 Expression by RT-PCR. First strand cDNA was generated from vital pool 1 (kidney, liver and lung), vital pool 2 (colon, pancreas and stomach), a pool of prostate cancer xenografts (LAPC-4AD, LAPC-4AI, LAPC-9AD and LAPC-9AI), prostate cancer pool, bladder cancer pool, kidney cancer pool, colon cancer pool, ovary cancer pool, breast cancer pool, and cancer metastasis pool Normalization was performed by PCR using primers to actin. Semi-quantitative PCR, using primers to 24P4C12, was performed at 26 and 30 cycles of amplification. Results show strong expression of 24P4C12 in prostate cancer pool and ovary cancer pool. Expression was also detected in prostate cancer xenografts, bladder cancer pool, kidney cancer pool, colon cancer pool, breast cancer pool, cancer metastasis pool, vital pool 1, and vital pool 2.

Figure 15. Expression of 24P4C12 in normal tissues. Two multiple tissue northern blots (Clontech) both with 2 ug of mRNAllane were probed with the 24P4C12 sequence. Size standards in kilobases (kb) are indicated on the side. Results show expression of 24P4C12 in prostate, kidney and colon. Lower expression is detected in pancreas, lung and placenta amongst all 16 normal tissues tested.

Figure 16. Expression of 24P4C12 in Prostate Cancer Xenografts and Cell Lines. RNA was extracted from a panel of cell lines and prostate cancer xenografts (PrEC, LAPC-4AD, LAPC-4AI, LAPC-9AD, LAPC-9AI, LNCaP, PC-3, DU145, TsuPr, and LAPC-4CL). Northern blot with 10 ug of total RNAllane was probed with 24P4C12 SSH sequence. Size standards in kilobases (kb) are indicated on the side. The 24P4C12 transcript was detected in LAPC-4AD, LAPC-4AI, LAPC- 9AD, LAPC-9AI, LNCaP, and LAPC-4 CL.

Figure 17. Expression of 24P4C12 in Patient Cancer Specimens and Normal Tissues. RNA was extracted from a pool of prostate cancer specimens, bladder cancer specimens, colon cancer specimens, ovary cancer specimens, breast cancer specimens and cancer metastasis specimens, as well as from normal prostate normal bladder normal kidney and normal colon Northern blot with 10 pg of total RNAllane was probed with 24P4C12 SSH sequence.

Size standards in kilobases (kb) are indicated on the side. Strong expression of 24P4C12 transcript was detected in the patient cancer pool specimens, and in normal prostate but not in the other normal tissues tested.

Figure 18. Expression of 24P4C12 in Prostate Cancer Patient Specimens. RNA was extracted from normal prostate prostate cancer patient tumors and their matched normal adjacent tissues (Nat). Norther blots with 10 ug of total RNA were probed with the 24P4C12 SSH fragment. Size standards in kilobases are on the side. Results show expression of 24P4C12 in normal prostate and all prostate patient tumors tested.

Figure 19. Expression of 24P4C12 in Colon Cancer Patient Specimens. RNA was extracted from colon cancer cell lines (CL: Colo 205, LoVo, and SK-CO-), normal colon colon'cancer patient tumors and their matched normal adjacent tissues (Nat). Northem blots with 10 ug of total RNA were probed with the 24P4C12 SSH fragment. Size standards in kilobases are on the side. Results show expression of 24P4C12 in normal colon and all colon patient tumors tested. Expression was detected in the cell lines Colo 205 and SK-CO-, but not in LoVo.

WO 2004/050828 PCT/US2002/038264 Figure 20. Expression of 24P4C12 in Lung Cancer Patient Specimens. RNA was extracted from lung cancer cell lines (CL: CALU-1, A427, NCI-H82, NCI-H146), normal lung lung cancer patient tumors and their matched normal adjacent tissues (Nat), Northern blots with 10 ug of total RNA were probed with the 24P4C12 SSH fragment. Size standards in kilobases are on the side. Results show expression of 24P4C12 in lung patient tumors tested, but not in normal lung. Expression was also detected in CALU-1, but not in the other cell lines A427, NCI-H82, and NCI-H146.

Figure 21. Expression of 24P4C12 in breast and stomach human cancer specimens. Expression of 24P4C12 was assayed in a panel of human stomach and breast cancers and their respective matched normal tissues on RNA dot blots. 24P4C12 expression was seen in both stomach and breast cancers. The expression detected in normal adjacent tissues (isolated from diseased tissues) but not in normal tissues (isolated from healthy donors) may indicate that these tissues are not fully normal and that 24P4C12 may be expressed in early stage tumors.

Figure 22. 24P4C12 Expression in a large panel of Patient Cancer Specimens. First strand cDNA was prepared from a panel of ovary patient cancer specimens uterus patient cancer specimens prostate cancer specimens bladder cancer patient specimens lung cancer patient specimens pancreas cancer patient specimens colon cancer specimens and kidney cancer specimens Normalization was performed by PCR using primers to actin.

Semi-quantitative PCR, using primers to 24P4C12, was performed at 26 and 30 cycles of amplification. Samples were run on an agarose gel, and PCR products were quantitated using the Alphalmager software. Expression was recorded as absent, low, medium or strong. Results show expression of 24P4C12 in the majority of patient cancer specimens tested, 73.3% of ovary patient cancer specimens, 83.3% of uterus patient cancer specimens, 95.0% of prostate cancer specimens, 61.1% of bladder cancer patient specimens, 80.6% of lung cancer patient specimens, 87.5% of pancreas cancer patient specimens, 87.5% of colon cancer specimens, 68.4% of of clear cell renal carcinoma, 100% of papillary renal cell carcinoma.

Figure 23. 24P4C12 expression in transduced cells. PC3 prostate cancer cells, NIH-3T3 mouse cells and 300.19 mouse cells were transduced with 24P4C12 .pSRa retroviral vector. Cells were selected in neomycin for the generation of stable cell lines. RNA was extracted following selection in neomycin. Northern blots with 10 ug of total RNA were probed with the 24P4C12 SSH fragment. Results show strong expression of 24P4C12 in 24P4C12.pSRa transduced PC3, 3T3 and 300.19 cells, but not in the control cells transduced with the parental pSRa construct.

Figure 24. Expression of 24P4C12 in 293T cells. 293T cell were transiently transfected with either pCDNA3.1 Myc-His tagged expression vector, the pSRD expression vector each encoding the 24P4C12 variant 1 cDNA or a control neo vector. Cells were harvested 2 days later and analyzed by Western blot with anti-24P4C12 pAb or by Flow cytometry (B) on fixed and permeabilized 293T cells with either the anti-24P4C12 pAb or anti-His pAb followed by a PE-conjugated antirabbit IgG secondary Ab. Shown is expression of the monomeric and aggregated forms of 24P4C12 by Western blot and a fluorescent shift of 24P4C12-293T cells compared to control neo cells when stained with the anti-24P4C12 and anti-His pAbs which are directed to the intracellular NH3 and COOH termini, respectively.

Figure 25. Expression and detection of 24P4012 in stably transduced PC3 cells. PC3 cells were infected with retrovirus encoding the 24P4C12 variant 1 cDNA and stably transduced cells were derived by G418 selection. Cells were then analyzed by Westem blot or immunohistochemistry with anti-24P4C12 pAb. Shown with an arrow on the Western blot is expression of a -94 kD band representing 24P4C12 expressed in PC3-24P4C12 cells but not in control neo cells. Immunohistochemical analysis shows specific staining of 24P4C12-PC3 cells and not PC3-neo cells which is competed away competitor peptide to which the pAb was derived.

Figure 26. Expression of recombinant 24P4C12 antigens in 293T cells. 293T cells were transiently transfected with Tag5 His-tagged expression vectors encoding either amino acids 59-227 or 319-453 of 24P4C12 variant 1 or a control vector. 2 days later supematants were collected and cells harvested and lysed. Supernatants and lysates were then subjected to Western blot analysis using an anti-His pAb. Shown is expression of the recombinant Tag5 59-227 protein in WO 2004/050828 PCT/US2002/038264 both the supematant and lysate and the Tag5 319-453 protein in the cell lysate. These proteins are purified and used as antigens for generation of 24P4C12-specific antibodies.

Figure 27. Monoclonal antibodies detect 24P4C12 protein expression in 293T cells by low cytometry. 293T cells were transfected with either pCDNA 3.1 His-tagged expression vector for 24P4C12 or a control neo vector and harvested 2 days later. Cells were fixed, permeabilized, and stained with a 1:2 dilution of supernatants of the indicated hybridomas generated from mice immunized with 300.19-24P4C12 cells or with anti-His pAb. Cells were then stained with a PEconjugated secondary Ab and analyzed by flow cytometry. Shown is a fluorescent shift of 293T-24P4C12 cells but not control neo cells demonstrating specific recognition of 24P4C12 protein by the hybridoma supernatants.

Figure 28. Shows expression of 24P4C12 Enhances Proliferation. PC3 and 3T3 were grown overnight in low FBS. Cells were then incubated in low or 10% FBS as indicated. Proliferation was measured by Alamar Blue.

Figure 29. Detection of 24P4C12 protein by immunohistochemistry in prostate cancer patient specimens.

Prostate adenocarcinoma tissue and its matched normal adjacent tissue were obtained from prostate cancer patients. The results showed strong expression of 24P4C12 in the tumor cells and normal epithelium of the prostate cancer patients' tissue (panels low grade prostate adenocarcinoma, high grade prostate adenocarcinoma, normal tissue adjacent to tumor). The expression was detected mostly around the cell membrane indicating that 24P4C12 is membrane associated in prostate tissues.

Figure 30. Detection of 24P4C12 protein by immunohistochemistry in various cancer patient specimens. Tissue was obtained from patients with colon adenocarcinoma, breast ductal carcinoma, lung adenocardnoma, bladder transitional cell carcinoma, renal clear cell carcinoma and pancreatic adenocarcinoma. The results showed expression of 24P4C12 in the tumor cells of the cancer patients' tissue (panel colon adenocarcinoma, lung adenocarcinoma, breast ductal carcinoma, bladder transitional carcinoma, renal clear cell carcinoma, pancreatic adenocarcinoma).

Figure 31. Shows 24P4C12 Enhances Tumor Growth in SCID Mice. 1 x 106 PC3-24P4C12 cells were mixed with Matrigel and injected on the right and left subcutaneous flanks of 4 male SCID mice per group. Each data point represents mean tumor volume Figure 32. Shows 24P4C12 Enhances Tumor Growth in SCID Mice. 1 x 106 3T3-24P4C12 cells were mixed with Matrigel and injected on the right subcutaneous flanks of 7 male SCID mice per group. Each data point represents mean tumor volume DETAILED DESCRIPTION OF THE INVENTION Outline of Sections Definitions II.) 24P4C12 Polynucleotides II.A.) Uses of 24P4C12 Polynucleotides II.A.1.) Monitoring of Genetic Abnormalities II.A.2.) Antisense Embodiments II.A.3.) Primers and Primer Pairs II.A.4.) Isolation of 24P4C12-Encoding Nucleic Acid Molecules Recombinant Nucleic Acid Molecules and Host-Vector Systems Il.) 24P4C12-related Proteins III.A.) Motif-bearing Protein Embodiments 111.8.) Expression of 24P4Ci2-related Proteins II.C.) Modifications of 24P4C12-related Proteins III.D.) Uses of 24P4C12-related Proteins WO 2004/050828 PCT/US2002/038264 IV.) 24P4C12 Antibodies 24P4C12 Cellular Immune Responses VI.) 24P4C12 Transgenic Animals VII.) Methods for the Detection of 24P4C12 VIII.) Methods for Monitoring the Status of 24P4C12-related Genes and Their Products IX.) Identification of Molecules That Interact With 24P4C12 Therapeutic Methods and Compositions Anti-Cancer Vaccines 24P4C12 as a Target for Antibody-Based Therapy 24P4C12 as a Target for Cellular Immune Responses X.C.1. Minigene Vaccines X.C.2. Combinations of CTL Peptides with Helper Peptides X.C.3. Combinations of CTL Peptides with T Cell Priming Agents X.C.4. Vaccine Compositions Comprising DC Pulsed with CTL andlor HTL Peptides Adoptive Immunotherapy Administration of Vaccines for Therapeutic or Prophylactic Purposes XI.) Diagnostic and Prognostic Embodiments of 24P4C12.

XII.) Inhibition of 24P4C12 Protein Function XII.A.) Inhibition of 24P4C12 With Intracellular Antibodies XII.B.) Inhibition of 24P4C12 with Recombinant Proteins XII.C.) Inhibition of 24P4C12 Transcription or Translation XII.D.) General Considerations for Therapeutic Strategies XIII.) Identification, Characterization and Use of Modulators of 24P4C12 XIV.) KITSIArticles of Manufacture Definitions: Unless otherwise defined, all terms of art, notations and other scientific terms or terminology used herein are intended to have the meanings commonly understood by those of skill in the art to which this invention pertains. In some cases, terms with commonly understood meanings are defined herein for clarity and/or for ready reference, and the inclusion of such definitions herein should not necessarily be construed to represent a substantial difference over what is generally understood in the art. Many of the techniques and procedures described or referenced herein are well understood and commonly employed using conventional methodology by those skilled in the art, such as, for example, the widely utilized molecular cloning methodologies described in Sambrook et at, Molecular Cloning: A Laboratory Manual 2nd, edition (1989) Cold.Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. As appropriate, procedures involving the use of commercially available kits and reagents are generally carried out in accordance with manufacturer defined protocols and/or parameters unless otherwise noted.

The terms "advanced prostate cancer", 'locally advanced prostate cancer", "advanced disease" and "locally advanced disease" mean prostate cancers that have extended through the prostate capsule, and are meant to include stage C disease under the American Urological Association (AUA) system, stage C1 C2 disease under the Whitmore-Jewett system, and stage T3 T4 and N+ disease under the TNM (tumor, node, metastasis) system. In general, surgery is not recommended for patients with locally advanced disease, and these patients have substantially less favorable outcomes compared to patients having clinically localized (organ-confined) prostate cancer. Locally advanced disease is clinically WO 2004/050828 PCT/US2002/038264 identified by palpable evidence of induration beyond the lateral border of the prostate, or asymmetry or induration above the prostate base. Locally advanced prostate cancer is presently diagnosed pathologically following radical prostatectomy if the tumor invades or penetrates the prostatic capsule, extends into the surgical margin, or invades the seminal vesicles.

"Altering, the native glycosylation pattern" is intended for purposes herein to mean deleting one or more carbohydrate moieties found in native sequence 24P4C12 (either by removing the underlying glycosylation site or by deleting the glycosylation by chemical andlor enzymatic means), and/or adding one or more glycosylation sites that are not present in the native sequence 24P4C12. In addition, the phrase includes qualitative changes in the glycosylation of the native proteins, involving a change in the nature and proportions of the various carbohydrate moieties present.

The term "analog" refers to a molecule which is structurally similar or shares similar or corresponding attributes with another molecule a 24P4C12-related protein). For example, an analog of a 24P4C12 protein can be specifically bound by an antibody or T cell that specifically binds to 24P4C12.

The term 'antibody" is used in the broadest sense. Therefore, an "antibody" can be naturally occurring or man-made such as monoclonal antibodies produced by conventional hybridoma technology. Anti-24P4C12 antibodies comprise monoclonal and polydonal antibodies as well as fragments containing the antigen-binding domain and/or one or more complementarity determining regions of these antibodies.

An 'antibody fragment" is defined as at least a portion of the variable region of the immunoglobulin molecule that binds to its target, the antigen-binding region. In one embodiment it specifically covers single ant-24P4C12 antibodies and clones thereof (including agonist, antagonist and neutralizing antbodies) and anti-24P4C12 antibody compositions with polyepitopic specificity.

The term "codon optimized sequences" refers to nucleotide sequences that have been optimized for a particular host species by replacing any codons having a usage frequency of less than about 20%. Nucleotide sequences that have been optimized for expression in a given host species by elimination of spurious polyadenylation sequences, elimination of exon/intron splicing signals, elimination of transposon-like repeats and/or optimization of GC content in addition to codon optimization are referred to herein as an "expression enhanced sequences." A "combinatorial library" is a collection of diverse chemical compounds generated by either chemical synthesis or biological synthesis by combining a number of chemical "building blocks" such as reagents. For example, a linear combinatorial chemical library, such as a polypeptide mutein) library, is formed by combining a set of chemical building blocks called amino acids in every possible way for a given compound length the number of amino acids in a polypeptide compound). Numerous chemical compounds are synthesized through such combinatorial mixing of chemical building blocks (Gallop et al., J. Med. Chem. 37(9): 1233-1251 (1994)).

Preparation and screening of combinatorial libraries is well known to those of skill in the art. Such combinatorial chemical libraries include, but are not limited to, peptide libraries (see, U.S. Patent No. 5,010,175, Furka, Pept. Prot.

Res. 37:487-493 (1991), Houghton et al., Nature, 354:84-88 (1991)), peptoids (PCT Publication No WO 91/19735), encoded peptides (PCT Publication WO 93/20242), random bio- oligomers (PCT Publication WO 92/00091), benzodiazepines (U.S.

Pat. No. 5,288,514), diversomers such as hydantoins, benzodiazepines and dipeptides (Hobbs et al., Proc. Nat. Acad. Sci.

,USA 90:6909-6913(1993)), vinylogous polypeptides (Hagihara et al., J. Amer. Chem. Soc. 114:6568 (1992)), nonpeptidal peptidomimetics with a Beta-D-Glucose scaffolding (Hirschmann et al., J. Amer. Chem. Soc. 114:9217-9218 (1992)), analogous organic syntheses of small compound libraries (Chen et al., J. Amer. Chem. Soc. 116:2661 (1994)), oligocarbamates (Cho, et al., Science 261:1303 (1993)), and/or peptidyl phosphonates (Campbell et al., J. Org. Chem.

59:658 (1994)). See, generally, Gordon et al., J. Med. Chem. 37:1385 (1994), nucleic acid libraries (see, Stratagene, Corp.), peptide nucleic acid libraries (see, U.S. Patent 5,539,083), antibody libraries (see, Vaughn et al., Nature Biotechnology 14(3): 309-314 (1996), and PCT/US96/10287), carbohydrate libraries (see, Liang et al., Science WO 2004/050828 PCT/US2002/038264 274:1520-1522 (1996), and U.S. Patent No. 5,593,853), and small organic molecule libraries (see, benzodiazepines, Baum, C&EN, Jan 18, page 33 (1993); isoprenoids, U.S. Patent No. 5,569,588; thiazolidinones and metathiazanones, U.S.

Patent No. 5,549,974; pyrrolidines, U.S. Patent Nos. 5,525,735 and 5,519,134; morpholino compounds, U.S. Patent No.

5,506, 337; benzodiazepines, U.S. Patent No. 5,288,514; and the like).

Devices for the preparation of combinatorial libraries are commercially available (see, 357 NIPS, 390 NIPS, Advanced Chem Tech, Louisville KY; Symphony, Rainin, Woburn, MA; 433A, Applied Biosystems, Foster City, CA; 9050, Plus, Millipore, Bedford, NIA). A number of well-known robotic systems have also been developed for solution phase chemistries. These systems include automated workstations such as the automated synthesis apparatus developed by Takeda Chemical Industries, LTD. (Osaka, Japan) and many robotic systems utilizing robotic arms (Zymate H, Zymark Corporation, Hopkinton, Mass.; Orca, Hewlett-Packard, Palo Alto, Calif.), which mimic the manual synthetic operations performed by a chemist. Any of the above devices are suitable for use with the present invention. The nature and implementation of modifications to these devices (if any) so that they can operate as discussed herein will be apparent to persons skilled in the relevant art. In addition, numerous combinatorial libraries are themselves commercially available (see, ComGenex, Princeton, NJ; Asinex, Moscow, RU; Tripos, Inc., St. Louis, MO; ChemStar, Ltd, Moscow, RU; 3D Pharmaceuticals, Exton, PA; Martek Biosciences, Columbia, MD; etc.).

The term "cytotoxic agent" refers to a substance that inhibits or prevents the expression activity of cells, function of cells andfor causes destruction of cells. The term is intended to include radioactive isotopes chemotherapeutic agents, and toxins such as small molecule toxins or enzymatically active toxins of bacterial, fungal, plant or animal origin, including fragments and/or variants thereof. Examples of cytotoxic agents include, but are not limited to auristatins, auromycins, maytansinoids, yttrium, bismuth, ricin, ricin A-chain, combrestatin, duocarmycins, dolostatins, doxorubicin, daunorubicin, taxol, cisplatin, cc1065, ethidium bromide, mitomycin, etoposide, tenoposide, vincristine, vinblastine, colchicine, dihydroxy anthracin dione, actinomycin, diphtheria toxin, Pseudomonas exotoxin (PE) A, PE40, abrin, abrin A chain, modeccin A chain, alpha-sarcin, gelonin, mitogellin, retstrictocin, phenomycin, enomycin, curicin, crotin, calicheamicin, Sapaonaria officinalis inhibitor, and glucocorticoid and other chemotherapeutic agents, as well as radioisotopes such as At 211 131, 125, Y90, y Rei, Re 1 Sm' 5 3 Bi 21 2 o' 21 3, P32 and radioactive isotopes of Lu including Lul T 7 Antibodies may also be conjugated to an anticancer pro-drug activating enzyme capable of converting the pro-drug to its active form.

The "gene product" is sometimes referred to herein as a protein or mRNA. For example, a "gene product of the invention" is sometimes referred to herein as a "cancer amino acid sequence", "cancer protein", 'protein of a cancer listed in Table a "cancer mRNA", "mRNA of a cancer listed in Table etc. In one embodiment, the cancer protein is encoded by a nucleic acid of Figure 2. The cancer protein can be a fragment, or alternatively, be the full-length protein to the fragment encoded by the nucleic acids of Figure 2. In one embodiment, a cancer amino acid sequence is used to determine sequence identity or similarity. In another embodiment, the sequences are naturally occurring allelic variants of a protein encoded by a nucleic acid of Figure 2. In another embodiment, the sequences are sequence variants as further described herein.

"High throughput screening" assays for the presence, absence, quantification, or other properties of particular :nucleic acids or protein products are well known to those of skill in the art. Similarly, binding assays and reporter gene assays are similarly well known. Thus, U.S. Patent No. 5,559,410 discloses high throughput screening methods for proteins; U.S. Patent No. 5,585,639 discloses high throughput screening methods for nucleic acid binding in arrays); while U.S. Patent Nos. 5,576,220 and 5,541,061 disclose high throughput methods of screening for ligand/antibody binding.

In addition, high throughput screening systems are commercially available (see, Amersham Biosciences, Piscataway, NJ; Zymark Corp., Hopkinton, MA; Air Technical Industries, Mentor, OH; Beckman Instruments, Inc. Fullerton, CA; Precision Systems, Inc., Natck, MA; etc.). These systems typically automate entire procedures, including all sample WO 2004/050828 PCT/US2002/038264 and reagent pipetting, liquid dispensing, timed incubatons, and final readings of the microplate in detector(s) appropriate for the assay. These configurable systems provide high throughput and rapid start up as well as a high degree of flexibility and customization. The manufacturers of such systems provide detailed protocols for various high throughput systems. Thus, Zymark Corp. provides technical bulletins describing screening systems for detecting the modulation of gene transcription, ligand binding, and the like.

The term "homolog" refers to a molecule which exhibits homology to another molecule, by for example, having sequences of chemical residues that are the same or similar at corresponding positions.

"Human Leukocyte Antigen" or "HLA" is a human class I or class II Major Histocompatibility Complex (MHC) protein (see, Stites, et al, IMMUNOLOGY, 8" ED., Lange Publishing, Los Altos, CA (1994).

The terms "hybridize", "hybridizing", "hybridizes" and the like, used in the context of polynucleotides, are meant to refer to conventional hybridization conditions, preferably such as hybridization in 50% formamidel6XSSCO/.1'% SDSI100 pg/ml ssDNA, in which temperatures for hybridization are above 37 degrees C and temperatures for washing in 0.1XSSC/0.1 SDS are above 55 degrees C.

The phrases "isolated" or "biologically pure" refer to material which is substantially or essentially free from components which normally accompany the material as it is found in its native state. Thus, isolated peptides in accordance with the invention preferably do not contain materials normally associated with the peptides in their in situ environment. For example, a polynucleotide is said to be "isolated" when it is substantially separated from contaminant polynucleotides that correspond or are complementary to genes other than the 24P4C12 genes or that encode polypeptides other than 24P4C12 gene product or fragments thereof. A skilled artisan can readily employ nucleic acid isolation procedures to obtain an isolated 24P4C12 polynucleotide. A protein is said to be "isolated," for example, when physical, mechanical or chemical methods are employed to remove the 24P4C12 proteins from cellular constituents that are normally associated with the protein. A skilled artisan can readily employ standard purfication methods to obtain an isolated 24P4C12 protein. Alternatively, an isolated protein can be prepared by chemical means.

The term "mammal" refers to any organism classified as a mammal, incuding mice, rats, rabbits, dogs, cats, cows, horses and humans. In one embodiment of the invention, the mammal is a mouse. In another embodiment of the invention, the mammal is a human.

The terms "metastatic prostate cancer" and "metastatic disease" mean prostate cancers that have spread to .regional lymph nodes or to distant sites, and are meant to include stage D disease under the AUA system and stage TxNxM+ under the TNM system. As is the case with locally advanced prostate cancer, surgery is generally not indicated for patients with metastatic disease, and hormonal (androgen ablation) therapy is a preferred treatment modality. Patients with metastatic prostate cancer eventually develop an androgen-refractory state within 12 to 18 months of treatment initiation.

Approximately half of these androgen-refractory patients die within 6 months after developing that status. The most common site for prostate cancer metastasis is bone. Prostate cancer bone metastases are often osteoblastic rather than osteolytic resulting in net bone formation). Bone metastases are found most frequently in the spine, followed by the femur, pelvis, rib cage, skull and humerus. Other common sites for metastasis include lymph nodes, lung, liver and brain. Metastatic ;prostate cancer is typically diagnosed by open or laparoscopic pelvic lymphadenectomy, whole body radionuclide scans, skeletal radiography, and/or bone lesion biopsy.

The term "modulator' or "test compound" or 'drug candidate" or grammatical equivalents as used herein describe any molecule, protein, oligopeptide, small organic molecule, polysaccharide, polynudeotide, etc., to be tested for the capacity to directly or indirectly alter the cancer phenotype or the expression of a cancer sequence, a nucleic acid or protein sequences, or effects of cancer sequences signaing, gene expression, protein interaction, etc.) In one aspect, a modulator will neutralize the effect of a cancer protein of the invention. By "neutralize" is meant that an activity of a protein WO 2004/050828 PCT/US2002/038264 is Inhibited or blocked, along with the consequent effect on the cell. In another aspect, a modulator will neutralize the effect of a gene, and its corresponding protein, of the invention by normalizing levels of said protein. In preferred embodiments, modulators alter expression profiles, or expression profile nucleic adds or proteins provided herein, or downstream effector pathways. In one embodiment, the modulator suppresses a cancer phenotype, e.g. to a normal tissue fingerprint. In another embodiment, a modulator induced a cancer phenotype. Generally, a plurality of assay mixlures is run in parallel with different agent concentrations to obtain a differential response to the various concentrations. Typically, one of these concentrations serves as a negative control, at zero concentration or below the level of detection.

Modulators, drug candidates or test compounds encompass numerous chemical classes, though typically they are organic molecules, preferably small organic compounds having a molecular weight of more than 100 and less than about 2,500 Daltons. Preferred small molecules are less than 2000, or less than 1500 or less than 1000 or less than 500 D.

Candidate agents comprise functional groups necessary for structural interaction with proteins, particularly hydrogen bonding, and typically include at least an amine, carbonyl, hydroxyl or carboxyl group, preferably at least two of the functional chemical groups. The candidate agents often comprise cyclical carbon or heterocyclic structures and/or aromatic or polyaromatic structures substituted with one or more of the above functional groups. Modulators also comprise biomolecules such as peptides, saccharides, fatty acids, steroids, purines, pyrimidines, derivatives, structural analogs or combinations thereof. Particularly preferred are peptides. One class of modulators are peptides, for example of from about five to about amino acids, with from about five to about 20 amino acids being preferred, and from about 7 to about 15 being particularly preferred. Preferably, the cancer modulatory protein is soluble, includes a non-lransmembrane region, and/or, has an Nterminal Cys to aid in solubility. In one embodiment, the C-terminus of the fragment is kept as a free acid and the N-terminus is a free amine to aid in coupling, to cysteine. In one embodiment, a cancer protein of the invention is conjugated to an immunogenic agent as discussed herein. In one embodiment, the cancer protein is conjugated to BSA. The peptides of the invention, of preferred lengths, can be linked to each other or to other amino acids to create a longer peptide/protein.

The modulatory peptides can be digests of naturally occurring proteins as is outlined above, random peptides, or "biased' random peptides. In a preferred embodiment, peptide/protein-based modulators are antibodies, and fragments thereof, as defined herein.

Modulators of cancer can also be nucleic acids. Nucleic acid modulating agents can be naturally occurring nucleic acids, random nucleic acids, or "biased" random nucleic acids. For example, digests of prokaryotic or eukaryotic genomes .can be used in an approach analogous to that outlined above for proteins.

The term "monoclonal antibody' refers to an antibody obtained from a population of substantially homogeneous antibodies, the antibodies comprising the population are identical except for possible naturally occurring mutations that are present in minor amounts.

A "motif, as in biological motif of a 24P4C12-related protein, refers to any pattern of amino acids forming part of the primary sequence of a protein, that is associated with a particular function protein-protein interaction, protein-DNA interaction, etc) or modification that is phosphorylated, glycosylated or amidated), or localization secretory sequence, nuclear localization sequence, etc.) or a sequence that is correlated with being immunogenic, either humorally or cellularly. A motif can be either contiguous or capable of being aligned to certain positions that are generally correlated with a certain function or property. In the context of HLA motifs, "motif" refers to the pattern of residues in a peptide of defined length, usually a peptide of from about 8 to about 13 amino acids for a class I HLA motif and from about 6 to about 25 amino acids for a class II HLA motif, which is recognized by a particular HLA molecule. Peptide motifs for HLA binding are typically different for each protein encoded by each human HLA allele and differ in the pattern of the primary and secondary anchor residues.

WO 2004/050828 PCT/US2002/038264 A "pharmaceutical excipient comprises a material such as an adjuvant, a carrier, pH-adjusting and buffering agents, tonicity adjusting agents, wetting agents, preservative, and the like.

"Pharmaceutically acceptable" refers to a non-toxic, inert, and/or composition that is physiologically compatible with humans or other mammals.

The term "polynucleotide" means a polymeric form of nucleotides of at least 10 bases or base pairs in length, either ribonucleotides or deoxynucleotides or a modified form of either type of nucleotide, and is meant to include single and double stranded forms of DNA and/or RNA. In the art, this term if often used interchangeably with "oligonucleotide". A polynucleotide can comprise a nucleotide sequence disclosed herein wherein thymidine as shown for example in Figure 2, can also be uracil this definition pertains to the differences between the chemical structures of DNA and RNA, in particular the observation that one of the four major bases in RNA is uracil instead of thymidine The term "polypeptide" means a polymer of at least about 4, 5, 6, 7, or 8 amino acids. Throughout the specification, standard three letter or single letter designations for amino acids are used. In the art, this term is often used interchangeably with "peptide" or "protein".

An HLA "primary anchor residue" is an amino acid at a specific position along a peptide sequence which is understood to provide a contact point between the immunogenic peptide and the HLA molecule. One to three, usually two, primary anchor residues within a peptide of defined length generally defines a "motif for an immunogenic peptide. These residues are understood to fit in close contact with peptide binding groove of an HLA molecule, with their side chains buried in specific pockets of the binding groove. In one embodiment, for example, the primary anchor residues for an HLA class I molecule are located at position 2 (from the amino terminal position) and at the carboxyl terminal position of a 8, 9, 10, 11, or 12 residue peptide epitope in accordance with the invention. Alternatively, in another embodiment, the primary anchor residues of a peptide binds an HLA class II molecule are spaced relative to each other, rather than to the termini of a peptide, where the peptide is generally of at least 9 amino acids in length. The primary anchor positions for each motif and supermotif are set forth in Table IV. For example, analog peptides can be created by altering the presence or absence of particular residues in the primary and/or secondary anchor positions shown in Table IV. Such analogs are used to modulate the binding affinity and/or population coverage of a peptide comprising a particular HLA motif or supermotif.

"Radioisotopes" include, but are not limited to the following (non-limiting exemplary uses are also set forth): Examples of Medical Isotopes: Isotope Description of use Actinium-225 (AC-225) See Thorium-229 (Th-229) Actinium-227 (AC-227) Parent of Radium-223 (Ra-223) which is an alpha emitter used to treat metastases in the skeleton resulting from cancer breast and prostate cancers), and cancer radioimmunotherapy Bismuth-212 (Bi.212) See Thorium-228 (Th-228) Bismuth-213 (Bi-213) See Thorium-229 (Th-229) Cadmium-109 (Cd-109) Cancer detection WO 2004/050828 PCT/US2002/038264 Radiation source for radiotherapy of cancer, for food irradiators, and for sterilization of medical supplies Copper-64 (Cu-64) A positron emitter used for cancer therapy and SPECT imaging Copper-67 (Cu-67) Betalgamma emitter used in cancer radioimmunotherapy and diagnostic studies breast and colon cancers, and lymphoma) Dysprosium-166 (Dy-166) Cancer radioimmunotherapy Erbium-169 (Er-169) Rheumatoid arthritis treatment, particularly for the small joints associated with fingers and toes Europium-152 (Eu-152) Radiation source for food irradiation and for sterilization of medical supplies Europium-154 (Eu-154) Radiation source for food irradiation and for sterilization of medical supplies Gadolinium-153 (Gd-153) Osteoporosis detection and nuclear medical quality assurance devices Gold-198 (Au-198) Implant and intracavity therapy of ovarian, prostate, and brain cancers Holmium-166 (Ho-166) Multiple myeloma treatment in targeted skeletal therapy, cancer radioimmunotherapy, bone marrow ablation, and rheumatoid arthritis treatment Iodine-125 (1-125) Osteoporosis detection, diagnostic imaging, tracer drugs, brain cancer treatment, radiolabeling, tumor imaging, mapping of receptors in the brain, interstitial radiation therapy, brachytherapy for treatment of prostate cancer, determination of glomerular filtration rate (GFR), determination of plasma volume, detection of deep vein thrombosis of the legs Iodine-131 (1-131) Thyroid function evaluation, thyroid disease detection, treatment of thyroid cancer as well as other nonmalignant thyroid diseases Graves disease, goiters, and hyperthyroidism), treatment of leukemia, lymphoma, and other forms of cancer breast cancer) using radioimmunotherapy lridium-192 (lr-192) Brachytherapy, brain and spinal cord tumor treatment, treatment of blocked arteries arteriosclerosis and restenosis), and implants for breast and prostate tumors Lutetium-177 (Lu-177) WO 2004/050828 PCT/US2002/038264 Cancer radioimmunotherapy and treatment of blocked arteries arteriosclerosis and restenosis) Molybdenum-99 (Mo-99) Parent of Technetium-99m (Tc-99m) which is used for imaging the brain, liver, lungs, heart, and other organs.

Currently, Tc-99m is the most widely used radioisotope used for diagnostic imaging of various cancers and diseases involving the brain, heart, liver, lungs; also used in detection of deep vein thrombosis of the legs Osmium-194 (Os-194) Cancer radioimmunotherapy Palladium-103 (Pd-103) Prostate cancer treatment Platinum-195m (Pt-195m) Studies on biodistribution and metabolism of cisplatin, a chemotherapeutic drug Phosphorus-32 (P-32) Polycythemia rubra vera (blood cell disease) and leukemia treatment, bone cancer diagnosis/treatment; colon, pancreatic, and liver cancer treatment; radiolabeling nucleic acids for in vitro research, diagnosis of superficial tumors, treatment of blocked arteries arteriosclerosis and restenosis), and intracavity therapy Phosphorus-33 (P-33) Leukemia treatment, bone disease diagnosis/treatment, radiolabeling, and treatment of blocked arteries arteriosclerosis and restenosis) Radium-223 (Ra-223) See Actinium-227 (Ac-227) Rhenium-186 (Re-186) Bone cancer pain relief, rheumatoid arthritis treatment, and diagnosis and treatment of lymphoma and bone, breast, colon, and liver cancers using radicimmunotherapy Rhenium-188 (Re-188) Cancer diagnosis and treatment using radioimmunotherapy, bone cancer pain relief, treatment of rheumatoid arthritis, and treatment of prostate cancer Rhodium-105 (Rh-105) Cancer radioimmunotherapy Samarium-145 (Sm-145) Ocular cancer treatment Samarium-153 (Sm-153) Cancer radioimmunotherapy and bone cancer pain relief Scandium-47 (Sc-47) Cancer radioimmunotherapy and bone cancer pain relief WO 2004/050828 PCT/US2002/038264 Radiotracer used in brain studies, imaging of adrenal cortex by gamma-scintigraphy, lateral localtions of steroid secreting tumors, pancreatic scanning, detection of hyperactive parathyroid glands, measure rate of bile acid loss from the endogenous pool Bone cancer detection and brain scans Strontium-89 (Sr-89) Bone cancer pain relief, multiple myeloma treatment, and osteoblastic therapy Technetium-99m (Tc-99m) See Molybdenum-99 (Mo-99) Thorium-228 (Th-228) Parent of Bismuth-212 (Bi-212) which is an alpha emitter used in cancer radioimmunotherapy Thorium-229 (Th-229) Parent of Actinium-225 (Ac-225) and grandparent of Bismuth-213 (Bi-213) which are alpha emitters used in cancer radioimmunotherapy Thulium-1 (Tm-170) Gamma source for blood irradiators, energy source for implanted med cal devices Tin-i 17m (Sn-117m) Cancer immunotherapy and bone cancer pain relief Tungsten-1 88 (W-188) Parent for Rhenium-188 (Re-188) which is used for cancer diagnostics/treatment, bone cancer pain relief, rheumatoid arthritis treatment, and treatment of blocked arteries arteriosclerosis and restenosis) Xenon-1 27 (Xe-127) Neuroimaging of brain disorders, high resolution SPECT studies, pulmonary function tests, and cerebral blood flow studies Yttorbium-175 (Yb.175) Cancer radioimmunotherapy Microseeds obtained from irradiating Yttrium-89 (Y-89) for liver cancer treatment Yttrium-91 (Y-91) A gamma-emitting label for Yttrium-90 (Y-90) which is used for cancer radioimmunotherapy ymphoma, breast, colon, kidney, lung, ovarian, prostate, pancrealic, and inoperable liver cancers) WO 2004/050828 PCT/US2002/038264 By "randomized" or grammatical equivalents as herein applied to nucleic acids and proteins is meant that each nucleic acid and peptide consists of essentially random nucleotides and amino acids, respectively. These random peptides (or nucleic acids, discussed herein) can incorporate any nucleotide or amino acid at any position. The synthetic process can be designed to generate randomized proteins or nucleic acids, to alow the formation of all or most of the possible combinations over the length of the sequence, thus forming a library of randomized candidate bioactive proteinaceous agents.

In one embodiment, a library is "fully randomized,' with no sequence preferences or constants at any position. In another embodiment, the library is a "biased random" library. That is, some positions within the sequence either are held constant, or are selected from a lim ted number of possibilities. For example, the nucleotides or amino acid residues are randomized within a defined class, of hydrophobic amino acids, hydrophilic residues, sterically biased (either small or large) residues, towards the creation of nucleic acid binding domains, the creation of cysteines, for cross-linking, prolines for SH-3 domains, serines, threonines, tyrosines or histidines for phosphorylation sites, etc., or to purines, etc.

A "recombinant" DNA or RNA molecule is a DNA or RNA molecule that has been subjected to molecular manipulation in vitro.

Non-limiting examples of small molecules include compounds that bind or interact with 24P4C12, ligands including hormones, neuropeptides, chemokines, odorants, phospholipids, and functional equivalents thereof that bind and preferably inhibit 24P4C12 protein function. Such non-limiting small molecules preferably have a molecular weight of less than about kDa, more preferably below about 9, about 8, about 7, about 6, about 5 or about 4 kDa. In certain embodiments, small molecules physically associate with, or bind, 24P4C12 protein; are not found in naturally occurring metabolic pathways; andlor are more soluble in aqueous than non-aqueous solutions "Stringency" of hybridization reactions is readily determinable by one of ordinary skill in the art, and generally is an empirical calculation dependent upon probe length, washing temperature, and salt concentration. In general, longer probes require higher temperatures for proper annealing, while shorter probes need lower temperatures. Hybridization generally depends on the ability of denatured nucleic acid sequences to reanneal when complementary strands are present in an environment below their melting temperature. The higher the degree of desired homology between the probe and hybridizable sequence, the higher the relative temperature that can be used. As a result, it follows that higher relative temperatures would tend to make the reaction conditions more stringent, while lower temperatures less so. For additional details and explanation of stringency of hybridization reactions, see Ausubel et al., Current Protocols in Molecular Biology, Wiley Interscience Publishers, (1995).

"Stringent conditions" or "high stringency conditions", as defined herein, are identified by, but not limited to, those that: employ low ionic strength and high temperature for washing, for example 0.015 M sodium chloride/0.0015 M sodium citratel0.1% sodium dodecyl sulfate at 500C; employ during hybridization a denaturing agent, such as formamide, for example, 50% (viv) formamide with 0.1% bovine serum albuminl0.1% Ficoll/0. 1% polyvinylpyrrolidonel50 mM sodium phosphate buffer at pH 6.5 with 750 mM sodium chloride, 75 mM sodium citrate at 42 OC; or employ 50% formamide, 5 x SSC (0.75 M NaCI, 0.075 M sodium citrate), 50 mM sodium phosphate (pH 0.1% sodium pyrophosphate, 5 x Denhardt's solution, sonicated salmon sperm DNA (50 ig/ml), 0.1% SDS, and 10% dextran sulfate at 42 oC, with washes at 42oC in 0.2 x SSC (sodium chloridelsodium. citrate) and 50% formamide at 55 oC, followed by a high-stringency wash consisting of 0.1 x SSC containing EDTA at 55 OC. "Moderately stringent conditions" are described by, but not limited to, those in Sambrook et al., Molecular Cloning: A Laboratory Manual, New York: Cold Spring Harbor Press, 1989, and include the use of washing solution and hybridization conditions temperature, ionic strength and %SDS) less stringent than those described above. An example of moderately stringent conditions is overnight incubation at 37oC in a solution comprising: 20% formamide, 5 x SSC (150 mM NaCI, 15 mM trisodium citrate), 50 mM sodium phosphate (pH 5 x WO 2004/050828 PCT/US2002/038264 Denhardrs solution, 10% dextran sulfate, and 20 mg/mL denatured sheared salmon sperm DNA, followed by washing the filters in 1 x SSC at about 37-50°C. The skilled artisan will recognize how to adjust the temperature, ionic strength, etc. as necessary to accommodate factors such as probe length and the like.

An HLA "supermotif is a peptide binding specificity shared by HLA molecules encoded by two or more HLA alleles.

Overall phenotypic frequencies of HLA-supertypes in different ethnic populations are set forth in Table IV The nonlimiting constituents of various supetypes are as follows: A2. A*0201, A*0202, A'0203, A*0204, A* 0205, A'0206, A*6802, A'6901, A*0207 A3: A3, A11, A31, A'3301, A*6801, A*0301, A*1101, A'3101 B: B7, B'3501-03, 8*51, B'5301, B*5401, B*5501, 8*5502, B*5601, B'6701, B'7801, 8*0702, B*5101, B'5602 B44: B*3701, B*4402, B*4403, B*60 (B*4001), B61 (8*4006) Ai: A*0102, A*2604, A'3601, A*4301, A'8001 A24: A'24, A*30, A*2403, A'2404, A*3002, A'3003 B27: B*1401-02, B*1503, B*1509, B'1510, B*1518, B*3801-02, 8*3901, B3902, 83903-04, B4801-02, B*7301, B'2701-08 B58: B*1516, B*1517, B*5701, B*5702, B58 B62: B*4601, B52, 8*1501 (862), B*1502 (B75), B*1513 (B77) Calculated population coverage afforded by different HLA-supertype combinations are set forth in Table IV As used herein 'to treat' or "therapeutic" and grammatically related terms, refer to any improvement of any consequence of disease, such as prolonged survival, less morbidity, and/or a lessening of side effects which are the byproducts of an alternative therapeutic modality; full eradication of disease is not required.

A "transgenic animal" a mouse or rat) is an animal having cells that contain a transgene, which transgene was introduced into the animal or an ancestor of the animal at a prenatal, an embryonic stage. A "transgene" is a DNA that is integrated into the genome of a cell from which a transgenic animal develops.

As used herein, an HLA or cellular immune response "vaccine" is a composition that contains or encodes one or more peptides of the invention. There are numerous embodiments of such vaccines, such as a cocktail of one or more individual peptides; one or more peptides of the invention comprised by a polyepitopic peptide; or nucleic acids that encode such individual peplides or polypeptides, a minigene that encodes a polyepitopic peptide. The "one or more peptides' can include any whole unit integer from 1-150 or more, at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 65, 70, 75, 80, 85, 90, 95, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, or 150 or more peptides of the invention.

The peptides or polypeptides can optionally be modified, such as by lipidation, addition of targeting or other sequences. HLA class I peptides of the invention can be admixed with, or linked to, HLA class II peptides, to facilitate activation of both cytotoxic T lymphocytes and helper T lymphocytes. HLA vaccines can also comprise peptide-pulsed antigen presenting cells, dendritic cells.

The term "variant" refers to a molecule that exhibits a variation from a described type or norm, such as a protein that has one or more different amino add residues in the corresponding position(s) of a specifically described protein the 24P4C12 protein shown in Figure 2 or Figure 3. An analog is an example of a variant protein. Splice isoforms and single nudeotides polymorphisms (SNPs) are further examples of variants.

The "24P4C12-related proteins" of the invention include those specifically identified herein, as well as allelic variants, conservative substitution variants, analogs and homologs that can be isolated/generated and characterized without undue experimentation following the methods outlined herein or readily available in the art. Fusion proteins that combine parts of different 24P4C12 proteins or fragments thereof, as well as fusion proteins of a 24P4C12 protein and a heterologous polypeptide WO 2004/050828 PCT/US2002/038264 are also induded. Such 24P4C12 proteins are collectively referred to as the 24P4C12-related proteins, the proteins of the invention, or 24P4C12. The term "24P4C12-related protein" refers to a polypeptide fragment or a 24P4C12 protein sequence of 4, 6, 7, 8, 9,10, 11, 12,13, 14,15, 16, 17,18, 19, 20, 21, 22, 23, 24, 25, or more than 25 amino adds; or, at least 30, 35, 40, 55, 60, 65,70, 80,85,90, 95,100,105,110,115,120,125,130, 135, 140,145,150,155,160,165,170,175,180,185, 190,195, 200, 225, 250, 275, 300, 325, 350, 375, 400, 425, 450,475, 500, 525, 550, 575, 600, 625, 650, or 664 or more amino acids.

II.) 24P4C12 Polynucleotides One aspect of the invention provides polynucleotides corresponding or complementary to all or part of a 24P4C12 gene, mRNA, and/or coding sequence, preferably in isolated form, including polynucleotides encoding a 24P4C12-related protein and fragments thereof, DNA, RNA, DNA/RNA hybrid, and related molecules, polynucleotides or oligonucleotides complementary to a 24P4C12 gene or mRNA sequence or a part thereof, and polynucleotides or oligonucleotides that hybridize to a 24P4C12 gene, mRNA, or to a 24P4C12 encoding polynucleotide (collectively, "24P4C12 polynucleotides"). In all instances when referred to in this section, T can also be U in Figure 2.

Embodiments of a 24P4C12 polynucleotide indude: a 24P4C12 polynucleotide having the sequence shown in Figure 2, the nucleotide sequence of 24P4C12 as shown in Figure 2 wherein T is U; at least 10 contiguous nucleotides of a polynucleotide having the sequence as shown in Figure 2; or, at least 10 contiguous nucleotides of a polynuclectide having the sequence as shown in Figure 2 where T is U. For example, embodiments of 24P4C12 nucleotides comprise, without limitation: a polynucleotide comprising, consisting essentially of, or consisting of a sequence as shown in Figure 2, wherein T can also be U; (II) a polynucleotide comprising, consisting essentially of, or consisting of the sequence as shown in Figure 2A, from nucleotide residue number 6 through nucleotide residue number 2138, including the stop codon, wherein Tcan also be U; (111) a polynucleotide comprising, consisting essentially of, or consisting of the sequence as shown in Figure 2B, from nucleotide residue number 6 through nucleotide residue number 2138, including the stop codon, wherein Tcan also be U; (IV) a polynucleotide comprising, consisting essentially of, or consisting of the sequence as shown in Figure 2C, from nucleotide residue number6 through nucleotide residue number 2138, including the a stop codon, wherein T can also be U; a polynucleotide comprising, consisting essentially of, or consisting of the sequence as shown in Figure 2D, from nucleotide residue number 6 through nucleotide residue number 2138, including the stop codon, wherein T can also be U; (VI) a polynucleotide comprising, consisting essentially of, or consisting of the sequence as shown in Figure 2E, from nucleotide residue number 6 through nucleotide residue number 2138, including the stop codon, wherein Tcan also be U; WO 2004/050828 PCT/US2002/038264 (VII) a polynucleotide comprising, consisting essentially of, or consisting of the sequence as shown in Figure 2F, from nucleotide residue number 6 through nucleotide residue number 2138, including the stop codon, wherein T can also be U; (VIII) a polynucleotide comprising, consisting essentially of, or consisting of the sequence as shown in Figure 2G, from nucleotide residue number 6 through nucleotide residue number 1802, including the stop codon, wherein T can also be U; (IX) a polynucleotide comprising, consisting essentially of, or consisting of the sequence as shown in Figure 2H, from nucleotide residue number 6 through nucleotide residue number 2174, including the stop codon, wherein T can also be U; a polynucleotide comprising, consisting essentially of, or consisting of the sequence as shown in Figure 21, from nudeotide residue number 6 through nucleotide residue number 2144, including the stop codon, wherein T can also be U; (XI) a polynucleotide that encodes a 24P4C12-related protein that is at least 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 or 100% homologous to an entire amino acid sequence shown in Figure 2A-I; (XII) a polynucleotide that encodes a 24P4C12-related protein that is at least 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 or 100% identical to an entire amino acid sequence shown in Figure 2A-I; (XIII) a polynucleotide that encodes at least one peptide set forth in Tables VIII-XXI and XXII-XLIX; (XIV) a polynucleotide that encodes a peptide region of at least 5, 6, 7, 8, 9, 10, 11, 12,13,14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a peptide of Figure 3A-D in any whole number increment up to 710 that includes at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value greater than in the Hydrophilicity profile of Figure (XV) a polynucleotide that encodes a peptide region of at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a peptide of Figure 3A-D in any whole number increment up to 710 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15,16, 17, 18, 19, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value less than 0.5 in the Hydropathicity profile of Figure 6; (XVI) a polynucleotide that encodes a peptide region of at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a peptide of Figure 3A-D in any whole number increment up to 710 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13,14, 15, 16, 17,18, 19, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Percent Accessible Residues profile of Figure 7; (XVII) a polynucleotide that encodes a peptide region of at least 5, 6, 7, 8, 9,10, 11, 12, 13,14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29,30, 31,32, 33, 34, 35 amino acids of a peptide of Figure 3A-D in any whole number increment up to 710 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, WO 2004/050828 PCT/US2002/038264 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Average Flexibility profile of Figure 8; (XVIII) a polynucleotide that encodes a peptide region of at least 5, 6, 7,8,9,10,11, 12,13,14,15,16,17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a peptide of Figure 3A-D in any whole number increment up to 710 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Beta-turn profile of Figure 9; (XIX) a polynucleotide that encodes a peptide region of at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a peptide of Figure 3E in any whole number increment up to 598 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,11,12,13,14,15, 16,17,18,19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Hydrophilicity profile of Figure (XX) a polynucleotide that encodes a peptide region of at least 5, 6, 7, 8,9, 10, 11, 12, 13, 14, 15,16,17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a peptide of Figure 3E in any whole number increment up to 598 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,11, 12, 13, 14, 15,16, 17,18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acd position(s) having a value less than 0.5 in the Hydropathicity profile of Figure 6; (XXI) a polynucleotide that encodes a peptide region of at least 5, 6, 7, 8, 9, 10, 11, 12, 13,14, 15, 16,17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a peptide of Figure 3E in any whole number increment up to 598 that includes 1, 2, 3, 4, 5, 6, 7 8, 9, 10,11,12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Percent Accessible Residues profile of Figure 7; (XXII) a polynucleotide that encodes a peptide region of at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16,17,18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a peptide of Figure 3E in any whole number increment up to 598 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,11, 12,13,14, 15,16,17,18, 19, 20,21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Average Flexibility profile of Figure 8; (XXIII) a polynucleotide that encodes a peptide region of at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a peptide of Figure 3E in any whole number increment up to 598 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15,16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Betaturn profile of Figure 9 (XXIV) a polynucleotide that encodes a peptide region of at least 5, 6, 7, 8, 9,10,11,12,13,14,15,16,17,18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a peptide of Figure 3F in any whole number increment up to 722 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,11, 12, 13, 14, 15,16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Hydrophilicity profile of Figure WO 2004/050828 PCT/US2002/038264 (XXV) a polynucleotide that encodes a peptide region of at least 5, 6, 7, 8, 9,10, 11,12, 13, 14,15, 16, 17,18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a peptide of Figure 3F in any whole number increment up to 722 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,11, 12,13,14, 15,16, 17, 18,19, 20, 21,22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value less than 0.5 in the Hydropathicity profile of Figure 6; (XXVI) a polynucleotide that encodes a peptide region of at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16,17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a peptide of Figure 3F in any whole number increment up to 722 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13,14, 15, 16, 17, 18,19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Percent Accessible Residues profile of Figure 7; (XXVII) a polynucleotide that encodes a peptide region of at least 5, 6, 7, 8, 9,10,11, 12,13,14,15, 16,17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a peptide of Figure 3F in any whole number increment up to 722 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18,19, 20, 21, 22, 23, 24, 25, 26. 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Average Flexibility profile of Figure 8; (XXVIII) a polynucleotide that encodes a peptide region of at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,15, 16,17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a peptide of Figure 3F in any whole number increment up to 722 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18,19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Betaturn profile of Figure 9 (XXIX) a polynudeotide that encodes a peptide region of at least 5, 6, 7, 8, 9, 10, 11, 12,13, 14,15, 16,17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a peptide of Figure 3G in any whole number increment up to 712 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18,19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Hydrophilicity profile of Figure (XXX) a polynudeotide that encodes a peptide region of at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a peptide of Figure 3G in any whole number increment up to 712 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18,19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value less than 0.5 in the Hydropathicity profile of Figure 6; (XXXI) a polynuceotide that encodes a peptide region of at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a peptide of Figure 3G in any whole number increment up to 712 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12,13, 14, 15, 16, 17, 18,19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Percent Accessible Residues profile of Figure 7; (XXXII) a polynudeotide that encodes a peptide region of at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a peptide of Figure 3G in any whole WO 2004/050828 PCT/US2002/038264 number increment up to 712 that includes 1, 2,3,4,5,6,7, 8, 9, 10, 11, 12,13,14,15,16,17,18,19,20, 21, 22, 23,24, 25, 26,27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Average Flexibility profile of Figure 8; (XXXIII) a polynucleotide that encodes a peptide region of at least 5, 6, 7, 8, 9,10,11,12,13,14,15,16,17,18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,32, 33, 34, 35 amino acids of a peptide of Figure 3G in any whole number increment up to 712 that includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,11, 12, 13, 14, 15, 16,17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Betaturn profile of Figure 9 (XXXIV) a polynucleotide that is fully complementary to a polynucleotide of any one of (I)-(XXXIII).

(XXXV) a peptide that is encoded by any of to (XXXIII); and (XXXVI) a composition comprising a polynucleotide of any of (I)-(XXXIV) or peptide of (XXXV) together with a pharmaceutical excipient and/or in a human unit dose form.

(XXXVII) a method of using a polynucleotide of any XXXIV) or peptide of (XXXV) or a composition of (XXXVI) in a method to modulate a cell expressing 24P4C12, (XXXVIII) a method of using a polynucleotide of any (I)-(XXXIV) or peptide of (XXXV) or a composition of (XXXVI) in a method to diagnose, prophylax, prognose, or treat an individual who bears a cell expressing 24P4C12 (XXXIX) a method of using a polynucleotide of any (I)-(XXXIV) or peptide of (XXXV) or a composition of (XXXVI) in a method to diagnose, prophylax, prognose, or treat an individual who bears a cell expressing 24P4C12, said cell from a cancer of a tissue listed in Table I; (XL) a method of using a polynucleotide of any (I)-(XXXIV) or peptide of (XXXV) or a composition of (XXXVI) in a method to diagnose, prophylax, prognose, or treat a a cancer; (XLI) a method of using a polynucleotide of any (I)-(XXXIV) or peptide of (XXXV) or a composition of (XXXVI) in a method to diagnose, prophylax, prognose, or treat a a cancer of a tissue listed in Table I; and, (XLII) a method of using a polynucleotide of any (I)-(XXXIV) or peptide of (XXXV) or a composition of (XXXVI) in a method to identify or characterize a modulator of a cell expressing 24P4C12.

As used herein, a range is understood to disclose specifically all whole unit positions thereof.

Typical embodiments of the invention disclosed herein include 24P4C12 polynucleotides that encode specific portions of 24P4C12 mRNA sequences (and those which are complementary to such sequences) such as those that encode the proteins and/or fragments thereof, for example: 4, 5, 6, 7, 8, 9, 10, 11, 12, 13,14, 15, 16, 17, 18,19, 20, 21, 22, 23, 24, 25, 30, 35, 40, 45, 50, 55, 60, 65, 80, 85, 90, 95, 100,105, 110,115,120,125,130,135,140, 145, 150, 155,160,165,170,175,180, 185, 190, 195, 200, 225, 250, 275, 300, 325, 350, 375, 400, 425, 450, 475, 500, 525, 550, 575, 600, 625, 650, 675, 700, 710 or more contiguous amino acids of 24P4C12 variant 1; the maximal lengths relevant for other variants are: variant 3, 710 amino acids; variant 710 amino acids, variant 6, 710 amino acids, variant 7, 598 amino acids, variant 8, 722 amino acids, and variant 9, 712 amino acids.

WO 2004/050828 PCT/US2002/038264 For example, representative embodiments of the invention disclosed herein include: polynucleotides and their encoded peptides themselves encoding about amino acid 1 to about amino acid 10 of the 24P4C12 protein shown in Figure 2 or Figure 3, polynucleotides encoding about amino acid 10 to about amino acid 20 of the 24P4C12 protein shown in Figure 2 or Figure 3, polynucleotides encoding about amino acid 20 to about amino acid 30 of the 24P4C12 protein shown in Figure 2 or Figure 3, polynucleotides encoding about amino acid 30 to about amino acid 40 of the 24P4C12 protein shown in Figure 2 or Figure 3, polynucleotides encoding about amino acid 40 to about amino acid 50 of the 24P4C12 protein shown in Figure 2 or Figure 3, polynucleotides encoding about amino acid 50 to about amino acid 60 of the 24P4C12 protein shown in Figure 2 or Figure 3, polynucleotides encoding about amino acid 60 to about amino acid 70 of the 24P4C12 protein shown in Figure 2 or Figure 3, polynucleotides encoding about amino acid 70 to about amino acid 80 of the 24P4C12 protein shown in Figure 2 or Figure 3, polynucleotides encoding about amino acid 80 to about amino acid 90 of the 24P4C12 protein shown in Figure 2 or Figure 3, polynucleotides encoding about amino acid 90 to about amino acid 100 of the 24P4C12 protein shown in Figure 2 or Figure 3, in increments of about 10 amino adds, ending at the carboxyl terminal amino acid set forth in Figure 2 or Figure 3. Accordingly, polynucleotides encoding portions of the amino acid sequence (of about 10 amino acids), of amino acids, 100 through the carboxyl terminal amino acid of the 24P4C12 protein are embodiments of the invention. Wherein it is understood that each particular amino acid position discloses that position plus or minus five amino acid residues.

Polynucleotides encoding relatively long portions of a 24P4C12 protein are also within the scope of the invention.

For example, polynucleotides encoding from about amino acid 1 (or 20 or 30 or 40 etc.) to about amino acid 20, (or 30, or or 50 etc.) of the 24P4C12 protein "or variant" shown in Figure 2 or Figure 3 can be generated by a variety of techniques well known in the art. These polynucleotide fragments can include any portion of the 24P4C12 sequence as shown in Figure 2.

Additional illustrative embodiments of the invention disclosed herein include 24P4C12 polynucleotide fragments encoding one or more of the biological motifs contained within a 24P4C12 protein "or variant" sequence, including one or more of the motif-bearing subsequences of a 24P4C12 protein "or variant" set forth in Tables VIII-XXI and XXII-XLIX In another embodiment, typical polynucleotide fragments of the invention encode one or more of the regions of 24P4C12 protein or variant that exhibit homology to a known molecule. In another embodiment of the invention, typical polynucleotide fragments can encode one or more of the 24P4C12 protein or variant N-glycosylation sites, cAMP and cGMP-dependent protein kinase phosphorylation sites, casein kinase II phosphorylation sites or N-myristoylation site and amidation sites.

Note that to determine the starting position of any peptide set forth in Tables VIII-XXI and Tables XXII to XLIX (collectively HLA Peptide Tables) respective to its parental protein, variant 1, variant 2, etc., reference is made to three factors: the particular variant, the length of the peptide in an HLA Peptide Table, and the Search Peptides listed in Table LVII. Generally, a unique Search Peptide is used to obtain HLA peptides for a particular variant. The position of each Search Peptide relative to its respective parent molecule Is listed In Table VII. Accordingly, if a Search Peptide begins at position one must add the value "X minus 1" to each position in Tables VIII-XXI and Tables XXII-IL to obtain the actual position of the HLA peptides in their parental molecule. For example if a particular Search Peptide begins at position 150 of its parental molecule, one must add 150 1, 149 to each HLA peptide amino acid position to calculate the position of that amino acid in the parent molecule.

II.A.) Uses of 24P4C12 Polynucleotides II.A.1.) Monitoring of Genetic Abnormalities The polynudeotides of the preceding paragraphs have a number of different specific uses. The human 24P4C12 gene maps to the chromosomal location set forth in the Example entitled "Chromosomal Mapping of 24P4C12." For example, because the 24P4C12 gene maps to this chromosome, polynucleotides that encode different regions of the 24P4C12 proteins are used to characterize cytogenetic abnormalities of this chromosomal locale, such as abnormalities that are identified as being associated with various cancers. In certain genes, a variety of chromosomal abnormalities including WO 2004/050828 PCT/US2002/038264 rearrangements have been identified as frequent cytogenetic abnormalities in a number of different cancers (see e.g.

Krajinovic et al., Mutat Res. 382(3-4): 81-83 (1998); Johansson etal., Blood 86(10): 3905-3914 (1995) and Finger et al., P.N.A.S. 85(23): 9158-9162 (1988)). Thus, polynuclectides encoding specific regions of the 24P4C12 proteins provide new tools that can be used to delineate, with greater precision than previously possible, cytogenetic abnormalities in the chromosomal region that encodes 24P4C12 that may contribute to the malignant phenotype. In this context, these polynucleotides satisfy a need in the art for expanding the sensitivity of chromosomal screening in order to identify more subtle and less common chromosomal abnormalities (see e.g. Evans of Am. J. Obstet. Gynecol 171(4): 1055-1057 (1994)).

Furthermore, as 24P4C12 was shown to be highly expressed in bladder and other cancers, 24P4C12 polynucleotides are used in methods assessing the status of 24P4C12 gene products in normal versus cancerous tissues.

Typically, polynucleotides that encode specific regions of the 24P4C12 proteins are used to assess the presence of perturbations (such as deletions, insertions, point mutations, or alterations resulting in a loss of an antigen etc.) in specific regions of the 24P4C12 gene, such as regions containing one or more motifs. Exemplary assays include both RT-PCR assays as well as single-strand conformation polymorphism (SSCP) analysis (see, Marrogi et at, J. Cutan. Pathol.

26(8): 369-378 (1999), both of which utilize polynucleotides encoding specific regions of a protein to examine these regions within the protein.

II.A.2.) Antisense Embodiments Other specifically contemplated nucleic acid related embodiments of the invention disclosed herein are genomic DNA, cDNAs, ribozymes, and antisense molecules, as well as nucleic acid molecules based on an altemative backbone, or including altemative bases, whether derived from natural sources or synthesized, and include molecules capable of inhibiting the RNA or protein expression of 24P4C12. For example, antisense molecules can be RNAs or other molecules, including peptide nucleic acids (PNAs) or non-nucleic acid molecules such as phosphorothioate derivatives that specifically bind DNA or RNA in a base pair-dependent manner. A skilled artisan can readily obtain these classes of nucleic acid molecules using the 24P4C12 polynucleotides and polynudeotide sequences disclosed herein.

Antisense technology entails the administration of exogenous oligonucleotides that bind to a target polynucleotide located within the cells. The term "antisense" refers to the fact that such oligonucleotides are complementary to their intracellular targets, 24P4C12. See for example, Jack Cohen, Oligodeoxynucleotides, Antisense Inhibitors of Gene Expression, CRC Press, 1989; and Synthesis 1:1-5 (1988). The 24P4C12 antisense oligonucleotides of the present invention include derivatives such as S-oligonucleotides (phosphorcthioate derivatives or S-oligos, see, Jack Cohen, supra), which exhibit enhanced cancer cell growth inhibitory action. S-oligos (nucleoside phosphorothioates) are isoelectronic analogs of an oligonucleotide (0-oligo) in which a nonbridging oxygen atom of the phosphate group is replaced by a sulfur atom. The S-oligos of the present invention can be prepared by treatment of the corresponding O-oligos with 3H-1,2benzodithiol-3-one-1,1-dioxide, which is a sulfur transfer reagent. See, lyer, R. P. et al, J. Org. Chem. 55:4693-4698 (1990); and lyer, R. P. et al, J. Am. Chem. Soc. 112:1253-1254 (1990). Additional 24P4C12 antisense oligonucleotides of the present invention include morpholino antisense oligonucleotides known in the art (see, Partridge etal., 1996, Antisense Nucleic Acid Drug Development 6: 169-175).

The 24P4C12 antisense oligonucleotides of the present invention typically can be RNA or DNA that is complementary to and stably hybridizes with the first 100 5' codons or last 100 3' codons of a 24P4C12 genomic sequence or the corresponding mRNA. Absolute complementarity is not required, although high degrees of complementarity are preferred. Use of an oligonuceotide complementary to this region allows for the selective hybridization to 24P4C12 mRNA and not to mRNA specifying other regulatory subunits of protein kinase. In one embodiment, 24P4C12 antisense oligonucleotides of the present invention are 15 to 30-mer fragments of the antisense DNA molecule that have a sequence WO 2004/050828 PCT/US2002/038264 that hybridizes to 24P4C12 mRNA. Optionally, 24P4C12 antisense oligonucleotide is a 30-mer oligonucleotide that is complementary to a region in the first 10 5' codons or last 10 3' codons of 24P4C12. Alternatively, the antisense molecules are modified to employ ribozymes in the inhibition of 24P4C12 expression, see, L. A. Couture D. T. Stinchcomb; Trends Genet 12: 510-515 (1996).

II.A.3.) Primers and Primer Pairs Further specific embodiments of these nucleotides of the invention include primers and primer pairs, which allow the specific amplification of polynucleotides of the invention or of any specific parts thereof, and probes that selectively or specifically hybridize to nucleic acid molecules of the invention or to any part thereof. Probes can be labeled with a detectable marker, such as, for example, a radioisotope, fluorescent compound, bioluminescent compound, a chemiluminescent compound, metal chelator or enzyme. Such probes and primers are used to detect the presence of a 24P4C12 polynudeotide in a sample and as a means for detecting a cell expressing a 24P4C12 protein.

Examples of such probes include polypeptides comprising all or part of the human 24P4C12 cDNA sequence shown in Figure 2. Examples of primer pairs capable of specifically amplifying 24P4C12 mRNAs are also described in the Examples. As will be understood by the skilled artisan, a great many different primers and probes can be prepared based on the sequences provided herein and used effectively to amplify and/or detect a 24P4C12 mRNA.

The 24P4C12 polynucleotides of the invention are useful for a variety of purposes, including but not limited to their use as probes and primers for the amplification and/or detection of the 24P4C12 gene(s), mRNA(s), or fragments thereof; as reagents for the diagnosis andfor prognosis of prostate cancer and other cancers; as coding sequences capable of directing the expression of 24P4C12 polypeptides; as tools for modulating or inhibiting the expression of the 24P4C12 gene(s) and/or translation of the 24P4C12 transcript(s); and as therapeutic agents.

The present invention includes the use of any probe as described herein to identify and isolate a 24P4C12 or 24P4C12 related nucleic acid sequence from a naturally occurring source, such as humans or other mammals, as well as the isolated nucleic acid sequence per se, which would comprise all or most of the sequences found in the probe used.

II.A.4.) Isolation of 24P4C12-Encoding Nucleic Acid Molecules The 24P4C12 cDNA sequences described herein enable the isolation of other polynucleotides encoding 24P4C12 gene product(s), as well as the isolation of polynucleotides encoding 24P4C12 gene product homologs, altematively spliced isoforms, allelic variants, and mutant forms of a 24P4C12 gene product as well as polynucleotides that encode analogs of 24P4C12-related proteins. Various molecular cloning methods that can be employed to isolate full length cDNAs encoding a 24P4C12 gene are well known (see, for example, Sambrook, J. et al, Molecular Cloning: A Laboratory Manual, 2d edition, Cold Spring Harbor Press, New York, 1989; Current Protocols in Molecular Biology. Ausubel ef al., Eds., Wiley and Sons, 1995). For example, lambda phage cloning methodologies can be conveniently employed, using commercially available cloning systems Lambda ZAP Express, Stratagene). Phage cones containing 24P4C12 gene cDNAs can be identified by probing with a labeled 24P4C12 cDNA or a fragment thereof. For example, in one embodiment, a 24P4C12 cDNA Figure 2) or a portion thereof can be synthesized and used as a probe to retrieve overlapping and full-length cDNAs corresponding to a 24P4C12 gene. A 24P4C12 gene itself can be isolated by screening genomic DNA libraries, bacterial artificial chromosome libraries (BACs), yeast artifidal chromosome libraries (YACs), and the like, with 24P4C12 DNA probes or primers.

Recombinant Nucleic Acid Molecules and Host-Vector Systems The invention also provides recombinant DNA or RNA molecules containing a 24P4C12 polynudeotide, a fragment, analog or homologue thereof, including but not limited to phages, plasmids, phagemids, cosmids, YACs, BACs, as well as various viral and non-viral vectors well known in the art, and cells transformed or transfected with such recombinant DNA or RNA molecules. Methods for generating such molecules are well known (see, for example, Sambrook et al., 1989, supra).

WO 2004/050828 PCT/US2002/038264 The invention further provides a host-vector system comprising a recombinant DNA molecule containing a 24P4C12 polynudeotide, fragment analog or homologue thereof within a suitable prokaryotic or eukaryotic host cell. Examples of suitable eukaryotic host cells include a yeast cell, a plant cell, or an animal cell, such as a mammalian cell or an insect cell a baculovirus-infeclible cell such as an Sf9 or HighFive cell). Examples of suitable mammalian cells include various prostate cancer cell lines such as DU145 and TsuPrl, other transfectable or transducible prostate cancer cell lines, primary cells (PrEC), as well as a number of mammalian cells routinely used for the expression of recombinant proteins COS, CHO, 293, 293T cells). More particularly, a polynucleotide comprising the coding sequence of 24P4C12 or a fragment, analog or homolog thereof can be used to generate 24P4C12 proteins or fragments thereof using any number of host-vector systems routinely used and widely known in the art.

A wide range of host-vector systems suitable for the expression of 24P4C12 proteins or fragments thereof are available, see for example, Sambrook et al., 19B9, supra; Current Protocols in Molecular Biology, 1995, supra). Preferred vectors for mammalian expression include but are not limited to pcDNA 3.1 myc-His-tag (Invitrogen) and the retroviral vector pSRatkneo (Muller et 1991, MCB 11:1785). Using these expression vectors, 24P4C12 can be expressed in several prostate cancer and non-prostate cell lines, including for example 293, 293T, rat-1, NIH 3T3 and TsuPrl. The host-vector systems of the invention are useful for the production of a 24P4C12 protein or fragment thereof. Such host-vector systems can be employed to study the functional properties of 24P4C12 and 24P4C12 mutations or analogs.

Recombinant human 24P4C12 protein or an analog or homolog or fragment thereof can be produced by mammalian cells transfected with a construct encoding a 24P4C12-related nucleotide. For example, 293T cells can be transfected with an expression plasmid encoding 24P4C12 or fragment, analog or homolog thereof, a 24P4C12-related protein is expressed in the 293T cells, and the recombinant 24P4C12 protein is isolated using standard purification methods affinity purification using anti-24P4C12 antibodies). In another embodiment, a 24P4C12 coding sequence is subcloned into the retroviral vector pSRuMSVIkneo and used to infect various mammalian cell lines, such as NIH 3T3, TsuPrl, 293 and rat-1 in order to establish 24P4C12 expressing cell ines. Various other expression systems well known in the art can also be employed. Expression constructs encoding a leader peptide joined in frame to a 24P4C12 coding sequence can be used for the generation of a secreted form of recombinant 24P4C12 protein.

As discussed herein, redundancy in the genetic code permits variation in 24P4C12 gene sequences. In particular, it is known in the art that specific host species often have specific codon preferences, and thus one can adapt the disclosed sequence as preferred for a desired host. For example, preferred analog codon sequences typically have rare codons codons having a usage frequency of less than about 20% in known sequences of the desired host) replaced with higher frequency codons. Codon preferences for a specific species are calculated, for example, by utilizing codon usage tables available on the INTERNET such as at URL dna.affrc.go.jpl~nakamura/codon.html.

Additional sequence modifications are known to enhance protein expression in a cellular host. These include elimination of sequences encoding spurious polyadenylalion signals, exon/intron splice site signals, transposon-like repeats, and/or other such well-characterized sequences that are deleterious to gene expression. The GC content of the sequence is adjusted to levels average for a given cellular host, as calculated by reference to known genes expressed in the host cell.

Where possible, the sequence is modified to avoid predicted hairpin secondary mRNA structures. Other useful modifications include the addition of a translational initiation consensus sequence at the start of the open reading frame, as described in Kozak, Mol. Cell Bid., 9:5073-5080 (1989). Skilled artisans understand that the general rule that eukaryotic ribosomes initiate translation exclusively at the 5' proximal AUG codon is abrogated only under rare conditions (see, Kozak PNAS 92(7): 2662-2666, (1995) and Kozak NAR 15(20): 8125-8148(1987)).

Ill.) 24P4C12-related Proteins WO 2004/050828 PCT/US2002/038264 Another aspect of the present invention provides 24P4C12-related proteins. Specific embodiments of 24P4C12 proteins comprise a polypeptide having all or part of the amino acid sequence of human 24P4C12 as shown in Figure 2 or Figure 3. Alternatively, embodiments of 24P4C12 proteins comprise variant, homolog or analog polypeptides that have alterations in the amino acid sequence of 24P4C12 shown in Figure 2 or Figure 3.

Embodiments of a 24P4C12 polypeptide include: a 24P4C12 polypeptide having a sequence shown in Figure 2, a peptide sequence of a 24P4C12 as shown in Figure 2 wherein T is U; at least 10 contiguous nucleotides of a polypeptide having the sequence as shown in Figure 2; or, at least 10 contiguous peptides of a polypeptide having the sequence as shown in Figure 2 where T is U. For example, embodiments of 24P4C12 peptides comprise, without limitation: a protein comprising, consisting essentially of, or consisting of an amino acid sequence as shown in Figure 2A-I or Figure 3A-G; (II) a 24P4C12-related protein that is at least 90, 91, 92, 93, 94, 95, 96, 97,98, 99 or 100% homologous to an entire amino acid sequence shown in Figure 2A-I; (111) a 24P4C12-related protein that is at least 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 or 100% identical to an entire amino acid sequence shown in Figure 2A-I or 3A-G; (IV) a protein that comprises at least one peptide set forth in Tables VIII to XLIX, optionally with a proviso that it is not an entire protein of Figure 2; a protein that comprises at least one peptide set forth in Tables VIII-XXI, collectively, which peptide is also set forth in Tables XXII to XLIX, collectively, optionally with a proviso that it is not an entire protein of Figure 2; (VI) a protein that comprises at least two peptides selected from the peptides set forth in Tables VIII-XLIX, optionally with a proviso that it is not an entire protein of Figure 2; (VII) a protein that comprises at least two peptides selected from the peptides set forth in Tables VIII to XLIX collectively, with a proviso that the protein is not a contiguous sequence from an amino acid sequence of Figure 2; (VIII) a protein that comprises at least one peptide selected from the peptides set forth in Tables VIII-XXI; and at least one peptide selected from the peptides set forth in Tables XXII to XLIX, with a proviso that the protein is not a contiguous sequence from an amino acid sequence of Figure 2; (IX) a polypeptide comprising at least 5, 6, 7, 8, 9, 10, 11,12,13, 14,15,16, 17, 18,19, 20, 21, 22, 23, 24, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of Figure 3A, 3B, 3C, 30, 3E, 3F, or 3G in any whole number increment up to 710, 710, 710, 710, 598, 722, or 712 respectively that includes at least 1, 2, 3, 4, 6, 7,8, 9, 10, 11, 12, 13, 14, 15, 16,17,18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Hydrophilicity profile of Figure a polypeptide comprising at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acids of a protein of Figure 3A, 3B, 3C, 3D, 3E, 3F, or 3G in any whole number increment up to 710, 710, 710, 710, 598, 722, or 712 respectively, that includes at least 1, 2, 3, 4, 6, 7,8, 9, 10, 11, 12, 13, 14, 15,16,17,18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value less than 0.5 in the Hydropathicity profile of Figure 6; WO 2004/050828 PCT/US2002/038264 (XI) a polypeptide comprising at least 5, 6, 7, B, 9,10,11,12,13, 14,15,16,17, 18,19, 20,21,22, 23,24, 26, 27, 28, 29, 30, 31,32, 33, 34, 35 amino acids of a protein of Figure 3A, 3B, 3C, 3D, 3E, 3F, or 3G in any whole number increment up to 710, 710, 710, 710, 598, 722, or 712 respectively, that includes at least 1, 2, 3, 4, 6, 7, 8, 9,10,11, 12,13, 14,15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Percent Accessible Residues profile of Figure 7; (XII) a polypeptide comprising at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21,22, 23, 24, 26, 27, 28, 29, 30, 31,32, 33, 34, 35 amino acids of a protein of Figure 3A, 3B, 3C, 3D, 3E, 3F, or 3G in any whole number increment up to 710, 710, 710, 710, 598, 722, or 712 respectively, that includes at least 1, 2, 3, 4, 6, 7, 8, 9, 10, 11, 12,13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Average Flexibility profile of Figure 8; (XIII) a polypeptide comprising at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21,22, 23, 24, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino adds of a protein of Figure 3A, 3B, 3C, 3D, 3E, 3F, or 3G in any whole number increment up to 710, 710, 710, 710, 598, 722, or 712 respectively, that includes at least 1, 2, 3, 4, 6, 7,8, 9,10,11, 12,13, 14,15, 16, 17,18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 amino acid position(s) having a value greater than 0.5 in the Beta-turn profile of Figure 9; (XIV) a peptide that occurs at least twice in Tables VIII-XXI and XXII to XLIX, collectively; (XV) a peptide that occurs at least three times in Tables VIII-XXI and XXII to XLIX, collectively; (XVI) a peptide that occurs at least four times in Tables VIII-XXI and XXII to XLIX, collectively; (XVII) a peptide that occurs at least five times in Tables VIII-XXI and XXII to XLIX, collectively; (XVIII) a peptide that occurs at least once in Tables VIII-XXI, and at least once in tables XXII to XLIX; (XIX) a peptide that occurs at least once in Tables VIII-XXI, and at least twice in tables XXII to XLIX; (XX) a peptide that occurs at least twice in Tables VIII-XXI, and at least once in tables XXII to XLIX; (XXI) a peptide that occurs at least twice in Tables VIII-XXI, and at least twice in tables XXII to XLIX; (XXII) a peptide which comprises one two, three, four, or five of the following characteristics, or an oligonudeotide encoding such peptide: i) a region of at least 5 amino acids of a particular peptide of Figure 3, in any whole number increment up to the full length of that protein in Figure 3, that includes an amino acid position having a value equal to or greater than 0.5, 0.6, 0.7, 0.8, 0.9, or having a value equal to 1.0, in the Hydrophilicity profile of Figure ii) a region of at least 5 amino acids of a particular peptide of Figure 3, in any whole number increment up to the full length of that protein in Figure 3, that includes an amino add position having a value equal to or less than 0.5, 0.4, 0.3, 0.2, 0.1, or having a value equal to 0.0, in the Hydropathicity profile of Figure 6; iii) a region of at least 5 amino acids of a particular peptide of Figure 3, in any whole number increment up to the full length of that protein in Figure 3, that includes an amino acid position having a value equal to or greater than 0.5, 0.6, 0.7, 0.8, 0.9, or having a value equal to 1.0, in the Percent Accessible Residues profile of Figure 7; iv) a region of at least 5 amino acids of a particular peptide of Figure 3, in any whole number increment up to the full length of that protein in Figure 3, that includes an amino acid position having a value equal to or greater than 0.5, 0.6, 0.7, 0.8, 0.9, or having a value equal to 1.0, in the Average Flexibility profile of Figure 8; or, WO 2004/050828 PCT/US2002/038264 v) a region of at least 5 amino acids of a particular peptide of Figure 3, in any whole number increment up to the full length of that protein in Figure 3, that includes an amino acid position having a value equal to or greater than 0.5, 0.6, 0.7,0.8, 0.9, or having a value equal to 1.0, in the Beta-turn profile of Figure 9; (XXIII) a composition comprising a peptide of (I)-(XXII) or an antibody or binding region thereof together with a pharmaceutical excipient and/or in a human unit dose form.

(XXIV) a method of using a peptide of or an antibody or binding region thereof or a composition of (XXIII) in a method to modulate a cell expressing 24P4C12, (XXV) a method of using a peptide of (I)-(XXII) or an antibody or binding region thereof or a composition of (XXIII) in a method to diagnose, prophylax, prognose, or treat an individual who bears a cell expressing 24P4C12 (XXVI) a method of using a peptide of (I)-(XXII) or an antibody or binding region thereof or a composition (XXIII) in a method to diagnose, prophylax, prognose, or treat an individual who bears a cell expressing 24P4C12, said cell from a cancer of a tissue listed in Table I; (XXVII) a method of using a peptide of (I)-(XXII) or an antibody or binding region thereof or a composition of (XXIII) in a method to diagnose, prophylax, prognose, or treat a a cancer; (XXVIII) a method of using a peptide of (I)-(XXII) or an antibody or binding region thereof or a composition of (XXIII) in a method to diagnose, prophylax, prognose, or treat a a cancer of a tissue listed in Table I; and, (XXIX) a method of using a a peptide of (1)-(XXII) or an antibody or binding region thereof or a composition (XXIII) in a method to identify or characterie a modulator of a cell expressing 24P4C12.

As used herein, a range is understood to specifically disclose all whole unit positions thereof.

Typical embodiments of the invention disclosed herein include 24P4C12 polynucleotides that encode specific portions of 24P4C12 mRNA sequences (and those which are complementary to such sequences) such as those that encode the proteins and/or fragments thereof, for example: 4, 5, 6, 7, 8, 9, 10,11, 12, 13, 14, 15, 16, 17, 18,19, 20, 21, 22, 23, 24, 25, 30, 35, 40, 45, 50, 55, 60, 65, 80, 85, 90, 95, 100, 105, 110, 115, 120,125,130,135,140,145, 150, 155, 160,165,170,175,180,185, 190,195, 200, 225, 250, 275, 300, 325, 350, 375, 400, 425, 450, 475, 500, 525, 550, 575, 600, 625, 650, 675, 700, 710 or more contiguous amino acids of 24P4C12 variant 1; the maximal lengths relevant for other variants are: variant 3, 710 amino acids; variant 710 amino acids, variant 6, 710, variant 7, 598 amino acids, variant 8, 722 amino acids, and variant 9, 712 amino acids..

In general, naturally occurring allelic variants of human 24P4C12 share a high degree of structural identity and homology 90% or more homology). Typically, allelic variants of a 24P4C12 protein contain conservative amino acid substitutions within the 24P4C12 sequences described herein or contain a substitution of an amino acid from a corresponding position in a homologue of 24P4C12. One class of 24P4C12 allelic variants are proteins that share a high degree of homology with at least a small region of a particular 24P4C12 amino acid sequence, butfurther contain a radical departure from the sequence, such as a non-conservative substitution, truncation, insertion or frame shift. In comparisons of protein sequences, the terms, similarity, identity, and homology each have a distinct meaning as appreciated in the field of genetics. Moreover, orthology and paralogy can be important concepts describing the relationship of members of a given protein family in one organism to the members of the same family in other organisms.

WO 2004/050828 PCT/US2002/038264 Amino acid abbreviations are provided in Table II. Conservative amino acid substitutions can frequently be made in a protein without altering either the conformation or the function of the protein. Proteins of the invention can comprise 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,11, 12, 13, 14, 15 conservative substitutions. Such changes include substituting any of isoleucine valine and leucine for any other of these hydrophobic amino acids; aspartic acid for glutamic acid and vice versa; glutamine for asparagine and vice versa; and serine for threonine and vice versa. Other substitutions can also be considered conservative, depending on the environment of the particular amino acid and its role in the threedimensional structure of the protein. For example, glycne and alanine can frequently be interchangeable, as can alanine and valine Methionine which is relatively hydrophobic, can frequently be interchanged with leucine and isoleucine, and sometimes with valine. Lysine and arginine are frequently interchangeable in locations in which the significant feature of the amino acid residue is its charge and the differing pK's of these two amino acid residues are not significant. Still other changes can be considered "conservative" in particular environments (see, e.g. Table III herein; pages 13-15 "Biochemistry" 2 nd ED. Lubert Stryered (Stanford University); Henikoff et at, PNAS 1992 Vol 89 10915-10919; Lei et al., J Biol Chem 1995 May 19; 270(20):11882-6).

Embodiments of the invention disclosed herein include a wide variety of art-accepted variants or analogs of 24P4C12 proteins such as polypeptides having amino acid insertions, deletions and substitutions. 24P4C12 variants can be made using methods known in the art such as site-directed mutagenesis, alanine scanning, and PCR mutagenesis. Sitedirected mutagenesis (Carter et al., Nuc!. Acids Res., 13:4331 (1986); Zoller etal., Nucl. Acids Res., 10:6487 (1987)), cassette mutagenesis (Wells et Gene, 34:315 (1985)), restriction selection mutagenesis (Wells et al., Philos. Trans. R.

Soc. London SerA, 317:415 (1986)) or other known techniques can be performed on the cloned DNA to produce the 24P4C12 variant DNA.

Scanning amino acid analysis can also be employed to identify one or more amino acids along a contiguous sequence that is involved in a specific biological activity such as a protein-protein interaction. Among the preferred scanning amino acids are relatively small, neutral amino acds. Such amino acids include alanine, glycine, serine, and cysteine.

Alanine is typically a preferred scanning amino acid among this group because it eliminates the side-chain beyond the betacarbon and is less likely to alter the main-chain conformation of the variant. Alanine is also typically preferred because it is the most common amino acid. Further, it is frequently found in both buried and exposed positions (Creighton, The Proteins, Freeman Co., Chothia, J. Mol. Biol., 150:1 (1976)). If alanine substitution does not yield adequate amounts of variant, an isosteric amino acid can be used.

As defined herein, 24P4C12 variants, analogs or homologs, have the distinguishing attribute of having at least one epitope that is "cross reactive" with a 24P4C12 protein having an amino acid sequence of Figure 3. As used in this sentence, 'cross reactive' means that an antibody or T cell that specifically binds to a 24P4C12 variant also specifically binds to a 24P4C12 protein having an amino acid sequence set forth in Figure 3. A polypeptide ceases to be a variant of a protein shown in Figure 3, when it no longer contains any epitope capable of being recognized by an antibody or T cell that specifically binds to the starting 24P4C12 protein. Those skilled in the art understand that antibodies that recognize proteins bind to epitopes of varying size, and a grouping of the order of about four or five amino adds, contiguous or not, is regarded as a typical number of amino acids in a minimal epitope. See, Nair et al., J. Immunol 2000 165(12): 6949-6955; Hebbes eta/., Mol Immunol (1989) 26(9):865-73; Schwartz et al, J Immunol (1985) 135(4):2598-608.

Other classes of 24P4C12-related protein variants share 70%, 75%, 80%, 85% or 90% or more similarity with an amino acid sequence of Figure 3, or a fragment thereof. Another specific class of 24P4C12 protein variants or analogs comprises one or more of the 24P4C12 biological motifs described herein or presently known in the art. Thus, encompassed by the present invention are analogs of 24P4C12 fragments (nucleic or amino acid) that have altered functional (e.g.

WO 2004/050828 PCT/US2002/038264 immunogenic) properties relative to the starting fragment. Itis to be appreciated that motifs now or which become part of the art are to be applied to the nucleic or amino acid sequences of Figure 2 or Figure 3.

As discussed herein, embodiments of the claimed invention include polypeptides containing less than the full amino acid sequence of a 24P4C12 protein shown in Figure 2 or Figure 3. For example, representative embodiments of the invention comprise peptides/proteins having any 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,15 or more contiguous amino acids of a 24P4C12 protein shown in Figure 2 or Figure 3.

Moreover, representative embodiments of the invention disclosed herein include polypeptides consisting of about amino acid 1 to about amino acid 10 of a 24P4C12 protein shown in Figure 2 or Figure 3, polypeptides consisting of about amino acid 10 to about amino acid 20 of a 24P4C12 protein shown in Figure 2 or Figure 3, polypeptides consisting of about amino acid 20 to about amino acid 30 of a 24P4C12 protein shown in Figure 2 or Figure 3, polypeptides consisting of about amino acid 30 to about amino acid 40 of a 24P4C12 protein shown in Figure 2 or Figure 3, polypeptides consisting of about amino acid 40 to about amino acid 50 of a 24P4C12 protein shown in Figure 2 or Figure 3, polypeptides consisting of about amino acid 50 to about amino acid 60 of a 24P4C12 protein shown in Figure 2 or Figure 3, polypeptides consisting of about amino acid 60 to about amino acid 70 of a 24P4C12 protein shown in Figure 2 or Figure 3, polypeptides consisting of about amino acid 70 to about amino acid 80 of a 24P4C12 protein shown in Figure 2 or Figure 3, polypeptides consisting of about amino acid 80 to about amino acid 90 of a 24P4C12 protein shown in Figure 2 or Figure 3, polypeptides consisting of about amino acid 90 to about amino acid 100 of a 24P4C12 protein shown in Figure 2 or Figure 3, etc. throughout the entirety of a 24P4C12 amino acid sequence. Moreover, polypeptides consisting of about amino acid 1 (or 20 or 30 or 40 etc.) to about amino acid 20, (or 130, or 140 or 150 etc.) of a 24P4C12 protein shown in Figure 2 or Figure 3 are embodiments of the invention. It is to be appreciated that the starting and stopping positions in this paragraph refer to the specified position as well as that position plus or minus 5 residues.

24P4C12-related proteins are generated using standard peptide synthesis technology or using chemical cleavage methods well known in the art. Altematively, recombinant methods can be used to generate nucleic acid molecules that encode a 24P4C12-related protein. In one embodiment, nucleic acd molecules provide a means to generate defined fragments of a 24P4C12 protein (or variants, homologs or analogs thereof).

IIIA.) Motif-bearing Protein Embodiments Additional illustrative embodiments of the invention disclosed herein include 24P4C12 polypeptides comprising the amino acid residues of one or more of the biological motifs contained within a 24P4C12 polypeptide sequence set forth in Figure 2 or Figure 3. Various motifs are known in the art, and a protein can be evaluated for the presence of such motifs by a number of publicly available Internet sites (see, URL addresses: pfam.wustl.edul; searchlauncher.bcm.tmc.edulseqsearch/struc-predict.html; psort.ims.u-tokyo.acjpl; cbs.dtu.dk/; ebi.ac.uklinterpro/scan.html; expasy.ch/tools/scnpsitl.html; Epimatrix M and Epimer

T

Brown University, brown.edulResearchTB-H IV_Lab/epimatrixepimarix.html; and BIMAS, bimas.dcrt.nih.gov/.).

Motif bearing subsequences of all 24P4C12 variant proteins are set forth and identified in Tables VIII-XXI and XXII-

XLIX.

Table V sets forth several frequently occurring motifs based on pfam searches (see URL address pfam.wustl.edu/).

The columns of Table V list motif name abbreviation, percent identity found amongst the different member of the motif family, motif name or description and most common function; location information is included if the motif is relevant for location.

Polypeptides comprising one or more of the 24P4C12 motifs discussed above are useful in elucidating the specific characteristics of a malignant phenotype in view of the observation that the 24P4C12 motifs discussed above are associated with growth dysregulation and because 24P4C12 is overexpressed in certain cancers (See, Table Casein kinase II, WO 2004/050828 PCT/US2002/038264 cAMP and camp-dependent protein kinase, and Protein Kinase C, for example, are enzymes known to be associated with the development of the malignant phenotype (see e.g. Chen et Lab Invest, 78(2): 165-174 (1998); Gaiddon of al., Endocrinology 136(10): 4331-4338 (1995); Hall et at., Nucleic Acids Research 24(6): 1119-1126 (1996); Peterziel etal., Oncogene 18(46): 6322-6329 (1999) and OBrian, Oncol. Rep. 305-309 (1998)). Moreover, both glycosylation and myristoylation are protein modifications also associated with cancer and cancer progression (see e.g. Dennis et Biochem.

Biophys. Acta 1473(1):21-34 (1999); Raju et Exp. Cell Res. 235(1): 145-154 (1997)). Amidation is another protein modification also associated with cancer and cancer progression (see e.g. Treston et al., J. Nail. Cancer Inst. Monogr. (13): 169-175 (1992)).

In another embodiment, proteins of the invention comprise one or more of the immunoreactive epitopes identified in accordance with art-accepted methods, such as the peptides set forth in Tables Vll-XXI and XXII-XLIX. CTL epitopes can be determined using specific algorithms to identify peptides within a 24P4C12 protein that are capable of optimally binding to specified HLA alleles Table IV; Epimatrix T M and Epimer T M Brown University, URL brown,edu/Research/TB- HIVLab/epimatixepimatrix.html; and BIMAS, URL bimas.dcrtnih.gov/.) Moreover, processes for identifying peptides that have sufficient binding affinity for HLA molecules and which are correlated with being immunogenic epitopes, are well known in the art, and are carried out without undue experimentation. In addition, processes for identifying peptides that are immunogenic epitopes, are well known in the art, and are carried out without undue experimentation either in vitro or in vivo.

Also known in the art are principles for creating analogs of such epitopes in order to modulate immunogenicity. For example, one begins with an epitope that bears a CTL or HTL motif (see, the HLA Class I and HLA Class II molifslsupermotifs of Table IV). The epitope is analoged by substituting out an amino acid at one of the specified positions, and replacing it with another amino acid specified for that position. For example, on the basis of residues defined in Table IV, one can substitute out a deleterious residue in favor of any other residue, such as a preferred residue; substitute a lesspreferred residue with a preferred residue; or substitute an originally-occurring preferred residue with another preferred residue. Substitutions can occur at primary anchor positions or at other positions in a peptide; see, Table IV.

A variety of references reflect the art regarding the identification and generation of epitopes in a protein of interest as well as analogs thereof. See, for example, WO 97/33602 to Chesnut etal.; Sette, Immunogenetics 1999 50(3-4): 201- 212; Sette et J. Immunol. 2001 166(2): 1389-1397; Sidney et al., Hum. Immunol. 1997 58(1). 12-20; Kondo et al., Immunogenetics 1997 45(4): 249-258; Sidney et al., J. Immunol. 1996 157(8): 3480-90; and Falk et al, Nature 351: 290-6 (1991); Hunt etal., Science 255:1261-3 (1992); Parkeref al, J. Immunol. 149:3580-7 (1992); Parkeretal., J. Immunol.

152:163-75(1994)); Kast et al., 1994 152(8): 3904-12; Borras-Cuesta et al, Hum. Immunol. 2000 61(3): 266-278; Alexander et al., J. Immunol. 2000 164(3); 164(3): 1625-1633; Alexander etal., PMID: 7895164, UI: 95202582; O'Sullivan etat., J.

Immunol. 1991 147(8): 2663-2669; Alexander et al., Immunity 1994 751-761 and Alexander et al, Immunol. Res. 1998 18(2): 79-92.

Related embodiments of the invention indude polypeptides comprising combinations of the different motifs set forth in Table VI, and/or, one or more of the predicted CTL epitopes of Tables VllI-XXI and XXII-XLIX, and/or, one or more of the predicted HTL epitopes of Tables XLVI-XLIX, and/or, one or more of the T cell binding motifs known in the art Preferred embodiments contain no insertions, deletions or substitutions either within the motifs or within the intervening sequences of the polypeptides, In addition, embodiments which include a number of either N-terminal and/or C-terminal amino acid residues on either side of these motifs may be desirable (to, for example, include a greater portion of the polypeptide architecture in which the motif is located). Typically, the number of N-terminal and/or C-terminal amino acid residues on either side of a motif is between about 1 to about 100 amino acid residues, preferably 5 to about 50 amino acid residues.

24P4C12-related proteins are embodied in many forms, preferably in isolated form. A purified 24P4C12 protein molecule will be substantially free of other proteins or molecules that impair the binding of 24P4C12 to antibody, T cell or WO 2004/050828 PCT/US2002/038264 other ligand. The nature and degree of isolation and purification will depend on the intended use. Embodiments of a 24P4C12related proteins include purified 24P4C12-related proteins and functional, soluble 24P4C12-related proteins. In one embodiment, a functional, soluble 24P4C12 protein or fragment thereof retains the ability to be bound by antibody, T cell or other ligand.

The invention also provides 24P4C12 proteins comprising biologically active fragments of a 24P4C12 amino acid sequence shown in Figure 2 or Figure 3. Such proteins exhibit properties of the starting 24P4C12 protein, such as the ability to elicit the generation of antibodies that specifically bind an epitope associated with the starting 24P4C12 protein; to be bound by such antibodies; to elicit the activation of HTL or CTL; and/or, to be recognized by HTL or CTL that also specifically bind to the starting protein.

24P4C12-related polypeptides that contain particularly interesting structures can be predicted andlor identified using various analytical techniques well known in the art, including, for example, the methods of Chou-Fasman, Gamier-Robson, Kyle- Doolittle, Eisenberg, Karplus-Schultz or Jameson-Wolf analysis, or based on immunogenicity. Fragments that contain such structures are particularly useful in generating subunit-specific anti-24P4C12 antibodies or T cells or in identifying cellular factors that bind to 24P4C12. For example, hydrophilicity profiles can be generated, and immunogenic peptide fragments identified, using the method of Hopp, T.P. and Woods, 1981, Proc. Natl. Acad. Sci. U.S.A. 78:3824-3828. Hydropathicity profiles can be generated, and immunogenic peptide fragments identified, using the method of Kyte, J. and Doolittle, 1982, J.

Mol. Biol. 157:105-132. Percent(%) Accessible Residues profiles can be generated, and immunogenic peptide fragments identified, using the method of Janin 1979, Nature 277:491-492. Average Flexibility profiles can be generated, and immunogenic peptide fragments identified, using the method of Bhaskaran Ponnuswamy 1988, Int. J. Pept. Protein Res. 32:242-255. Beta-turn profiles can be generated, and immunogenic peptide fragments identified, using the method of Deleage, Roux 1987, Protein Engineering 1:289-294.

CTL epitopes can be determined using specific algorithms to identify peptides within a 24P4C12 protein that are capable of optimally binding to specified HLAalleles by using the SYFPEITHI site at World Wide Web URL syfpeithi.bmiheidelberg.coml; the listings in Table Epimatrix and Epimer T M Brown University, URL (brown.edulResearch/TB- HIVLablepimatrix/epimatrix.htm); and BIMAS, URL bimas.dat.nih.gov/). Illustrating this, peptide epitopes from 24P4C12 that are presented in the context of human MHC Class I molecules, HLA-A1, A2, A3, All, A24, B7 and 835 were predicted (see, Tables VIII-XXI, XXII-XLIX). Specifically, the complete amino acid sequence of the 24P4C12 protein and relevant portions of other variants, for HLA Class I predictions 9 flanking residues on either side of a point mutation or exon juction, and for HLA Class II predictions 14 flanking residues on either side of a point mutation or exon junction corresponding to that variant, were entered into the HLA Peptide Motif Search algorithm found in the Bioinformatics and Molecular Analysis Section (BIMAS) web site listed above; in addition to the site SYFPEITHI, at URL syfpeithi.bmiheidelberg.com/.

The HLA peptide motif search algorithm was developed by Dr. Ken Parker based on binding of specific peptide sequences in the groove of HLA Class I molecules, in particular HLA-A2 (see, Falk et al., Nature 351; 290-6 (1991); Hunt et Science 255:1261-3 (1992); Parker et J. Immunol. 149:3580-7 (1992); Parker et al., J. Immunol. 152:163-75 (1994)). This algorithm allows location and ranking of 8-mer, 9-mer, and 10-mer peptides from a complete protein sequence for predicted binding to HLA-A2 as well as numerous other HLA Class I molecules. Many HLAdass I binding peptides are 10 or 11-mers. For example, for Class I HLA-A2, the epitopes preferably contain a leucine or methionine at position 2 and a valine or leucine at the C-terminus (see, Parker etal., J. Immunol. 149:3580-7 (1992)).

Selected results of 24P4C12 predicted binding peptides are shown in Tables VIII-XXI and XXII-XLIX herein. In Tables VIll- XXI and XXII-XLVII, selected candidates, 9-mers and 10-mers, for each family member are shown along with their location, the amino acid sequence of each specific peptide, and an estimated binding score. In Tables XLVI-XLIX, selected WO 2004/050828 PCT/US2002/038264 candidates, 15-mers, for each family member are shown along with their location, the amino acid sequence of each specific peptide, and an estimated binding score. The binding score corresponds to the estimated half time of dissociation of complexes containing the peptide at 370C at pH 6.5. Peptides with the highest binding score are predicted to be the most tightly bound to HLA Class I on the cell surface for the greatest period of time and thus represent the best immunogenic targets for T-cell recognition.

Actual binding of peptides to an HLA allele can be evaluated by stabilization of HLA expression on the antigenprocessing defective cell line T2 (see, Xue et al, Prostate 30:73-8 (1997) and Peshwa et al., Prostate 36:129-38 (1998)). Immunogenicity of specific peptides can be evaluated in vitro by stimulation of CD8+ cytotoxic T lymphocytes (CTL) in the presence of antigen presenting cells such as dendritic cells.

It is to be appreciated that every epitope predicted by the BIMAS site, Epimer T M and Epimatrix

T

M sites, or specified by the HLA class I or class II motifs available in the art or which become part of the art such as set forth in Table IV (or determined using World Wide Web site URL syfpeithi.bmi-heidelberg.com/, or BIMAS, bimas.dcrt.nih.gov/) are to be "applied" to a 24P4C12 protein in accordance with the invention. As used in this context "applied" means that a 24P4C12 protein is evaluated, visually or by computer-based patterns finding methods, as appreciated by those of skill in the relevant art.

Every subsequence of a 24P4C12 protein of 8, 9, 10, or 11 amino acid residues that bears an HLA Class I motif, or a subsequence of 9 or more amino acid residues that bear an HLA Class II motif are within the scope of the invention 111.B.) Expression of 24P4C12-related Proteins In an embodiment described in the examples that follow, 24P4C12 can be conveniently expressed in cells (such as 293T cells) transfected with a commercially available expression vector such as a CMV-driven expression vector encoding 24P4C12 with a C-terminal 6XHis and MYC tag (pcDNA3.1/mycHIS, Invitrogen or Tag5, GenHunter Corporation, Nashville TN). The Tag5 vector provides an IgGK secretion signal that can be used to facilitate the production of a secreted 24P4C12 protein in transfected cells, The secreted HIS-tagged 24P4C12 in the culture media can be purified, using a nickel column using standard techniques.

III.C.) Modifications of 24P4C12-related Proteins Modifications of 24P4C12-related proteins such as covalent modifications are included within the scope of this invention. One type of covalent modification includes reacting targeted amino acid residues of a 24P4C12 polypeptide with an organic derivatizing agent that is capable of reacting with selected side chains or the N- or C- terminal residues of a 24P4C12 protein. Another type of covalent modification of a 24P4C12 polypeptide included within the scope of this invention comprises altering the native glycosylation pattern of a protein of the invention. Another type of covalent modification of 24P4C12 comprises linking a 24P4C12 polypeptide to one of a variety of nonproteinaceous polymers, polyethylene glycol (PEG), polypropylene glycol, or polyoxyalkylenes, in the manner set forth in U.S. Patent Nos. 4,640,835; 4,496,689; 4,3D1,144; 4,670,417; 4,791,192 or 4,179,337.

The 24P4C12-related proteins of the present invention can also be modified to form a chimeric molecule comprising 24P4C12 fused to another, heterologous polypeptide or amino acid sequence. Such a chimeric molecule can be synthesized chemically or recombinantly. A chimeric molecule can have a protein of the invention fused to another tumorassociated antigen or fragment thereof. Altematively, a protein in accordance with the invention can comprise a fusion of fragments of a 24P4C12 sequence (amino or nucleic acid) such that a molecule is created that is not, through its length, directly homologous to the amino or nucleic acid sequences shown in Figure 2 or Figure 3. Such a chimeric molecule can comprise multiples of the same subsequence of 24P4C12. A chimeric molecule can comprise a fusion of a 24P4C12-related protein with a polyhistidine epitope tag, which provides an epitope to which immobilized nickel can selectively bind, with WO 2004/050828 PCT/US2002/038264 cytokines or with growth factors. The epitope tag is generally placed at the amino- or carboxyl- terminus of a 24P4C12 protein. In an alternative embodiment, the chimeric molecule can comprise a fusion of a 24P4C12-related protein with an immunoglobulin or a particular region of an immunoglobulin. For a bivalent form of the chimeric molecule (also referred to as an "immunoadhesin"), such a fusion could be to the Fc region of an IgG molecule. The Ig fusions preferably include the substitution of a soluble (transmembrane domain deleted or inactivated) form of a 24P4C12 polypeptide in place of at least one variable region within an Ig molecule. In a preferred embodiment the immunoglobulin fusion includes the hinge, CH2 and CH3, or the hinge, CHI, CH2 and CH3 regions of an IgGI molecule. For the production of immunoglobulin fusions see, U.S. Patent No. 5,428,130 issued June 27,1995.

III.D.) Uses of 24P4C12-related Proteins The proteins of the invention have a number of different specific uses. As 24P4C12 is highly expressed in prostate and other cancers, 24P4C12-related proteins are used in methods that assess the status of 24P4C12 gene products in normal versus cancerous tissues, thereby elucidating the malignant phenotype. Typically, polypeptides from specific regions of a 24P4C12 protein are used to assess the presence of perturbations (such as deletions, insertions, point mutations etc.) in those regions (such as regions containing one or more motifs). Exemplary assays utilize antibodies or T cells targeting 24P4C12-related proteins comprising the amino acid residues of one or more of the biological motifs contained within a 24P4C12 polypeptide sequence in order to evaluate the characteristics of this region in normal versus cancerous tissues or to elicit an immune response to the epitope. Alternatively, 24P4C12-related proteins that contain the amino acid residues of one or more of the biological motifs in a 24P4C12 protein are used to screen for factors that interact with that region of 24P4C12.

24P4C12 protein fragmentstsubsequences are particularly useful in generating and characterizing domain-specific antibodies antibodies recognizing an extracellular orintracellular epitope of a 24P4C12 protein), for identifying agents or cellular factors that bind to 24P4C12 or a particular structural domain thereof, and in various therapeutic and diagnostic contexts, including but not limited to diagnostic assays, cancer vaccines and methods of preparing such vaccines.

Proteins encoded by the 24P4C12 genes, or by analogs, homologs or fragments thereof have a variety of uses, including but not limited to generating antibodies and in methods for identifying ligands and other agents and cellular constituents that bind to a 24P4C12 gene product Antibodies raised against a 24P4C12 protein or fragment thereof are useful in diagnostic and prognostic assays, and imaging methodologies in the management of human cancers characterized by expression of 24P4C12 protein, such as those listed in Table I. Such antibodies can be expressed intracellularly and used in methods of treating patients with such cancers. 24P4C12-related nucleic acids or proteins are also used in generating HTL or CTL responses.

Various immunological assays useful for the detection of 24P4C12 proteins are used, including but not limited to various types of radioimmunoassays, enzyme-linked immunosorbent assays (ELISA), enzyme-linked immunoluorescent assays (ELIFA), immunocytochemical methods, and the like. Antibodies can be labeled and used as immunological imaging reagents capable of detecting 24P4C12-expressing cells in radioscintigraphic imaging methods). 24P4C12 proteins are also particularly useful in generating cancer vaccines, as further described herein.

IV.) 24P4C12 Antibodies Another aspect of the invention provides antibodies that bind to 24P4C12-'elated proteins. Preferred antibodies specifically bind to a 24P4C12-related protein and do not bind (or bind weakly) to peptides or proteins that are not 24P4C12related proteins. For example, antibodies that bind 24P4C12 can bind 24P4C12-related proteins such as the homologs or analogs thereof.

WO 2004/050828 PCT/US2002/038264 24P4C12 antibodies of the invention are particularly useful in cancer (see, Table I) diagnostic and prognostic assays, and imaging methodologies. Similarly, such antibodies are useful in the treatment, diagnosis, andlor prognosis of other cancers, to the extent 24P4C12 is also expressed or overexpressed in these other cancers. Moreover, intracellularly expressed antibodies single chain antibodies) are therapeutically useful in treating cancers in which the expression of 24P4C12 is involved, such as advanced or metastatic prostate cancers.

The invention also provides various immunological assays useful for the detection and quantification of 24P4C12 and mutant 24P4C12-related proteins. Such assays can comprise one or more 24P4C12 antibodies capable of recognizing and binding a 24P4C12-related protein, as appropriate. These assays are performed within various immunological assay formats wel known in the art, including but not limited to various types of radioimmunoassays, enzyme-linked immunosorbent assays (ELISA), enzyme-linked immunofluorescent assays (ELIFA), and the like.

Immunological non-antibody assays of the invention also comprise T cell immunogenicity assays (inhibitory or stimulatory) as well as major histocompatibility complex (MHC) binding assays.

In addition, immunological imaging methods capable of detecting prostate cancer and other cancers expressing 24P4C12 are also provided by the invention, including but not limited to radioscintigraphic imaging methods using labeled 24P4C12 antibodies. Such assays are clinically useful in the detection, monitoring, and prognosis of 24P4C12 expressing cancers such as prostate cancer.

24P4C12 antibodies are also used in methods for purifying a 24P4C12-related protein and for isolating 24P4C12 homologues and related molecules. For example, a method of purifying a 24P4C12-related protein comprises incubating a 24P4C12 antibody, which has been coupled to a solid matrix, with a lysate or other solution containing a 24P4C12-related protein under conditions that permit the 24P4C12 antibody to bind to the 24P4C12-related protein; washing the solid matrix to eliminate impurities; and eluting the 24P4C12-related protein from the coupled antibody. Other uses of 24P4C12 antibodies in accordance with the invention include generating anti-idiotypic antibodies that mimic a 24P4C12 protein.

Various methods for the preparation of antibodies are well known in the art. For example, antibodies can be prepared by immunizing a suitable mammalian host using a 24P4C12-related protein, peptide, or fragment, in isolated or immunoconjugated form (Antibodies: A Laboratory Manual, CSH Press, Eds., Harlow, and Lane (1988); Harlow, Antibodies, Cold Spring Harbor Press, NY (1989)). In addition, fusion proteins of 24P4C12 can also be used, such as a 24P4C12 GST-fusion protein. In a particular embodiment, a GST fusion protein comprising all or most of the amino acid sequence of Figure 2 or Figure 3 is produced, then used as an immunogen to generate appropriate antibodies. In another embodiment, a 24P4C12-related protein is synthesized and used as an immunogen.

In addition, naked DNA immunization techniques known in the art are used (with or without purified 24P4C12-related protein or 24P4C12 expressing cells) to generate an immune response to the encoded immunogen (for review, see Donnelly et al., 1997, Ann. Rev. Immunol. 15: 617-648).

The amino acid sequence of a 24P4C12 protein as shown in Figure 2 or Figure 3 can be analyzed to select specific regions of the 24P4C12 protein for generating antibodies. For example, hydrophobicity and hydrophilicity analyses of a 24P4C12 amino acd sequence are used to identify hydrophilic regions in the 24P4C12 structure. Regions of a 24P4C12 protein that show immunogenic structure, as well as other regions and domains, can readily be identified using various other methods known in the art, such as Chou-Fasman, Gamier-Robson, Kyte-Doollttle, Elsenberg, Karplus-Schultz or Jameson-Wolf analysis. Hydrophilicity profiles can be generated using the method of Hopp, T.P. and Woods, 1981, Proc. Natl. Acad. Sci. U.S.A. 78:3824- 3828. Hydropathicity profiles can be generated using the method of Kyte, J. and Doolittle, RF., 1982, J. Mol. Biol. 157:105- 132. Percent Accessible Residues profiles can be generated using the method of Janin 1979, Nature 277:491-492.

Average Flexibility profiles can be generated using the method of Bhaskaran Ponnuswamy 1988, Int. J. Pept.

Protein Res. 32:242-255. Beta-turn profiles can be generated using the method of Deleage, Roux 1987, Protein WO 2004/050828 PCT/US2002/038264 Engineering 1:289-294. Thus, each region identified by anyof these programs or methods is within the scope of the present invention. Methods for the generation of 24P4C12 antibodies are further illustrated by way of the examples provided herein.

Methods for preparing a protein or polypeptide for use as an immunogen are well known in the art. Also well known in the art are methods for preparing immunogenic conjugates of a protein with a carrier, such as BSA, KLH or other carrier protein. In some circumstances, direct conjugation using, for example, carbodiimide reagents are used; in other instances linking reagents such as those supplied by Pierce Chemical Co., Rockford, IL, are effective. Administration of a 24P4C12 immunogen is often conducted by injection over a suitable time period and with use of a suitable adjuvant, as is understood in the art. During the immunization schedule, titers of antibodies can be taken to determine adequacy of antibody formation.

24P4C12 monoclonal antibodies can be produced by various means well known in the art For example, immortalized cell lines that secrete a desired monoclonal antibody are prepared using the standard hybridoma technology of Kohler and Milstein or modifications that immortalize antibody-producing B cells, as is generally known. Immortalized cell lines that secrete the desired antibodies are screened by immunoassay in which the antigen is a 24P4C12-related protein. When the appropriate immortalized cell culture is identified, the cells can be expanded and antibodies produced either from in vitro cultures or from ascites fluid.

The antibodies or fragments of the invention can also be produced, by recombinant means. Regions that bind specifically to the desired regions of a 24P4C12 protein can also be produced in the context of chimeric or complementaritydetermining region (CDR) grafted antibodies of multiple species origin. Humanized or human 24P4C12 antibodies can also be produced, and are preferred for use in therapeutic contexts. Methods for humanizing murine and other non-human antibodies, by substituting one or more of the non-human antibody CDRs for corresponding human antibody sequences, are well known (see for example, Jones et al., 1986, Nature 321: 522-525; Riechmann et al, 1988, Nature 332: 323-327; Verhoeyen et al., 1988, Science 239: 1534-1536). See also, Carter et al, 1993, Proc. Natl. Acad. Sci. USA 89:4285 and Sims et al., 1993, J. Immunol. 151: 2296.

Methods for producing fully human monoclonal antibodies include phage display and transgenic methods (for review, see Vaughan etal., 1998, Nature Biotechnology 16: 535-539). Fully human 24P4C12 monoclonal antibodiescan be generated using cloning technologies employing large human Ig gene combinatorial libraries phage display) (Griffiths and Hoogenboom, Building an in vtro immune system: human antibodies from phage display libraries. In: Protein Engineering of Antibody Molecules for Prophylactic and Therapeutic Applications in Man, Clark, M. Nottingham Academic, pp 45-64 (1993); Burton and Barbas, Human Antibodies from combinatorial libraries. Id., pp 65-82). Fully human 24P4C12 monoclonal antibodies can also be produced using transgenic mice engineered to contain human immunoglobulin gene loci as described in PCT Patent Application W098/24893, Kucherlapati and Jakobovits et al., published December 3, 1997 (see also, Jakobovits, 1998, Exp. Opin. Invest Drugs 607-614; U.S. patents 6,162,963 issued 19 December 2000; 6,150,584 issued 12 November 2000; and, 6,114598 issued 5 September 2000). This method avoids the in vitro manipulation required with phage display technology and efficiently produces high affinity authentic human antibodies.

Reactivity of 24P4C12 antibodies with a 24P4C12-related protein can be established by a number of well known means, including Western blot, immunoprecipitation, ELISA, and FACS analyses using, as appropriate, 24P4C12-related proteins, 24P4C12-expressing cells or extracts thereof. A 24P4C12 antibody or fragment thereof can be labeled with a detectable marker or conjugated to a second molecule. Suitable detectable markers include, but are not limited to, a radioisotope, a fluorescent compound, a bioluminescent compound, chemiluminescent compound, a metal chelator or an enzyme. Further, bi-specific antibodies specific for two or more 24P4C12 epitopes are generated using methods generally known in the art. Homodimeric antibodies can also be generated by cross-linking techniques known in the art Wolff et al., Cancer Res. 53: 2560-2565).

24P4C12 Cellular Immune Responses WO 2004/050828 PCT/US2002/038264 The mechanism by which T cells recognize antigens has been delineated. Efficacious peptide epitope vaccine compositions of the invention induce a therapeutic or prophylactic immune responses in very broad segments of the worldwide population. For an understanding of the value and efficacy of compositions of the invention that induce cellular immune responses, a brief review of immunology-related technology is provided.

A complex of an HLA molecule and a peptidic antigen acts as the ligand recognized by HLA-restricted T cells (Buus, S. et al., Cell 47:1071, 1986; Babbitt, B. P. et Nature 317:359, 1985; Townsend, A. and Bodmer, Annu. Rev.

Immunol. 7:601, 1989; Germain, R. Annu. Rev. Immunol. 11:403,1993). Through the study of single amino acid substituted antigen analogs and the sequencing of endogenously bound, naturally processed peptides, critical residues that correspond to motifs required for specific binding to HLA antigen molecules have been identified and are set forth in Table IV (see also, Southwood, et al., J. Immuno. 160:3363, 1998; Rammensee, et al., Immunogenetics 41:178, 1995; Rammensee e al., SYFPEITHI, access via World Wide Web at URL (134.2.96.221/scripts.hlaserver.dlllhome.htm); Sette, A.

and Sidney, J. Curr. Opin. Immunol. 10:478, 1998; Engelhard, V. Curr. Opin. Immunol. 6:13,1994; Sette, A. and Grey, H.

Curr. Opin. Immuno. 4:79,1992; Sinigaglia, F. and Hammer, J. Curr. Biol. 6:52, 1994; Ruppert et Cell 74:929-937, 1993; Kondo et al., J. Immunol. 155:4307-4312,1995; Sidney et al., J. Immunol. 157:3480-3490,1996; Sidney et al, Human Immunol. 45:79-93, 1996; Sette, A. and Sidney, J. Immunogenetics 1999 Nov; 50(3-4):201-12, Review).

Furthermore, x-ray crystallographic analyses of HLA-peptide complexes have revealed pockets within the peptide binding cleftigroove of HLA molecules which accommodate, in an allele-specific mode, residues bore by peptide ligands; these residues in turn determine the HLA binding capacity of the peptides in which they are present. (See, Madden, D.R. Annu. Rev. Immunol. 13:587, 1995; Smith, et al., Immunity 4:203, 1996; Fremont et al, Immunity 8:305, 1998; Stern et al., Structure 2:245, 1994; Jones, E.Y. Curr. Opin. Immuncl. 9:75, 1997; Brown, J. H. etal., Nature 364:33,1993; Guo, H. C.

et al., Proc. Natl. Acad. Sci. USA 90:8053, 1993; Guo, H. C. et al., Nature 360:364, 1992; Silver, M. L. et at, Nature 360:367, 1992; Matsumura, M. et al, Science 257:927, 1992; Madden ef al., Cel 70:1035, 1992; Fremont, D. H. et Science 257:919, 1992; Saper, M. A. Bjorkman, P. J. and Wiley, D. J. Mot. Biol. 219:277, 1991.) Accordingly, the definition of class I and class II allele-specific HLA binding motifs, or class I or class II supermotifs allows identification of regions within a protein that are correlated with binding to particular HLA antigen(s).

Thus, by a process of HLA motif identification, candidates for epitope-based vaccines have been identified; such candidates can be further evaluated by HLA-peptide binding assays to determine binding affinity andlor the time period of association of the epitope and its corresponding HLA molecule. Additional confirmatory work can be performed to select, amongst these vaccine candidates, epitopes with preferred characteristics in terms of population coverage, and/or immunogenicity.

Various strategies can be utilized to evaluate cellular immunogenicity, including: 1) Evaluation of primary T cell cultures from normal individuals (see, Wentworth, P. A. et al., Mot. !rnmunot.

32:603, 1995; Cells, E. etal., Proc. Natl. Acad. Sci. USA 91:2105,1994; Tsai, V. et J. Immunol. 158:1796, 1997; Kawashima, I. et al., Human Immunol. 59:1, 1998). This procedure involves the stimulation of peripheral blood lymphocytes (PBL) from normal subjects with a test peptide in the presence of antigen presenting cells in vitro over a period of several weeks. T cells specific for the peptide become activated during this time and are detected using, a lymphokine- or 1 Cr-release assay involving peptide sensitized target cells.

2) Immunization of HLA transgenic mice (see, Wentworth, P. A. et al, J. Immunol. 26:97,1996; Wentworth, P.

A. et Int. Immune. 8:651, 1996; Alexander, J. et al., J. Immunol. 159:4753, 1997). For example, in such methods peptides in incomplete Freund's adjuvant are administered subcutaneously to HLA transgenic mice. Several weeks following immunization, splenocytes are removed and cultured in vitro in the presence of test peptide for approximately one week.

WO 2004/050828 PCT/US2002/038264 Peptide-specific T cells are detected using, a 51 Cr-release assay involving peptide sensitized target cells and target cells expressing endogenously generated antigen.

3) Demonstration of recall T cell responses from immune individuals who have been either effectively vaccinated and/or from chronically ill patients (see, Rehermann, B. et al., J. Exp. Med. 181:1047,1995; Doolan, D. L. etal., Immunity7:97, 1997; Bertoni, R. et al., J. Clin. Invest. 100:503,1997; Threlkeld, S. C. et al., J. Immunol. 159:1648, 1997; Diepolder, H. M. et al., J. Virol 71:6011, 1997). Accordingly, recall responses are detected by culturing PBL from subjects that have been exposed to the antigen due to disease and thus have generated an immune response "naturally", or from patients who were vaccinated against the antigen. PBL from subjects are cultured in vitro for 1-2 weeks in the presence of test peptide plus antigen presenting cells (APC) to allow activation of "memory" T cells, as compared to naive" T cells. At the end of the culture period, T cell activity is detected using assays including 51 Cr release involving peptide-sensitized targets, T cell proliferation, or lymphokine release.

Vl.) 24P4C12 Transgenic Animals Nucleic acids that encode a 24P4C12-related protein can also be used to generate either transgenic animals or "knock out" animals that, in turn, are useful in the development and screening of therapeutically useful reagents. In accordance with established techniques, cDNA encoding 24P4C12 can be used to clone genomic DNA that encodes 24P4C12. The cloned genomic sequences can then be used to generate transgenic animals containing cells that express DNA that encode 24P4C12. Methods for generating transgenic animals, particularly animals such as mice or rats, have become conventional in the art and are described, for example, in U.S. Patent Nos. 4,736,866 issued 12 April 1988, and 4,870,009 issued 26 September 1989. Typically, particular cells would be targeted for 24P4C12 transgene incorporation with tissue-specific enhancers.

Transgenic animals that include a copy of a transgene encoding 24P4C12 can be used to examine the effect of increased expression of DNA that encodes 24P4C12. Such animals can be used as tester animals for reagents thought to confer protection from, for example, pathological conditions associated with its overexpression. In accordance with this aspect of the invention, an animal is treated with a reagent and a reduced incidence of a pathological condition, compared to untreated animals that bear the transgene, would indicate a potential therapeutic intervention for the pathological condition.

Alternatively, non-human homologues of 24P4C12 can be used to construct a 24P4C12 "knock out" animal that has a defective or altered gene encoding 24P4C12 as a result of homologous recombination between the endogenous gene encoding 24P4C12 and altered genomic DNA encoding 24P4C12 introduced into an embryonic cell of the animal. For example,.cDNA that encodes 24P4C12 can be used to clone genomic DNA encoding 24P4C12 in accordance with established techniques. A portion of the genomic DNA encoding 24P4C12 can be deleted or replaced with another gene, such as a gene encoding a selectable marker that can be used to monitor integration. Typically, several kilobases of unaltered flanking DNA (both at the 5 and 3' ends) are included in the vector (see, Thomas and Capecchi, Cell, 51:503 (1987) for a description of homologous recombination vectors). The vector is introduced into an embryonic stem cell line by electroporation) and cells in which the introduced DNA has homologously recombined with the endogenous DNA are selected (see, U et al., Cell, 69:915 (1992)). The selected cells are then injected into a blastocyst of an animal a mouse or rat) to form aggregation chimeras (see, Bradley, in Teratocarcinomas and Emblyonic Stem Cells: A Practical Approach, E. J. Robertson, ed. (IRL, Oxford, 1987), pp. 113-152). A chimeric embryo can then be implanted into a suitable pseudopregnant female foster animal, and the embryo brought to term to create a "knock out animal. Progeny harboring the homologously recombined DNA in their germ cells can be identified by standard techniques and used to breed animals in which all cells of the animal contain the homologously recombined DNA. Knock out animals can be characterized, WO 2004/050828 PCT/US2002/038264 for example, for their ability to defend against certain pathological conditions or for their development of pathological conditions due to absence of a 24P4C12 polypeptide.

VII.) Methods for the Detection of 24P4C12 Another aspect of the present invention relates to methods for detecting 24P4C12 polynudeotides and 24P4C12related proteins, as well as methods for identifying a cell that expresses 24P4C12. The expression profile of 24P4C12 makes it a diagnostic marker for metastasized disease. Accordingly, the status of 24P4C12 gene products provides information useful for predicting a variety of factors including susceptibility to advanced stage disease, rate of progression, and/or tumor aggressiveness. As discussed in detail herein, the status of 24P4C12 gene products in patient samples can be analyzed by a variety protocols that are well known in the art including immunohistochemical analysis, the variety of Northern blotting techniques including in situ hybridization, RT-PCR analysis (for example on laser capture micro-dissected samples), Western blot analysis and tissue array analysis.

More particularly, the invention provides assays for the detection of 24P4C12 polynicleotides in a biological sample, such as serum, bone, prostate, and other tissues, urine, semen, cell preparations, and the like. Detectable 24P4C12 polynudeotides include, for example, a 24P4C12 gene or fragment thereof, 24P4C12 mRNA, alternative splice variant 24P4C12 mRNAs, and recombinant DNA or RNA molecules that contain a 24P4C12 polynucleotide. A number of methods for amplifying and/or detecting the presence of 24P4C12 polynuceotides are well known in the art and can be employed in the practice of this aspect of the invention.

In one embodiment, a method for detecting a 24P4C12 mRNA in a biological sample comprises producing cDNA from the sample by reverse transcription using at least one primer; amplifying the cDNA so produced using a 24P4C12 polynucleotides as sense and antisense primers to amplify 24P4C12 cDNAs therein; and detecting the presence of the amplified 24P4C12 cDNA. Optionally, the sequence of the amplified 24P4C12 cDNA can be determined.

In another embodiment, a method of detecting a 24P4C12 gene in a biological sample comprises first isolating genomic DNA from the sample; amplifying the isolated genomic DNA using 24P4C12 polynudeotides as sense and antisense primers; and detecting the presence of the amplified 24P4C12 gene. Any number of appropriate sense and antisense probe combinations can be designed from a 24P4C12 nucleotide sequence (see, Figure 2) and used for this purpose.

The invention also provides assays for detecting the presence of a 24P4C12 protein in a tissue or other biological sample such as serum, semen, bone, prostate, urine, cel preparations, and the like. Methods for detecting a 24P4C12-related protein are also well known and include, for example, immunoprecipitation, immunohistochemical analysis, Western blot analysis, molecular binding assays, ELISA, ELIFA and the like. For example, a method of detecting the presence of a 24P4C12-related protein in a biological sample comprises first contacting the sample with a 24P4C12 antibody, a 24P4C12-reactive fragment thereof, or a recombinant protein containing an antigen-binding region of a 24P4C12 antibody; and then detecting the binding of 24P4C12-related protein in the sample.

Methods for identifying a cell that expresses 24P4C12 are also within the scope of the invention. In one embodiment, an assay for identifying a cell that expresses a 24P4C12 gene comprises detecting the presence of 24P4C12 mRNA in the cell.

Methods for the detection of particular mRNAs in cells are well known and include, for example, hybridization assays using complementary DNA probes (such as in situ hybridization using labeled 24P4C12 riboprobes, Northern blot and related techniques) and various nucleic acid amplification assays (such as RT-PCR using complementary primers specific for 24P4C12, and other amplification type detection methods, such as, for example, branched DNA, SISBA, TMA and the like). Alternatively, an assay for identifying a cell that expresses a 24P4C12 gene comprises detecting the presence of 24P4C12-related protein in the WO 2004/050828 PCT/US2002/038264 cell or secreted by the cell. Various methods for the detection of proteins are well known in the art and are employed for the detection of 24P4C12-related proteins and cells that express 24P4C12-related proteins.

24P4C12 expression analysis is also useful as a tool for identifying and evaluating agents that modulate 24P4C12 gene expression. For example, 24P4C12 expression is significantly upregulated in prostate cancer, and is expressed in cancers of the tissues listed in Table I. Identification of a molecule or biological agent that inhibits 24P4C12 expression or overexpression in cancer cells is of therapeutic value. For example, such an agent can be identified by using a screen that quantifies 24P4C12 expression by RT-PCR, nucleic acid hybridization or antibody binding.

VIII. Methods for Monitoring the Status of 24P4C12-related Genes and Their Products Oncogenesis is known to be a multistep process where cellular growth becomes progressively dysregulated and cells progress from a normal physiological state to precancerous and then cancerous states (see, Alers et at., Lab Invest..77(5): 437-438(1997) and Isaacs etal, Cancer Surv. 23: 19-32 (1995)). In this context, examining a biological sample for evidence of dysregulated cell growth (such as aberrant 24P4C12 expression in cancers) allows for early detection of such aberrant physiology, before a pathologic state such as cancer has progressed to a stage that therapeutic options are more limited and or the prognosis is worse. In such examinations, the status of 24P4C12 in a biological sample of interest can be compared, for example, to the status of 24P4C12 in a corresponding normal sample a sample from that individual or alternatively another individual that is not affected by a pathology). An alteration in the status of 24P4C12 in the biological sample (as compared to the normal sample) provides evidence of dysregulated cellular growth. In addition to using a biological sample that is not affected by a pathology as a normal sample, one can also use a predetermined normative value such as a predetermined normal level of mRNA expression (see, Grever eta/., J. Comp. Neurol. 1996 Dec 9; 376(2): 306-14 and U.S. Patent No. 5,837,501) to compare 24P4C12 status in a sample.

The term "status" in this context is used according to its art accepted meaning and refers to the condition or state of a gene and its products. Typically, skilled artisans use a number of parameters to evaluate the condition or state of a gene and its products. These include, but are not limited to the location of expressed gene products (including the location of 24P4C12 expressing cells) as well as the level, and biological activity of expressed gene products (such as 24P4C12 mRNA, polynucleotides and polypeptides). Typically, an alteration in the status of 24P4C12 comprises a change in the location of 24P4C12 and/or 24P4C12 expressing cells and/or an increase in 24P4C12 mRNA and/or protein expression.

24P4C12 status in a sample can be analyzed by a number of means well known in the art, including without limitation, immunohistochemical analysis, in situ hybridization, RT-PCR analysis on laser capture micro-dissected samples, Western blot analysis, and tissue array analysis. Typical protocols for evaluating the status of a 24P4C12 gene and gene products are found, for example in Ausubel et al. eds., 1995, Current Protocols In Molecular Biology, Units 2 (Northem Blotting), 4 (Southern Blotting), 15 (Immunoblotting) and 18 (PCR Analysis). Thus, the status of 24P4C12 in a biological sample is evaluated by various methods utilized by skilled artisans including, but not limited to genomic Southern analysis (to examine, for example perturbations in a 24P4C12 gene), Northern analysis and/or PCR analysis of 24P4C12 mRNA (to examine, for example alterations in the polynucleotide sequences or expression levels of 24P4C12 mRNAs), and, Western and/or immunohistochemical analysis (to examine, for example alterations in polypeptide sequences, alterations in polypeptide localization within a sample, alterations in expression levels of 24P4C12 proteins and/or associations of 24P4C12 proteins with polypeptide binding partners). Detectable 24P4C12 polynucleotides include, for example, a 24P4C12 gene or fragment thereof, 24P4C12 mRNA, alternative splice variants, 24P4C12 mRNAs, and recombinant DNA or RNA molecules containing a 24P4C12 polynudeotide.

The expression profile of 24P4C12 makes it a diagnostic marker for local and/or metastasized disease, and provides information on the growth or oncogenic potential of a biological sample. In particular, the status of 24P4C12 provides WO 2004/050828 PCT/US2002/038264 information useful for predicting susceptibility to particular disease stages, progression, and/or tumor aggressiveness. The invention provides methods and assays for determining 24P4C12 status and diagnosing cancers that express 24P4C12, such as cancers of the tissues listed in Table I. For example, because 24P4C12 mRNA is so highly expressed in prostate and other cancers relative to normal prostate tissue, assays that evaluate the levels of 24P4C12 mRNA transcripts or proteins in a biological sample can be used to diagnose a disease associated with 24P4C12 dysregulation, and can provide prognostic information useful in defining appropriate therapeutic options.

The expression status of 24P4C12 provides information including the presence, stage and location of dysplastic, precancerous and cancerous cells, predicting susceptibility to various stages of disease, and/or for gauging tumor aggressiveness. Moreover, the expression profile makes it useful as an imaging reagent for metastasized disease.

Consequently, an aspect of the invention is directed to the various molecular prognostic and diagnostic methods for examining the status of 24P4C12 in biological samples such as those from individuals suffering from, or suspected of suffering from a pathology characterized by dysregulated cellular growth, such as cancer.

As described above, the status of 24P4C12 in a biological sample can be examined by a number of well-known procedures in the art. For example, the status of 24P4C12 in a biological sample taken from a specific location in the body can be examined by evaluating the sample for the presence or absence of 24P4C12 expressing cells those that express 24P4C12 mRNAs or proteins). This examination can provide evidence of dysregulated cellular growth, for example, when 24P4C12-expressing cells are found in a biological sample that does not normally contain such cells (such as a lymph node), because such alterations in the status of 24P4C12 in a biological sample are often associated with dysregulated cellular growth. Specifically, one indicator of dysregulated cellular growth is the metastases of cancer cells from an organ of origin.

(such as the prostate) to a different area of the body (such as a lymph node). In this context, evidence of dysregulated cellular growth is important for example because occult lymph node metastases can be detected in a substantial proportion of patients with prostate cancer, and such metastases are associated with known predictors of disease progression (see, Murphy et Prostate 42(4): 315-317 (2000);Su et Semin. Surg. Oncol. 18(1): 17-28 (2000) and Freeman et J Urol 1995 Aug 154(2 Pt 1):474-8).

In one aspect, the invention provides methods for monitoring 24P4C12 gene products by determining the status of 24P4C12 gene products expressed by cells from an individual suspected of having a disease associated with dysregulated cell growth (such as hyperplasia or cancer) and then comparing the status so determined to the status of 24P4C12 gene products in a corresponding normal sample. The presence of aberrant 24P4C12 gene products in the test sample relative to the normal sample provides an indication of the presence of dysregulated cell growth within the cells of the individual.

In another aspect, the invention provides assays useful in determining the presence of cancer in an individual, comprising detecting a significant increase in 24P4C12 mRNA or protein expression in a test cell or tissue sample relative to expression levels in the corresponding normal cell or tissue. The presence of 24P4C12 mRNA can, for example, be evaluated in tissues including but not limited to those listed in Table I. The presence of significant 24P4C12 expression in any of these tissues is useful to indicate the emergence, presence and/or severity of a cancer, since the corresponding normal tissues do not express 24P4C12 mRNA or express it at lower levels.

In a related embodiment, 24P4C12 status is determined at the protein level rather than at the nucleic acid level. For example, such a method comprises determining the level of 24P4C12 protein expressed by cells in atest tissue sample and comparing the level so determined to the level of 24P4C12 expressed in a corresponding normal sample. In one embodiment, the presence of 24P4C12 protein is evaluated, for example, using immunohistochemical methods. 24P4C12 antibodies or binding partners capable of detecting 24P4C12 protein expression are used in a variety of assay formats well known in the art for this purpose.

WO 2004/050828 PCT/US2002/038264 In a further embodiment, one can evaluate the status of 24P4C12 nucleotide and amino add sequences in a biological sample in order to identify perturbations in the structure of these molecules. These perturbations can include insertions, deletions, substitutions and the like. Such evaluations are useful because perturbations in the nucleotide and amino acid sequences are observed in a large number of proteins associated with a growth dysregulated phenotype (see, Marrogi et 1999, J.

Cutan. Pathol. 26(8):369-378). For example, a mutation in the sequence of 24P4C12 may be indicative of the presence or promotion of a tumor. Such assays therefore have diagnostic and predictive value where a mutation in 24P4C12 indicates a potential loss of function or increase in tumor growth.

A wide variety of assays for observing perturbations in nucleotide and amino acid sequences are well known in the art.

For example, the size and structure of nucleic acid or amino acid sequences of 24P4C12 gene products are observed by the Northem, Southem, Westem, PCR and DNA sequencing protocols discussed herein. In addition, other methods for observing perturbations in nucleotide and amino acid sequences such as single strand conformation polymorphism analysis are well known in the art (see, U.S. Patent Nos. 5,382,510 issued 7 September 1999, and 5,952,170 issued 17 January 1995).

Additionally, one can examine the methylation status of a 24P4C12 gene in a biological sample. Aberrant demethylaton and/or hypermethylation of CpG islands in gene 5' regulatory regions frequently occurs in immortalized and transformed cells, and can result in altered expression of various genes. For example, promoter hypermethylation of the pi-class glutathione S-transferase (a protein expressed in normal prostate but not expressed in >90% of prostate carcinomas) appears to permanently silence transcription of this gene and is the most frequently detected genomic alteration in prostate carcinomas (De Marzo e al., Am. J. Pathol. 155(6): 1985-1992 (1999)). In addition, this alteration is present in at least of cases of high-grade prostatic intraepithelial neoplasia (PIN) (Brooks et Cancer Epidemiol. Biomarkers Prev., 1998, 7:531-536). In another example, expression of the LAGE-I tumor specific gene (which is not expressed in normal prostate but is expressed in 25-50% of prostate cancers) is induced by deoxy-azacytidine in lymphoblastoid cells, suggesting that tumoral expression is due to demethylation (Lethe et al., Int. J. Cancer 76(6): 903-908 (1998)). A variety of assays for examining methylation status of a gene are well known in the art. For example, one can utilize, in Southem hybridization approaches, methylation-sensitive restriction enzymes that cannot deave sequences that contain methylated CpG sites to assess the methylaton status of CpG islands. In addition, MSP (methylation specific PCR) can rapidly profile the methylation status of all the CpG sites present in a CpG island of a given gene. This procedure involves initial modification of DNA by sodium bisulfite (which will convert all unmethylated cytosines to uracil) followed by amplification using primers specific for methylated versus unmethylated DNA. Protocols involving methylation interference can also be found for example in Current Protocols In Molecular Biology, Unit 12, Frederick M. Ausubel etal. eds., 1995.

Gene amplification is an additional method for assessing the status of 24P4C12. Gene amplification Is measured in a sample directly, for example, by conventional Southern blotting or Northern blotting to quantitate the transcription of mRNA (Thomas, 1980, Proc. Natl. Acad. Sci. USA, 77:5201-5205), dot blotting (DNA analysis), or in situ hybridization, using an appropriately labeled probe, based on the sequences provided herein. Alternatively, antibodies are employed that recognize specific duplexes, including DNA duplexes, RNA duplexes, and DNA-RNA hybrid duplexes or DNA-protein duplexes. The antibodies in turn are labeled and the assay carried out where the duplex is bound to a surface, so that upon the formation of duplex on the surface, the presence of antibody bound to the duplex can be detected.

Biopsied tissue or peripheral blood can be conveniently assayed for the presence of cancer cells using for example, Northem, dot blot or RT-PCR analysis to detect 24P4C12 expression. The presence of RT-PCR amplifiable 24P4C12 mRNA provides an indication of the presence of cancer, RT-PCR assays are well known in the art RT-PCR detection assays for tumor cells in peripheral blood are currently being evaluated for use in the diagnosis and management of a number of human solid tumors. In the prostate cancer field, these include RT-PCR assays for the detection of cells expressing PSA and PSM (Verkaik et WO 2004/050828 PCT/US2002/038264 al., 1997, Urol. Res. 25:373-384; Ghossein et 1995, J. Clin. Oncol. 13:1195-2000; Heston et 1995, Clin. Chem. 41:1687- 1688).

A further aspect of the invention is an assessment of the susceptibility that an individual has for developing cancer. In one embodiment, a method for predicting susceptibility to cancer comprises detecting 24P4C12 mRNA or 24P4C12 protein in a tissue sample, its presence indicating susceptibility to cancer, wherein the degree of 24P4C12 mRNA expression correlates to the degree of susceptibility. In a specific embodiment, the presence of 24P4C12 in prostate or other tissue is examined, with the presence of 24P4C12 in the sample providing an indication of prostate cancer susceptibility (or the emergence or existence of a prostate tumor). Similarly, one can evaluate the integrity 24P4C12 nucleotide and amino acid sequences in a biological sample, in order to identify perturbations in the structure of these molecules such as insertions, deletions, substitutions and the like. The presence of one or more perturbatons in 24P4C12 gene products in the sample is an indication of cancer susceptibility (or the emergence or existence of a tumor).

The invention also comprises methods for gauging tumor aggressiveness. In one embodiment, a method for gauging aggressiveness of a tumor comprises determining the level of 24P4C12 mRNA or 24P4C12 protein expressed by tumor cells, comparing the level so determined to the level of 24P4C12 mRNA or 24P4C12 protein expressed in a corresponding normal tissue taken from the same individual or a normal tissue reference sample, wherein the degree of 24P4C12 mRNA or 24P4C12 protein expression in the tumor sample relative to the normal sample indicates the degree of aggressiveness. In a specific embodiment, aggressiveness of a tumor is evaluated by determining the extent to which 24P4C12 is expressed in the tumor cells, with higher expression levels indicating more aggressive tumors. Another embodiment is the evaluation of the integrity of 24P4C12 nucleotide and amino acid sequences in a biological sample, in order to identify perturbations in the structure of these molecules such as insertions, deletions, substitutions and the like. The presence of one or more perturbations indicates more aggressive tumors.

Another embodiment of the invention is directed to methods for observing the progression of a malignancy in an individual over time. In one embodiment, methods for observing the progression of a malignancy in an individual over time comprise determining the level of 24P4C12 mRNA or 24P4C12 protein expressed by cells in a sample of the tumor, comparing the level so determined to the level of 24P4C12 mRNA cr 24P4C12 protein expressed in an equivalent tissue sample taken from the same individual at a different time, wherein the degree of 24P4C12 mRNA or 24P4C12 protein expression in the tumor sample over time provides information on the progression of the cancer. In a specific embodiment, the progression of a cancer is evaluated by determining 24P4C12 expression in the tumor cells over time, where increased expression overtime indicates a progression of the cancer. Also, one can evaluate the integrity 24P4C12 nudeotide and amino acid sequences in a biological sample in order to identify perturbations in the structure of these molecules such as insertions, deletions, substitutions and the like, where the presence of one or more perturbations indicates a progression of the cancer.

The above diagnostic approaches can be combined with any one of a wide variety of prognostic and diagnostic protocols known in the art For example, another embodiment of the invention is directed to methods for observing a coincidence between the expression of 24P4C12 gene and 24P4C12 gene products (or perturbations in 24P4C12 gene and 24P4C12 gene products) and a factor that is associated with malignancy, as a means for diagnosing and prognosticating the status of a tissue sample. A wide variety of factors associated with malignancy can be utilized, such as the expression of genes associated with malignancy PSA, PSCA and PSM expression for prostate cancer etc.) as well as gross cytological observations (see, e.g., Bocking et 1984, Anal. Quant. Cytol. 6(2):74-88; Epstein, 1995, Hum. Pathol. 26(2):223-9; Thorson etal., 1998, Mod.

Pathol. 11(6):543-51; Baisden et aL, 1999, Am. J. Surg, Pathol. 23(8):918-24). Methods forobserving a coincidence between the expression of 24P4C12 gene and 24P4C12 gene products (or perturbations in 24P4C12gene and 24P4C12 gene products) and another factor that is associated with malignancy are useful, for example, because the presence of a set of specific factors that coincide with disease provides information crucial for diagnosing and prognosticating the status of a tissue sample.

WO 2004/050828 PCT/US2002/038264 In one embodiment, methods for observing a coincidence between the expression of 24P4C12gene and 24P4C12 gene products (or perturbations in 24P4C12 gene and 24P4C12 gene products) and another factor associated with malignancy entails detecting the overexpression of 24P4C12 mRNA or protein in a tissue sample, detecting the overexpression of PSA mRNA or protein in a tissue sample (or PSCA or PSM expression), and observing a coincidence of 24P4C12 mRNA or protein and PSA mRNA or protein overexpression (or PSCA or PSM expression). In a specific embodiment, the expression of 24P4C12 and PSA mRNA in prostate tissue is examined, where the coincidence of 24P4C12 and PSA mRNA overexpression in the sample indicates the existence of prostate cancer, prostate cancer susceptbility or the emergence or status of a prostate tumor.

Methods for detecting and quantifying the expression of 24P4C12 mRNA or protein are described herein, and standard nudeic acid and protein detection and quantification technologies are well known in the art Standard methods for the detection and quantification of 24P4C12 mRNA include in situ hybridization using labeled 24P4C12 riboprobes, Northern blot and related techniques using 24P4C12 polynudeotide probes, RT-PCR analysis using primers specific for 24P4C12, and other amplification type detection methods, such as, for example, branched DNA, SISBA, TMA and the like. In a specific embodiment, semiquantitative RT-PCR is used to detect and quantify 24P4C12 mRNA expression. Any number of primers capable of amplifying 24P4C12 can be used for this purpose, including but not limited to the various primer sets specifically described herein. In a specific embodiment, polyclonal or monoclonal antibodies specifically reactive with the wild-type 24P4C12 protein can be used in an immunohistochemical assay of biopsied tissue.

IX.) Identification of Molecules That Interact With 24P4C12 The 24P4C12 protein and nucleic acid sequences disclosed herein allow a skilled artisan to identify proteins, small molecules and other agents that interact with 24P4C12, as well as pathways activated by 24P4C12 via any one of a variety of art accepted protocols. For example, one can utilize one of the so-called interaction trap systems (also referred to as the "two-hybrid assay'). In such systems, molecules interact and reconstitute a transcription factor which directs expression of a reporter gene, whereupon the expression of the reporter gene is assayed. Other systems identify protein-protein interactions in vivo through reconstitution of a eukaryotic transcriptional activator, see, U.S. Patent Nos. 5,955,280 issued 21 September 1999, 5,925,523 issued 20 July 1999, 5,846,722 issued 8 December 1998 and 6,004,746 issued 21 December 1999. Algorithms are also available in the art for genome-based predictions of protein function (see, Marcotte, et a., Nature 402: 4 November 1999, 83-86).

Alternatively one can screen peptide libraries to identify molecules that interact with 24P4C12 protein sequences.

In such methods, peptides that bind to 24P4C12 are identified by screening libraries that encode a random or controlled collection of amino acids. Peptides encoded by the libraries are expressed as fusion proteins of bacteriophage coat proteins, the bacteriophage particles are then screened against the 24P4C12 protein(s).

Accordingly, peptides having a wide variety of uses, such as therapeutic, prognostic or diagnostic reagents, are thus identified without any prior information on the structure of the expected ligand or receptor molecule. Typical peptide libraries and screening methods that can be used to identify molecules that interact with 24P4C12 protein sequences are disclosed for example in U.S. Patent Nos. 5,723,286 issued 3 March 1998 and 5,733,731 issued 31 March 1998.

Alternatively, cell lines that express 24P4C12 are used to identify protein-protein interactions mediated by 24P4C12. Such interactions can be examined using immunoprecipitation techniques (see, Hamilton etal.

Biochem. Biophys. Res. Commun. 1999, 261:646-51). 24P4C12 protein can be immunoprecipitated from 24P4C12expressing cell lines using anti-24P4C12 antibodies. Alternatively, antibodies against His-tag can be used in a cell line engineered to express fusions of 24P4C12 and a His-tag (vectors mentioned above). The immunoprecipitated complex can be examined for protein association by procedures such as Western blotting, 3S-methionine labeling of proteins, protein microsequencing, silver staining and two-dimensional gel electrophoresis.

WO 2004/050828 PCT/US2002/038264 Small molecules and ligands that interact with 24P4C12 can be identified through related embodiments of such screening assays For example, small molecules can be identified that interfere with protein function, including molecules that interfere with 24P4C12's ability to mediate phosphorylation and de-phosphorylation, interaction with DNA or RNA molecules as an indication of regulation of cell cycles, second messenger signaling or tumorigenesis. Similarly, small molecules that modulate 24P4C12-related ion channel, protein pump, or cell communication functions are identified and used to treat patients that have a cancer that expresses 24P4C12 (see, Hille, Ionic Channels of Excitable Membranes 2nd Ed., Sinauer Assoc., Sunderland, MA, 1992). Moreover, ligands that regulate 24P4C12 function can be identified based on their ability to bind 24P4C12 and activate a reporter construct. Typical methods are discussed for example in U.S. Patent No. 5,928,868 issued 27 July 1999, and include methods for forming hybrid ligands in which at least one ligand is a small molecule. In an illustrative embodiment, cells engineered to express a fusion protein of 24P4C12 and a DNA-binding protein are used to co-express a fusion protein of a hybrid ligandismall molecule and a cDNA library transcriptional activator protein. The cells further contain a reporter gene, the expression of which is conditioned on the proximity of the first and second fusion proteins to each other, an event that occurs only if the hybrid ligand binds to target sites on both hybrid proteins. Those cells that express the reporter gene are selected and the unknown small molecule or the unknown ligand is identified. This method provides a means of identifying modulators, which activate or inhibit 24P4C12.

An embodiment of this invention comprises a method of screening for a molecule that interacts with a 24P4C12 amino acid sequence shown in Figure 2 or Figure 3, comprising the steps of contacting a population of molecules with a 24P4C12 amino acid sequence, allowing the population of molecules and the 24P4C12 amino acid sequence to interact under conditions that facilitate an interaction, determining the presence of a molecule that interacts with the 24P4C12 amino acid sequence, and then separating molecules that do not interact with the 24P4C12 amino acid sequence from molecules that do. In a specific embodiment, the method further comprises purifying, characterizing and identifying a molecule that interacts with the 24P4C12 amino acid sequence. The identified molecule can be used to modulate a function performed by 24P4C12. In a preferred embodiment, the 24P4C12 amino acid sequence is contacted with a library of peptides.

Therapeutic Methods and Compositions The identification of 24P4C12 as a protein that is normally expressed in a restricted set of tissues, but which is also expressed in prostate and other cancers, opens a number of therapeutic approaches to the treatment of such cancers. As contemplated herein, 24P4C12 functions as a transcription factor involved in activating tumor-promoting genes or repressing genes that block tumorigenesis.

Accordingly, therapeutic approaches that inhibit the activity of a 24P4C12 protein are useful for patients suffering from a cancer that expresses 24P4C12. These therapeutic approaches generally fall into two classes. One class comprises various methods for inhibiting the binding or association of a 24P4C12 protein with its binding partner or with other proteins.

Another class comprises a variety of methods for inhibiting the transcription of a 24P4C12 gene or translation of 24P4C12 mRNA.

Anti-Cancer Vaccines The invention provides cancer vaccines comprising a 24P4C12-related protein or 24P4C12-related nucleic acid. In view of the expression of 24P4012, cancer vaccines prevent and/or treat 24P4C12-expressing cancers with minimal or no effects on non-target tissues. The use of a tumor antigen in a vaccine that generates humoral and/or cell-mediated immune responses as anti-cancer therapy is well known in the art and has been employed in prostate cancer using human PSMA and rodent PAP immunogens (Hodge et al, 1995, Int. J. Cancer63:231-237; Fong et al., 1997, J. Immunol. 159:3113-3117).

WO 2004/050828 PCT/US2002/038264 Such methods can be readily practiced by employing a 24P4C12-related protein, or a 24P4C12-encoding nucleic acid molecule and recombinant vectors capable of expressing and presenting the 24P4C12 immunogen (which typically comprises a number of antibody or T cell epitopes). Skilled artisans understand that a wide variety of vaccine systems for delivery of immunoreactive epitopes are known in the art (see, Heryln et at., Ann Med 1999 Feb 31(1):66-78; Maruyama et al, Cancer Immunol Immunother 2000 Jun 49(3):123-32) Briefly, such methods of generating an immune response (e.g.

humoral and/or cell-mediated) in a mammal, comprise the steps of: exposing the mammal's immune system to an immunoreactive epitope an epitope present in a 24P4C12 protein shown in Figure 3 or analog or homolog thereof) so that the mammal generates an immune response that is specific for that epitope generates antibodies that specifically recognize that epitope). In a preferred method, a 24P4C12 immunogen contains a biological motif, see Tables VIII-XXI and XXII-XLIX, or a peptide of a size range from 24P4C12 indicated in Figure 5, Figure 6, Figure 7, Figure 8, and Figure 9.

The entire 24P4C12 protein, immunogenic regions or epitopes thereof can be combined and delivered by various means. Such vaccine compositions can include, for example, lipopeptides (e.g.,Vitiello, A. et al., J. Clin. Invest. 95:341, 1995), peptide compositions encapsulated in poly(DL-lactide-co-glycolide) microspheres (see, Eldridge, et at, Molec. Immunol. 28:287-294, 1991: Alonso et Vaccine 12:299-306,1994; Jones et al., Vaccine 13:675-681, 1995), peptide compositions contained in immune stimulating complexes (ISCOMS) (see, Takahashi etal., Nature 344:873- 875,1990; Hu etal., Clin Exp Immunol. 113:235-243, 1998), multiple antigen peptide systems (MAPs) (see Tam, J. P., Proc. Natl. Acad. Sci. U.S.A. 85:5409-5413, 1988; Tam, J. Immunol. Methods 196:17-32, 1996), peptides formulated as multivalent peptides; peptides for use in ballistic delivery systems, typically crystallized peptides, viral delivery vectors (Perkus, M. E. et at., In: Concepts in vaccine development, Kaufmann, S. H. ed., p. 379, 1996; Chakrabarti, S. et al, Nature 320:535, 1986; Hu, S. L et Nature 320:537, 1986; Kieny, et al., AIDS Bio/Technology4:790, 1986; Top, F.

H. et al., J. Infect. Dis. 124:148, 1971; Chanda, P. K. et al, Virology 175:535, 1990), particles of viral or synthetic origin Kofler, N. et al, J. Immunol. Methods. 192:25,1996; Eldridge, J. H. et al., Sem. Hematol. 30:16, 1993; Falo, L. Jr. et a., Nature Med. 7:649,1995), adjuvants (Warren, H. Vogel, F. and Chedid, L. A. Annu. Rev. Immunol. 4:369,1986; Gupta, R. K. et at, Vaccine 11:293, 1993), liposomes (Reddy, R. et al, J. Immunol. 148:1585, 1992; Rock, K. L, ImmuncL Today 17:131, 1996), or, naked or particle absorbed cDNA (Ulmer, J. B. et al, Science 259:1745, 1993; Robinson, H. L., Hunt, L. and Webster, R. Vaccine 11:957,1993; Shiver, J. W. et In: Concepts in vaccine development, Kaufmann, S. H. ed., p. 423, 1996; Cease, K. and Berzofsky, J. Annu. Rev. Immunol. 12:923, 1994 and Eldridge, J. H. et a., Sem. Hematol. 30:16, 1993). Toxin-targeted delivery technologies, also known as receptor mediated targeting, such as those of Avant Immunotherapeutics, Inc. (Needham, Massachusetts) may also be used.

In patients with 24P4C12-associated cancer, the vaccine compositions of the invention can also be used in conjunction with other treatments used for cancer, surgery, chemotherapy, drug therapies, radiation therapies, etc.

including use in combination with immune adjuvants such as IL-2, IL-12, GM-CSF, and the like.

Cellular Vaccines: CTL epitopes can be determined using specific algorithms to identify peptides within 24P4C12 protein that bind corresponding HLA alleles (see Table IV; Epimerm and Epimatrix

T

Brown University (URL brown.edulResearch/TB- HIV_Lablepimatix/epimatrix.html); and, BIMAS, (URL bimas.dcrtnih.govl; SYFPEITHI at URL syfpeithi.bmi-heidelberg.coml).

In a preferred embodiment, a 24P4C12 immunogen contains one or more amino acid sequences identified using techniques well known in the art, such as the sequences shown in Tables VIII-XXI and XXII-XLIX or a peptide of 8, 9, 10 or 11 amino acids specified by an HLA Class I motiftsupermotif Table IV Table IV or Table IV and/or a peplide of at least 9 amino acids that comprises an HLA Class II motiflsupermotif Table IV or Table IV As is appreciated in the art, the HLA Class I binding groove is essentially closed ended so that peptides of only a particular size range can fit into the groove and be bound, generally HLA Class I epitopes are 8, 9, 10, or 11 amino acids long. In contrast, the HLA Class II WO 2004/050828 PCT/US20021038264 binding groove is essentially open ended; therefore a peptide of about 9 or more amino acids can be bound by an HLA Class II molecule. Due to the binding groove differences between HLA Class I and II, HLA Class I motifs are length specific, i.e., position two of a Class I motif is the second amino acid in ar amino to carboxyl direction of the peptide. The amino acid positions in a Class II motif are relative only to each other, not the overall peptide, additional amino acids can be attached to the amino and/or carboxyl termini of a motif-bearing sequence. HLA Class II epitopes are often 9, 10, 11, 12,13, 14, 15,16,17,18, 19, 20, 21, 22, 23, 24, or 25 amino acids long, or longer than 25 amino acids.

Antibody-based Vaccines A wide variety of methods for generating an immune response in a mammal are known in the art (for example as the first step in the generation of hybridomas). Methods of generating an immune response in a mammal comprise exposing the mammal's immune system to an immunogenic epitope on a protein a 24P4C12 protein) so that an immune response is generated. A typical embodiment consists of a method for generating an immune response to 24P4C12 in a host, by contacting the host with a sufficient amount of at least one 24P4C12 B cell or cytotoxic T-cell epitope or analog thereof; and at least one periodic interval thereafter re-contacting the host with the 24P4C12 B cell or cytotoxic T-cell epitope or analog thereof. A specific embodiment consists of a method of generating an immune response against a 24P4C12related protein or a man-made multiepitopic peptide comprising: administering 24P4C12 immunogen a 24P4C12 protein or a peptide fragment thereof, a 24P4C12 fusion protein or analog etc.) in a vaccine preparation to a human or another mammal. Typically, such vaccine preparations further contain a suitable adjuvant (see, U.S. Patent No.

6,146,635) or a universal helper epitope such as a PADRE M peptide (Epimmune Inc., San Diego, CA; see, Alexander et al., J. Immunol. 2000 164(3); 164(3): 1625-1633; Alexander et al., Immunity 1994 751-761 and Alexander etaL., Immunol. Res. 1998 18(2): 79-92). An alternative method comprises generating an immune response in an individual against a 24P4C12 immunogen by: administering in vivo to muscle or skin of the individual's body a DNA molecule that comprises a DNA sequence that encodes a 24P4C12 immunogen, the DNA sequence operatively linked to regulatory sequences which control the expression of the DNA sequence; wherein the DNA molecule is taken up by cells, the DNA sequence is expressed in the cells and an immune response is generated against the immunogen (see, U.S. Patent No.

5,962,428). Optionally a genetic vaccine facilitator such as anionic lipids; saponins; lectins; estrogenic compounds; hydroxylated lower alkyls; dimethyl sulfoxide; and urea is also administered. In addition, an antiidiotypic antibody can be administered that mimics 24P4C12, in order to generate a response to the target antigen.

Nucleic Add Vaccines: Vaccine compositions of the invention include nucleic acid-mediated modalities. DNA or RNA that encode protein(s) of the invention can be administered to a patient. Genetic immunization methods can be employed to generate prophylactic or therapeutic humoral and cellular immune responses directed against cancer cells expressing 24P4C12.

Constructs comprising DNA encoding a 24P4C12-related proteinlimmuncgen and appropriate regulatory sequences can be injected directly into muscle or skin of an individual, such that the cells of the muscle or skin take-up the construct and express the encoded 24P4C12 protein/immunogen. Alternatively, a vaccine comprises a 24P4C12-related protein.

Expression of the 24P4C12-related protein immunogen results in the generation of prophylactic or therapeutic humoral and cellular immunity against cells that bear a 24P4C12 protein. Various prophylactic and therapeutic genetic immunization techniques known in the art can be used (for review, see information and references published at Intemet address genweb.com). Nucleic acid-based delivery is described, for instance, in Wolff et. Science 247:1465 (1990) as well as U.S. Patent Nos. 5,580,859; 5,589,466; 5,804,566; 5,739,118; 5,736,524; 5,679,647; WO 98/04720. Examples of DNAbased delivery technologies include "naked DNA", facilitated (bupivicaine, polymers, peptide-mediated) delivery, catonic lipid complexes, and particle-mediated ("gene gun") or pressure-mediated delivery (see, U.S. Patent No. 5,922,687).

WO 2004/050828 PCT/US2002/038264 For therapeutic or prophylactic immunization purposes, proteins of the invention can be expressed via viral or bacterial vectors. Various viral gene delivery systems that can be used in the practice of the invention include, but are not limited to, vaccinia, fowlpox, canarypox, adenovirus, influenza, poliovirus, adeno-assocated virus, lentivirus, and sindbis virus (see, e.g., Restifo, 1996, Curr. Opin. Immunol. 8:658-663; Tsang etal. J. Nail. Cancer Inst 87:982-990 (1995)). Non-viral delivery systems can also be employed by introducing naked DNA encoding a 24P4C12-related protein into the patient intramuscularly or intradermally) to induce an anti-tumor response.

Vaccinia virus is used, for example, as a vector to express nucleotide sequences that encode the peptides of the invention. Upon introduction into a host, the recombinant vaccinia virus expresses the protein immunogenic peptide, and thereby elicits a host immune response. Vaccinia vectors and methods useful in immunization protocols are described in, U.S. Patent No. 4,722,848. Another vector is BCG (Bacille Calmette Guerin). BCG vectors are described in Stover et al., Nature 351:456-460 (1991). A wide variety of other vectors useful for therapeutic administration or immunization of the peptides of the invention, e.g. adeno and adeno-associated virus vectors, retroviral vectors, Salmonella typhi vectors, detoxified anthrax toxin vectors, and the like, will be apparent to those skilled in the art from the description herein.

Thus, gene delivery systems are used to deliver a 24P4C12-related nucleic acid molecule. In one embodiment, the fulllength human 24P4C12 cDNA is employed. In another embodiment, 24P4C12 nucleic acid molecules encoding specific cytotoxic T lymphocyte (CTL) and/or antibody epitopes are employed.

Ex Vivo Vaccines Various ex vivo strategies can also be employed to generate an immune response. One approach involves the use of antigen presenting cells (APCs) such as dendritic cells (DC) to present 24P4C12 antigen to a patient's immune system. Dendritic cells express MHC class I and II molecules, B7 co-stimulator, and IL-12, and are thus highly specialized antigen presenting cells.

In prostate cancer, autologous dendritic cells pulsed with peptides of the prostate-specific membrane antigen (PSMA) are being used in a Phase I clinical trial to stimulate prostate cancer patients' immune systems (Tjoa et al., 1996, Prostate 28:65- 69; Murphy et al., 1996, Prostate 29:371-380). Thus, dendritic cells can be used to present 24P4C12 peptides to T cells in the context of MHC class I or II molecules. In one embodiment, autologous dendritic cells are pulsed with 24P4C12 peptides capable of binding to MHC class I and/or class II molecules. In another embodiment, dendritic cells are pulsed with the complete 24P4C12 protein. Yet another embodiment involves engineering the overexpression of a 24P4C12 gene in dendritic cells using various implementing vectors known in the art, such as adenovirus (Arthur etal., 1997, Cancer Gene Ther. 4:17-25), retrovirus (Henderson el al., 1996, Cancer Res. 56:3763-3770), lentivirus, adeno-associated virus, DNA transfection (Ribas et al., 1997, Cancer Res. 57:2865-2869), or tumor-derived RNA transfection (Ashley et al., 1997, J. Exp.

Med. 186:1177-1182). Cells that express 24P4C12 can also be engineered to express immune modulators, such as GM- CSF, and used as immunizing agents.

24P4C12 as a Target for Antibody-based Therapy 24P4C12 is an attractive target for antibody-based therapeutic strategies. A number of antibody strategies are known in the art for targeting both extracellular and intracellular molecules (see, complement and ADCC mediated killing as well as the use of intrabodies). Because 24P4C12 is expressed by cancer cells of various lineages relative to corresponding normal cells, systemic administration of 24P4C12-immunoreactive compositions are prepared that exhibit excellent sensitivity without toxic, non-specific and/or non-target effects caused by binding of the immunoreactive composition to non-target organs and tissues. Antibodies specifically reactive with domains of 24P4C12 are useful to treat 24P4C12-expressing cancers systemically, either as conjugates with a toxin or therapeutic agent, or as naked antibodies capable of inhibiting cell proliferation or function.

WO 2004/050828 PCT/US2002/038264 24P4C12 antibodies can be introduced into a patient such that the antibody binds to 24P4C12 and modulates a function, such as an interaction with a binding partner, and consequently mediates destruction of the tumor cells and/or inhibits the growth of the tumor cells. Mechanisms by which such antibodies exert a therapeutic effect can include complement-mediated cytolysis, antibody-dependent cellular cytotoxicity, modulation of the physiological function of 24P4C12, inhibition of ligand binding or signal Iransduction pathways, modulation of tumor cell differentiation, alteration of tumor angiogenesis factor profiles, and/or apoptosis.

Those skilled in the art understand that antibodies can be used to specifically target and bind immunogenic molecules such as an immunogenic region of a 24P4C12 sequence shown in Figure 2 or Figure 3. In addition, skilled artisans understand that it is routine to conjugate antibodies to cytotoxic agents (see, Slevers at at. Blood 93:11 3678- 3684 (June 1, 1999)). When cytotoxic and/or therapeutic agents are delivered directly to cells, such as by conjugating them to antibodies specific for a molecule expressed by that cell 24P4C12), the cytotoxic agent will exert its known biological effect cytotoxicity) on those cells.

A wide variety of compositions and methods for using antibody-cytotoxic agent conjugates to kill cells are known in the art. In the context of cancers, typical methods entail administering to an animal having a tumor a biologically effective amount of a conjugate comprising a selected cytotoxic and/or therapeutic agent linked to a targeting agent an anti- 24P4C12 antibody) that binds to a marker 24P4C12) expressed, accessible to binding or localized on the cell surfaces.

A typical embodiment is a method of delivering a cytotoxic and/or therapeutic agent to a cell expressing 24P4C12, comprising conjugating the cytotoxic agent to an antibody that immunospecifically binds to a 24P4C 12 epitope, and, exposing the cell to the antibody-agent conjugate. Another illustrative embodiment is a method of treating an individual suspected of sjffering from metastasized cancer, comprising a step of administering parenterally to said individual a pharmaceutical composition comprising a therapeutically effective amount of an antibody conjugated to a cytotoxic and/or therapeutic agent.

Cancer immunotherapy using anti-24P4C12 antibodies can be done in accordance with various approaches that have been successfully employed in the treatment of other types of cancer, including but not limited to colon cancer (Arlen el al., 1998, Crit. Rev. Immunol. 18:133-138), multiple myeloma (Ozaki etal., 1997, Blood 90:3179-3186, Tsunenari etal., 1997, Blood 90:2437-2444), gastric cancer (Kasprzyk et al., 1992, Cancer Res. 52:2771-2776), B-cell lymphoma (Funakoshi etal., 1996, J. Immunother. Emphasis Tumor Immunol. 19:93-101), leukemia (Zhong et al., 1996, Leuk. Res. 20:581-589), colorectal cancer (Moun et al., 1994, Cancer Res. 54:6160-6166, Velders et 1995, Cancer Res. 55:4398-4403), and breast cancer (Shepard et al., 1991, J. Clin. Immunol. 11:117-127). Some therapeutic approaches involve conjugation of naked antibody to a toxin or radioisotope, such as the conjugation of Y91 or I 13 to anti-CD20 antibodies Zevalin

T

IDEC

Pharmaceuticals Corp. or BexxarTM, Coulter Pharmaceuticals), while others involve co-administration of antibodies and other therapeutic agents, such as Herceptin T M (trastuzumab) with paclitaxel (Genentech, Inc.). The antibodies can be conjugated to a therapeutic agent. To treat prostate cancer, for example, 24P4C12 antibodies can be administered in conjunction with radiation, chemotherapy or hormone ablation. Also, antibodies can be conjugated to a toxin such as calicheamicin Mylotarg

T

Wyeth-Ayerst, Madison, NJ, a recombinant humanized IgG4 kappa antibody conjugated to antitumor antibiotic calicheamicin) or a maytansinoid taxane-based Tumor-Activated Prodrug, TAP, platform, ImmunoGen, Cambridge, MA, also see US Patent 5,416,064).

Although 24P4C12 antibody therapy is useful for all stages of cancer, antibody therapy can be particularly appropriate in advanced or metastatic cancers. Treatment with the antibody therapy of the invention is indicated for patients who have received one or more rounds of chemotherapy. Alternatively, antibody therapy of the invention is combined with a chemotherapeutic or radiation regimen for patients who have not received chemotherapeutic treatment. Additionally, antibody therapy can enable the use of reduced dosages of concomitant chemotherapy, particularly for patients who do not WO 2004/050828 PCT/US2002/038264 tolerate the toxicity of the chemotherapeutic agent very well. Fan et al. (Cancer Res. 53:4637-4642, 1993), Prewett et al.

(International J. of Onco. 9:217-224, 1996), and Hancock et al. (Cancer Res. 51:4575-4580, 1991) describe the use of various antibodies together with chemotherapeutic agents.

Although 24P4C12 antibody therapy is useful for all stages of cancer, antibody therapy can be particularly appropriate in advanced or metastatic cancers. Treatment with the antibody therapy of the invention is indicated for patients who have received one or more rounds of chemotherapy. Alternatively, antibody therapy of the invention is combined with a chemotherapeutic or radiation regimen for patients who have not received chemotherapeutic treatment. Additionally, antibody therapy can enable the use of reduced dosages of concomitant chemotherapy, particularly for patients who do not tolerate the toxicity of the chemotherapeutic agent very well.

Cancer patients can be evaluated for the presence and level of 24P4C12 expression, preferably using immunohistochemical assessments of tumor tissue, quantitative 24P4C12 imaging, or other techniques that reliably indicate the presence and degree of 24P4C12 expression. Immunohistochemical analysis of tumor biopsies or surgical specimens is preferred for this purpose. Methods for immunohistochemical analysis of tumor tissues are well known in the art.

Anti-24P4C12 monoclonal antibodies that treat prostate and other cancers include those that initiate a potent immune response against the tumor or those that are directly cytotoxic. In this regard, anti-24P4C12 monoclonal antibodies (mAbs) can elicit tumor cell lysis by either complement-mediated or antibody-dependent cell cytotoxicity (ADCC) mechanisms, both of which require an intact Fc portion of the immunoglobulin molecule for interaction with effector cell Fc receptor sites on complement proteins. In addition, anti-24P4C12 mAbs that exert a direct biological effect on tumor growth are useful to treat cancers that express 24P4C12, Mechanisms by which directly cytotoxic mAbs act include: inhibition of cell growth, modulation of cellular differentiation, modulation of tumor angiogenesis factor profiles, and the induction of apoptosis.

The mechanism(s) by which a particular anti-24P4C12 mAb exerts an anti-tumor effect is evaluated using any number of in vitro assays that evaluate cell death such as ADCC, ADMMC, complement-mediated cell lysis, and so forth, as is generally known in the art.

In some patients, the use of murine or other non-human monoclonal antibodies, or human/mouse chimeric mAbs can induce moderate to strong immune responses against the non-human antibody. This can result in clearance of the antibody from circulation and reduced efficacy. In the most severe cases, such an immune response can lead to the extensive formation of immune complexes which, potentially, can cause renal failure. Accordingly, preferred monoclonal antibodies used in the therapeutic methods of the invention are those that are either fully human or humanized and that bind specifically to the target 24P4C12 antigen with high affinity but exhibit low or no antigenicity in the patient.

Therapeutic methods of the invention contemplate the administration of single anti-24P4C12 mAbs as well as combinations, or cocktails, of different mAbs. Such mAb cocktails can have certain advantages inasmuch as they contain mAbs that target different epitopes, exploit different effector mechanisms or combine directly cytotoxic mAbs with mAbs that rely on immune effector functionality. Such mAbs in combination can exhibit synergistic therapeutic effects. In addition, anti- 24P4C12 mAbs can be administered concomitantly with other therapeutic modalities, including but not limited to various chemotherapeutic agents, androgen-blockers, immune modulators IL-2, GM-CSF), surgery or radiation. The anti- 24P4C12 mAbs are administered in their "naked" or unconjugated form, or can have a therapeutic agent(s) conjugated to them.

Anti-24P4C12 antibody formulations are administered via any route capable of delivering the antibodies to a tumor cell. Routes of administration include, but are not limited to, intravenous, intraperitoneal, intramuscular, intratumor, intradermal, and the like. Treatment generally involves repeated administration of the anti-24P4C12 antibody preparation, via an acceptable route of administration such as intravenous injection typically ata dose in the range of about 0.1, .2, WO 2004/050828 PCT/US2002/038264 1,2,3,4,5,6,7, 8, 9, 10, 15,20, or 25 mg/kg body weight. In general, doses in the range of 10-1000 mg mAb per week are effective and well tolerated.

Based on clinical experience with the HerceptinTM mAb in the treatment of metastatic breast cancer, an initial loading dose of approximately 4 mg/kg patient body weight IV, followed by weekly doses of about 2 mg/kg IV of the anti- 24P4C12 mAb preparation represents an acceptable dosing regimen. Preferably, the initial loading dose is administered as a 90-minute or longer infusion. The periodic maintenance dose is administered as a 30 minute or longer infusion, provided the initial dose was well tolerated. As appreciated by those of skill in the art, various factors can influence the ideal dose regimen in a particular case. Such factors include, for example, the binding affinity and half life of the Ab or mAbs used, the degree of 24P4C12 expression in the patient, the extent of circulating shed 24P4C12 antigen, the desired steady-state antibody concentration level, frequency of treatment, and the influence of chemotherapeutic or other agents used in combination with the treatment method of the invention, as well as the health status of a particular patient.

Optionally, patients should be evaluated for the levels of 24P4C12 in a given sample the levels of circulating 24P4C12 antigen and/or 24P4C12 expressing cells) in order to assist in the determination of the most effective dosing regimen, etc. Such evaluations are also used for monitoring purposes throughout therapy, and are useful to gauge therapeutic success in combination with the evaluation of other parameters (for example, urine cytology and/or ImmunoCyt levels in bladder cancer therapy, or by analogy, serum PSA levels in prostate cancer therapy).

Anti-idiotypic anti-24P4C12 antibodies can also be used in anti-cancer therapy as a vaccine for inducing an immune response to cells expressing a 24P4C12-related protein. In particular, the generation of anti-idiotypic antibodies is well known in the art; this methodology can readily be adapted to generate anti-idiotypic anti-24P4C12 antibodies that mimic an epitope on a 24P4C12-related protein (see, for example, Wagner etal., 1997, Hybridoma 16: 33-40; Foon et 1995, J.

Clin. Invest. 96:334-342; Herlyn et at., 1996, Cancer Immunol. Immunother. 43:65-76). Such an anti-idiotypic antibody can be used in cancer vaccine strategies.

24P4C12 as a Target for Cellular Immune Responses Vaccines and methods of preparing vaccines that contain an immunogenically effective amount of one or more HLA-binding peptides as described herein are further embodiments of the invention. Furthermore, vaccines in accordance with the invention encompass compositions of one or more of the claimed peptides. A peptide can be present in a vaccine individually. Alternatively, the peptide can exist as a homopolymer comprising multiple copies of the same peptide, or as a heteropolymer of various peptides. Polymers have the advantage of increased immunological reaction and, where different peptide epitopes are used to make up the polymer, the additional ability to induce antibodies and/or CTLs that react with different antigenic determinants of the pathogenic organism or tumor-related peptide targeted for an immune response. The composition can be a naturally occurring region of an antigen or can be prepared, recombinantly or by chemical synthesis.

Carriers that can be used with vaccines of the invention are well known in the art, and include, thyroglobulin, albumins such as human serum albumin, tetanus toxoid, polyamino acids such as poly L-lysine, poly L-glutamic acid, influenza, hepatitis 8 virus core protein, and the like. The vaccines can contain a physiologically tolerable acceptable) diluent such as water, or saline, preferably phosphate buffered saline. The vaccines also typically include an adjuvant.

Adjuvants such as incomplete Freund's adjuvant, aluminum phosphate, aluminum hydroxide, or alum are examples of materials well known in the art. Additionally, as disclosed herein, CTL responses can be primed by conjugating peptides of the invention to lipids, such as tripalmitoyl-S-glycerylcysteinlyseryl- serine (P3CSS). Moreover, an adjuvant such as a synthetic cytosine-phosphorothiolated-guanine-containing (CpG) oligonudeotides has been found to increase CTL responses 10-to 100-fold. (see, e.g. Davila and Cells, J. Immunol. 165:539-547 (2000)) WO 2004/050828 PCT/US2002/038264 Upon immunization with a peptide composition in accordance with the invention, via injection, aerosol, oral, transdermal, transmucosal, intrapleural, intrathecal, or other suitable routes, the immune system of the host responds to the vaccine by producing large amounts of CTLs and/or HTLs specific for the desired antigen. Consequently, the host becomes at least partially immune to later development of cells that express or overexpress 24P4C12 antigen, or derives at least some therapeutic benefit when the antigen was tumor-associated.

SIn some embodiments, it may be desirable to combine the class I peptide components with components that induce or facilitate neutralizing antibody and or helper T cell responses directed to the target antigen. A preferred embodiment of such a composition comprises class I and class II epitopes in accordance with the invention. An alternative embodiment of such a composition comprises a class I and/or class II epitope in accordance with the invention, along with a cross reactive HTL epitope such as PADRE T M (Epimmune, San Diego, CA) molecule (described in U.S. Patent Number 5,736,142).

A vaccine of the invention can also include antigen-presenting cells (APC), such as dendritic cells as a vehicle to present peptides of the invention. Vaccine compositions can be created in vitro, following dendritic cell mobilization and harvesting, whereby loading of dendritic cells occurs in vitro. For example, dendritic cells are transfected, with a minigene in accordance with the invention, or are pulsed with peptides. The dendritic cell can then be administered to a patient to elicit immune responses in vivo. Vaccine compositions, either DNA- or peptide-based, can also be administered in vivo in combination with dendritic cell mobilization whereby loading of dendritic cells occurs in vivo.

Preferably, the following principles are utilized when selecting an array of epitopes for inclusion in a polyepitopic composition for use in a vaccine, or for selecting discrete epitopes to be included in a vaccine and/or to be encoded by nucleic acids such as a minigene. It is preferred that each of the following principles be balanced in order to make the selection. The multiple epitopes to be incorporated in a given vaccine composition may be, but need not be, contiguous in sequence in the native antigen from which the epitopes are derived.

Epitopes are selected which, upon administration, mimic immune responses that have been observed to be correlated with tumor clearance. For HLA Class I this includes 3-4 epitopes that come from at least one tumor associated antigen (TAA). For HLA Class II a similar rationale is employed; again 3-4 epitopes are selected from at least one TAA (see, Rosenberg et al., Science 278:1447-1450). Epitopes from one TAA may be used in combination with epilopes from one or more additional TAAs to produce a vaccine that targets tumors with varying expression patterns of frequently-expressed TAAs.

Epitopes are selected that have the requisite binding affinity established to be correlated with immunogenicity: for HLA Class I an ICso of 500 nM or less, often 200 nM or less; and for Class II an ICso of 1000 nM or less.

Sufficient supermotif bearing-peptides, or a sufficient array of allele-specific motif-bearing peptides, are selected to give broad population coverage. For example, it is preferable to have at least 80% population coverage. A Monte Carlo analysis, a statistical evaluation known in the art, can be employed to assess the breadth, or redundancy of, population coverage.

When selecting epitopes from cancer-related antigens it is often useful to select analogs because the patient may have developed tolerance to the native epitope.

Of particular relevance are epitopes referred to as "nested epitopes." Nested epitopes occur where at least two epitopes overlap in a given peptide sequence. A nested peptide sequence can comprise B cell, HLA class I and/or HLA class II epitopes. When providing nested epitopes, a general objective is to provide the greatest number of epitopes per sequence. Thus, an aspect is to avoid providing a peptide that is any longer than the amino terminus of the amino terminal epitope and the carboxyl terminus of the carboxyl terminal epitope in the peptide. When providing a multi-epitopic sequence, WO 2004/050828 PCT/US2002/038264 such as a sequence comprising nested epitopes, it is generally important to screen the sequence in order to insure that it does not have pathological or other deleterious biological properties.

If a polyepitopic protein is created, or when creating a minigene, an objective is to generate the smallest peptide that encompasses the epitopes of interest. This principle is similar, if not the same as that employed when selecting a peptide comprising nested epitopes. However, with an artificial polyepitopic peptide, the size minimization objective is balanced against the need to integrate any spacer sequences between epitopes in the polyepitopic protein. Spacer amino add residues can, for example, be introduced to avoid junctional epitopes (an epitope recognized by the immune system, not present in the target antigen, and only created by the man-made juxtaposition of epitopes), or to facilitate cleavage between epitopes and thereby enhance epitope presentation. Junctional epitopes are generally to be avoided because the recipient may generate an immune response to that non-native epitope. Of particular concern is a junctional epitope that is a "dominant epitope." A dominant epitope may lead to such a zealous response that immune responses to other epitopes are diminished or suppressed.

Where the sequences of multiple variants of the same target protein are present, potential peptide epitopes can also be selected on the basis of their conservancy. For example, a criterion for conservancy may define that the entire sequence of an HLA class I binding peptide or the entire 9-mer core of a class II binding peptide be conserved in a designated percentage of the sequences evaluated for a specific protein antigen.

X.C.1. Minigene Vaccines A number of different approaches are available which allow simultaneous delivery of multiple epitopes. Nucleic acids encoding the peptides of the invention are a particularly useful embodiment of the invention. Epitopes for inclusion in a minigene are preferably selected according to the guidelines set forth in the previous section. A preferred means of administering nucleic acids encoding the peptides of the invention uses minigene constructs encoding a peptide comprising one or multiple epitopes of the invention.

The use of multi-epitope minigenes is described below and in, Ishioka et aL, J. Immunol. 162:3915-3925, 1999; An, L. and Whitton, J. J. Virol. 71:2292, 1997; Thomson, S. A. et al, J. Immunol. 157:822, 1996; Whitton, J. L. et al., J. Virol.

67:348, 1993; Hanke, R. et Vaccine 16:426, 1998. For example, a multi-epitope DNA plasmid encoding supermotifand/or motif-bearing epitopes derived 24P4C12, the PADRE® universal helper T cell epitope or multiple HTL epitopes from 24P4C12 (see Tables VIII-XXI and XXII to XLIX), and an endoplasmic reticulum-translocating signal sequence can be engineered. A vaccine may also comprise epitopes that are derived from other TAAs.

The immunogenicity of a multi-epitopic minigene can be confirmed in transgenic mice to evaluate the magnitude of CTL induction responses against the epitopes tested. Further, the immunogenicity of DNA-encoded epitopes in vive can be correlated with the in vitro responses of specific CTL lines against target cells transfected with the DNA plasmid. Thus, these experiments can show that the minigene serves to both: generate a CTL response and that the induced CTLs recognized cells expressing the encoded epitopes.

For example, to create a DNA sequence encoding the selected epitopes (minigene) for expression in human cells, the amino acid sequences of the epitopes may be reverse translated. A human codon usage table can be used to guide the codon choice for each amino acid. These epitope-encoding DNA sequences may be directly adjoined, so that when translated, a continuous polypeptide sequence is created. To optimize expression and/or immunogenicity, additional elements can be incorporated into the minigene design. Examples of amino acid sequences that can be reverse translated and included in the minigene sequence include: HLA class I epitopes, HLA class II epitopes, antibody epitopes, a ubiquitination signal sequence, and/or an endoplasmic reticulum targeting signal. In addition, HLA presentation of CTL and HTL epitopes may be improved by including synthetic poly-alanine) or naturally-occurring flanking sequences adjacent to the CTL or HTL epitopes; these larger peptides comprising the epitope(s) are within the scope of the invention.

WO 2004/050828 PCT/US20021038264 The minigene sequence may be converted to DNA by assembling oligonucleotides that encode the plus and minus strands of the minigene. Overlapping oligonucleotides (30-100 bases long) may be synthesized, phosphorylated, purified and annealed under appropriate conditions using well known techniques. The ends of the oligonucleotides can be joined, for example, using T4 DNA ligase. This synthetic minigene, encoding the epitope polypeptide, can then be cloned into a desired expression vector.

Standard regulatory sequences well known to those of skill in the art are preferably included in the vector to ensure expression in the target cells. Several vector elements are desirable: a promoter with a down-stream cloning site for minigene insertion; a polyadenylation signal for efficient transcription termination; an E coJi origin of replication; and an E.

coli selectable marker ampicillin or kanamycin resistance). Numerous promoters can be used for this purpose, the human cytomegalovirus (hCMV) promoter. See, U.S. Patent Nos. 5,580,859 and 5,589,466 for other suitable promoter sequences.

Additional vector modifications may be desired to optimize minigene expression and immunogenicity. In some cases, introns are required for efficient gene expression, and one or more synthetic or naturally-occurring introns could be incorporated into the transcribed region of the minigene. The inclusion of mRNA stabilizaton sequences and sequences for replication in mammalian cells may also be considered for increasing minigene expression.

Once an expression vector is selected, the minigene is cloned into the polylinker region downstream of the promoter. This plasmid is transformed into an appropriate E coli strain, and DNA is prepared using standard techniques.

The orientation and DNA sequence of the minigene, as well as all other elements included in the vector, are confirmed using restriction mapping and DNA sequence analysis. Bacterial cells harboring the correct plasmid can be stored as a master cell bank and a working cell bank.

In addition, immunostimulatory sequences (ISSs or CpGs) appear to play a role in the immunogenicity of DNA vaccines. These sequences may be included in the vector, outside the minigene coding sequence, if desired to enhance immunogenicity.

In some embodiments, a bi-cistronic expression vector which allows production of both the minigene-encoded epitopes and a second protein (included to enhance or decrease immunogenicity) can be used. Examples of proteins or polypeptides that could beneficially enhance the immune response if co-expressed include cytokines IL-2, IL-12, GM- CSF), cytokine-inducing molecules LelF), costimulatory molecules, or for HTL responses, pan-DR binding proteins

(PADRE

T

Epimmune, San Diego, CA). Helper (HTL) epitopes can be joined to intracellular targeting signals and expressed separately from expressed CTL epitopes; this allows direction of the HTL epitopes to a cell compartment different than that of the CTL epitopes. If required, this could facilitate more efficient entry of HTL epitopes into the HLA class II pathway, thereby improving HTL induction. In contrast to HTL or CTL induction, specifically decreasing the immune response by co-expression of immunosuppressive molecules TGF-p) may be beneficial in certain diseases.

Therapeutic quantities of plasmid DNA can be produced for example, by fermentation in E. coli, followed by purification. Aliquots from the working cell bank are used to inoculate growth medium, and grown to saturation in shaker flasks or a bioreactor according to well-known techniques. Plasmid DNA can be purified using standard bioseparation technologies such as solid phase anion-exchange resins supplied by QIAGEN, Inc. (Valencia, California). If required, supercoiled DNA can be isolated from the open circular and linear forms using gel electrophoresis or other methods.

Purified plasmid DNA can be prepared for injection using a variety of formulations. The simplest of these is reconstitution of lyophilized DNA in sterile phosphate-buffer saline (PBS). This approach, known as "naked DNA." is currently being used for intramuscular (IM) administration in clinical trials. To maximize the immunotherapeutic effects of minigene DNA vaccines, an alternative method for formulating purified plasmid DNA may be desirable. A variety of methods have been described, and new techniques may become available. Cationic lipids, glycolipids, and fusogenic liposomes can WO 2004/050828 PCT/US2002/038264 also be used in the formulation (see, as described by WO 93/24640; Mannino Gould-Fogerite, BioTechniques 6(7): 682 (1988); U.S. Pat No. 5,279,833; WO 91/06309; and Feigner, et al, Proc. Natl Acad. Sci. USA 84:7413 (1987). In addition, peptides and compounds referred to collectively as protective, interactive, non-condensing compounds (PINC) could also be complexed to purified plasmid DNA to influence variables such as stability, intramuscular dispersion, or trafficking to specific organs or cell types.

Target cell sensitization can be used as a functional assay for expression and HLA class I presentation of minigene-encoded CTL epitopes. For example, the plasmid DNA is introduced into a mammalian cell line that is suitable as a target for standard CTL chromium release assays. The transfection method used will be dependent on the final formulation. Elcctroporation can be used for "naked" DNA, whereas cationic lipids allow direct in vitro transfection. A plasmid expressing green fluorescent protein (GFP) can be co-transfected to allow enrichment of transfected cells using fluorescence activated cell sorting (FACS). These cells are then chromium-51 5 Cr) labeled and used as target cells for epitope-specific CTL lines; cytolysis, detected by 51Cr release, indicates both production of, and HLA presentation of, minigene-encoded CTL epitopes. Expression of HTL epitopes may be evaluated in an analogous manner using assays to assess HTL activity.

In vivo immunogenicity is a second approach for functional testing of minigene DNA formulations. Transgenic mice expressing appropriate human HLA proteins are immunized with the DNA product. The dose and route of administration are formulation dependent IM for DNA in PBS, intraperitoneal for lipid-complexed DNA). Twenty-one days after immunization, splenocytes are harvested and restimulated for one week in the presence of peptides encoding each epitope being tested. Thereafter, for CTL effector cells, assays are conducted for cytolysis of peptide-loaded, 51 Cr-labeled target cells using standard techniques. Lysis of target cells that were sensitized by HLA loaded with peptide epitopes, corresponding to minigene-encoded epitopes, demonstrates DNA vaccine function for in vive induction of CTLs.

Immunogenicity of HTL epitopes is confirmed in transgenic mice in an analogous manner.

Alternatively, the nucleic acids can be administered using ballistic delivery as described, for instance, in U.S.

Patent No. 5,204,253. Using this technique, particles comprised solely of DNA are administered. In a further alternative embodiment, DNA can be adhered to particles, such as gold particles.

Minigenes can also be delivered using other bacterial or viral delivery systems well known in the art, an expression construct encoding epitopes of the invention can be incorporated into a viral vector such as vaccinia.

X.C.2. Combinations of CTL Peptides with Helper Peptides Vaccine compositions comprising CTL peptides of the invention can be modified, analoged, to provide desired attributes, such as improved serum half life, broadened population coverage or enhanced immunogenicity.

For instance, the ability of a peptide to induce CTL activity can be enhanced by linking the peptide to a sequence which contains at least one epitope that is capable of inducing a T helper cell response. Although a CTL peptide can be directly linked to a T helper peptide, often CTL epitopelHTL epitope conjugates are linked by a spacer molecule. The spacer is typically comprised of relatively small, neutral molecules, such as amino acids or amino acid mimetics, which are substantially uncharged under physiological conditions. The spacers are typically selected from, Ala, Gly, or other neutral spacers of nonpolar amino acids or neutral polar amino acids. It will be understood that the optionally present spacer need not be comprised of the same residues and thus may be a hetero- or homo-oligomer. When present, the spacer will usually be at least one or two residues, more usually three to six residues and sometimes 10 or more residues. The CTL peptide epitope can be linked to the T helper peptide epitope either directly or via a spacer either at the amino or carboxy terminus of the CTL peptide. The amino terminus of either the immunogenic peptide or the T helper peptide may be acylated.

WO 2004/050828 PCT/US2002/038264 In certain embodiments, the T helper peptide is one that is recognized by T helper cells present in a majority of a genetically diverse population. This can be accomplished by selecting peptides that bind to many, most, or all of the HLA class II molecules. Examples of such amino acid bind many HLA Class II molecules include sequences from antigens such as tetanus toxoid at positions 830-843 (QYIKANSKFIGITE; SEQ ID NO: 29), Plasmodium falciparum circumsporozoite (CS) protein at positions 373-398 (DIEKKIAKMEKASSVFNWNS; SEQ ID NO: 30), and Streptococcus 18kD protein at positions 116-131 (GAVDSILGGVATYGAA; SEQ ID NO: 31). Other examples include peptides bearing a DR 1-4-7 supermotif, or either of the DR3 motifs.

Alternatively, it is possible to prepare synthetic peptides capable of stimulating T helper lymphocytes, in a loosely HLA-restricted fashion, using amino acid sequences not found in nature (see, PCT publication WO 95/07707). These synthetic compounds called Pan-DR-binding epitopes PADRE

T

Epimmune, Inc., San Diego, CA) are designed, most preferably, to bind most HLA-DR (human HLA class II) molecules. For instance, a pan-DR-binding epitope peptide having the formula: AKXVAAWTLKAAA (SEQ ID NO: 32), where is either cyclohexylalanine, phenylalanine, or tyrosine, and a is either o-alanine or L-alanine, has been found to bind to most HLA-DR alleles, and to stimulate the response of T helper lymphocytes from most individuals, regardless of their HLA type. An alternative of a pan-DR binding epitope comprises all natural amino acids and can be provided in the form of nucleic acids that encode the epitope.

HTL peptide epitopes can also be modified to alter their biological properties. For example, they can be modified to include D-amino acids to increase their resistance to proteases and thus extend their serum half life, or they can be conjugated to other molecules such as lipids, proteins, carbohydrates, and the like to increase their biological activity. For example, a T helper peptide can be conjugated to one or more palmitic acid chains at either the amino or carboxyl termini.

X.C.3. Combinations of CTL Peptides with T Cell Priming Agents In some embodiments it may be desirable to include in the pharmaceutical compositions of the invention at least one component which primes B lymphocytes or T lymphocytes. Lipids have been identified as agents capable of priming CTL in vivo. For example, palmitic acid residues can be attached to the e-and a- amino groups of a lysine residue and then linked, via one or more linking residues such as Gly, Gly-Gly-, Ser, Ser-Ser, or the like, to an immunogenic peptide.

The lipidated peptide can then be administered either directly in a micelle or particle, incorporated into a liposome, or emulsified in an adjuvant, incomplete Freund's adjuvant. In a preferred embodiment, a particularly effective immunogenic composition comprises palmitic acid attached to e- and ax- amino groups of Lys, which is attached via linkage, Ser-Ser, to the amino terminus of the immunogenic peptide.

As another example of lipid priming of CTL responses, E. coil lipoproteins, such as tripalmitoyl-Sglycerylcysteinlyseryl- serine (P3CSS) can be used to prime virus specific CTL when covalently attached to an appropriate peptide (see, Deres, et Nature 342:561, 1989). Peptides of the invention can be coupled to P3CSS, for example, and the lipopeptide administered to an individual to prime specifically an immune response to the target antigen. Moreover, because the induction of neutralizing antibodies can also be primed with PsCSS-conjugated epitopes, two such compositions can be combined to more effectively elicit both humoral and cell-mediated responses.

X.C.4. Vaccine Compositions Comprising DC Pulsed with CTL and/or HTL Peptides An embodiment of a vaccine composition in accordance with the invention comprises ex vivo administration of a cocktail of epitope-bearing peptides to PBMC, or isolated DC therefrom, from the patient's blood. A pharmaceutical to facilitate harvesting of DC can be used, such as Progenipoietin T M (Pharmacia-Monsanto, St. Louis, MO) or GM-CSFIIL-4.

After pulsing the DC with peptides and prior to reinfusion into patients, the DC are washed to remove unbound peptides. In this embodiment, a vaccine comprises peptide-pulsed DCs which present the pulsed peptide epitopes complexed with HLA molecules on their surfaces.

WO 2004/050828 PCT/US2002/038264 The DC can be pulsed ex vivo with a cocktail of peptides, some of which stimulate CTL responses to 24P4C12.

Optionally, a helper T cell (HTL) peptide, such as a natural or artificial loosely restricted HLA Class II peptide, can be included to facilitate the CTL response. Thus, a vaccine in accordance with the invention is used to treat a cancer which expresses or overexpresses 24P4C12.

X.D. Adoptive Immunotherapy Antigenic 24P4C12-related peptides are used to elicit a CTL andlor HTL response ex vivo, as well. The resulting CTL or HTL cells, can be used to treat tumors in patients that do not respond to other conventional forms of therapy, or will not respond to a therapeutic vaccine peptide or nucleic acid in accordance with the invention. Ex vivo CTLor HTL responses to a particular antigen are induced by incubating in tissue culture the patient's, or genetically compatible, CTL or HTL precursor cells together with a source of antigen-presenting cells (APC), such as dendritic cells, and the appropriate immunogenic peptide. After an appropriate incubation time (typically about 7-28 days), in which the precursor cells are activated and expanded into effector cells, the cells are infused back into the patient, where they will destroy (CTL) or facilitate destruction (HTL) of their specific target cell a tumor cell). Transfected dendritic cells may also be used as antigen presenting cells.

X.E. Administration of Vaccines for Therapeutic or Prophylactic Purposes Pharmaceutical and vaccine compositions of the invention are typically used to treat and/or prevent a cancer that expresses or overexpresses 24P4C12. In therapeutic applications, peptide and/or nucleic acid compositions are administered to a patient in an amount sufficient to elicit an effective B cell, CTL and/or HTL response to the antigen and to cure or at least partially arrest or slow symptoms and/or complications. An amount adequate to accomplish this is defined as "therapeutically effective dose." Amounts effective for this use will depend on, the particular composition administered, the manner of administration, the stage and severity of the disease being treated, the weight and general state of health of the patient, and the judgment of the prescribing physician.

For pharmaceutical compositions, the immunogenic peptides of the invention, or DNA encoding them, are generally administered to an individual already bearing a tumor that expresses 24P4C12. The peptides or DNA encoding them can be administered individually or as fusions of one or more peptide sequences. Patients can be treated with the immunogenic peptides separately or in conjunction with other treatments, such as surgery, as appropriate.

For therapeutic use, administration should generally begin at the first diagnosis of 24P4C12-associated cancer.

This is followed by boosting doses until at least symptoms are substantially abated and for a period thereafter. The embodiment of the vaccine composition including, but not limited to embodiments such as peptide cocktails, polyepitopic polypeptides, minigenes, or TAA-specific CTLs or pulsed dendritic cells) delivered to the patient may vary according to the stage of the disease or the patient's health status. For example, in a patient with a tumor that expresses 24P4C12, a vaccine comprising 24P4C12-specific CTL may be more efficacious in killing tumor cells in patient with advanced disease than alternative embodiments.

It is generally important to provide an amount of the peptide epitope delivered by a mode of administration sufficient to stimulate effectively a cytotoxic T call response; compositions which stimulate helper T cell responses can also be given in accordance with this embodiment of the invention.

The dosage for an initial therapeutic immunization generally occurs in a unit dosage range where the lower value is about 1, 5, 50, 500, or 1,000 pg and the higher value is about 10,000; 20,000; 30,000; or 50,000 pg. Dosage values for a human typically range from about 500 pg to about 50,000 pg per 70 kilogram patient. Boosting dosages of between about ig to about 50,000 pg of peptide pursuant to a boosting regimen over weeks to months may be administered depending WO 2004/050828 PCT/US2002/038264 upon the patient's response and condition as determined by measuring the specific activity of CTL and HTL obtained from the patient's blood. Administration should continue until at least clinical symptoms or laboratory tests indicate that the neoplasla, has been eliminated or reduced and for a period thereafter. The dosages, routes of administration, and dose schedules are adjusted in accordance with methodologies known in the art.

In certain embodiments, the peptides and compositions of the present invention are employed in serious disease states, that is, life-threatening or potentially life threatening situations. In such cases, as a result of the minimal amounts of extraneous substances and the relative nontoxic nature of the peptides in preferred compositions of the invention, it is possible and may be felt desirable by the treating physician to administer substantial excesses of these peptide compositions relative to these stated dosage amounts.

The vaccine compositions of the invention can also be used purely as prophylactic agents. Generally the dosage for an initial prophylactic immunization generally occurs in a unit dosage range where the lower value is about 1, 5, 50, 500, or 1000 pg and the higher value is about 10,000; 20,000; 30,000; or 50,000 pg. Dosage values for a human typically range from about 500 Ig to about 50,000 pg per 70 kilogram patient. This is followed by boosting dosages of between about lig to about 50,000 Aig of peptide administered at defined intervals from about four weeks to six months after the initial administration of vaccine. The immunogenicity of the vaccine can be assessed by measuring the specific activity of CTL and HTL obtained from a sample of the patient's blood.

The pharmaceutical compositions for therapeutic treatment are intended for parenteral, topical, oral, nasal, intrathecal, or local as a cream or topical ointment) administration. Preferably, the pharmaceutical compositions are administered parentally, intravenously, subcutaneously, intradermally, or intramuscularly. Thus, the invention provides compositions for parenteral administration which comprise a solution of the immunogenic peptides dissolved or suspended in an acceptable carrier, preferably an aqueous carrier.

A variety of aqueous carriers may be used, water, buffered water, 0.8% saline, 0.3% glycine, hyaluronic acid and the like. These compositions may be sterilized by conventional, well-known sterilization techniques, or may be sterile filtered. The resulting aqueous solutions may be packaged for use as is, or lyophilized, the lyophilized preparation being combined with a sterile solution prior to administration.

The compositions may contain pharmaceutically acceptable auxiliary substances as required to approximate physiological conditions, such as pH-adjusting and buffering agents, tonicity adjusting agents, wetting agents, preservatives, and the like, for example, sodium acetate, sodium lactate, sodium chloride, potassium chloride, calcium chloride, sorbitan monolaurate, triethanolamine oleate, etc.

The concentration of peptides of the invention in the pharmaceutical formulations can vary widely, from less than about usually at or at least about 2% to as much as 20% to 50% or more by weight, and will be selected primarily by fluid volumes, viscosities, etc., in accordance with the particular mode of administration selected.

A human unit dose form of a composition is typically included in a pharmaceutical composition that comprises a human unit dose of an acceptable carrier, in one embodiment an aqueous carrier, and is administered in a volume/quantity that is known by those of skill in the art to be used for administration of such compositions to humans (see, Remington's Pharmaceutical Sciences, 17 1 Edition, A. Gennaro, Editor, Mack Publishing Co., Easton, Pennsylvania, 1985). For example a peptide dose for initial immunization can be from about 1 to about 50,000 4g, generally 100-5,000 pg, for a 70 kg patient.

For example, for nucleic acids an initial immunization may be performed using an expression vector in the form of naked nucleic acid administered IM (or SC or ID) in the amounts of 0.5-5 mg at multiple sites. The nucleic acid (0.1 to 1000 pg) can also be administered using a gene gun. Following an incubation period of 3-4 weeks, a booster dose is then administered. The booster can be recombinant fowlpox virus administered at a dose of 5-10 7 to 5x109 pfu.

WO 2004/050828 PCT/US20021038264 For antibodies, a treatment generally involves repeated administration of the anti-24P4C12 antibody preparation, via an acceptable route of administration such as intravenous injection typically at a dose in the range of about 0.1 to about 10 mg/kg body weight. In general, doses in the range of 10-500 mg mAb per week are effective and well tolerated.

Moreover, an initial loading dose of approximately 4 mg/kg patient body weight IV, followed by weekly doses of about 2 mg/kg IV of the anti- 24P4C12 mAb preparation represents an acceptable dosing regimen. As appreciated by those of skill in the art, various factors can influence the ideal dose in a particular case. Such factors include, for example, half life of a composition, the binding affinity of an Ab, the immunogenicity of a substance, the degree of 24P4C12 expression in the patient, the extent of circulating shed 24P4C12 antigen, the desired steady-state concentration level, frequency of treatment, and the influence of chemotherapeutic or other agents used in combination with the treatment method of the invention, as well as the health status of a particular patient. Non-limiting preferred human unit doses are, for example, 500pg 1mg, 1mg 50mg, 50mg 100mg, 100mg 200mg, 200mg 300mg, 400mg 500mg, 500mg 600mg, 600mg 700mg, 700mg 800mg, 800mg 900mg, 900mg lg, or img 700mg. In certain embodiments, the dose is in a range of 2-5 mg/kg body weight, with follow on weekly doses of 1-3 mg/kg; 0.5mg, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10mg/kg body weight followed, in two, three or four weeks by weekly doses; 0.5 -10mg/kg body weight, followed in two, three or four weeks by weekly doses; 225, 250, 275, 300, 325, 350, 375, 400mg m 2 of body area weekly; 1-600mg m 2 of body area weekly; 225-400mg m 2 of body area weekly; these does can be followed by weekly doses for 2, 3, 4, 5, 6, 7, 8, 9, 19, 11, 12 or more weeks.

In one embodiment, human unit dose forms of polynucleotides comprise a suitable dosage range or effective amount that provides any therapeutic effect. As appreciated by one of ordinary skill in the art a therapeutic effect depends on a number of factors, including the sequence of the polynucleotide, molecular weight of the polynucleotide and route of administration. Dosages are generally selected by the physician or other health care professional in accordance with a variety of parameters known in the art, such as severity of symptoms, history of the patient and the like. Generally, for a polynucleotide of about 20 bases, a dosage range may be selected from, for example, an independently selected lower limit such as about 0.1, 0.25, 0.5, 1, 2, 5, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 200, 300, 400 or 500 mg/kg up to an independently selected upper limit, greater than the lower limit, of about 60, 80, 100, 200, 300, 400, 500, 750, 1000, 1500, 2000, 3000, 4000, 5000, 6000, 7000, 8000, 9000 or 10,000 mg/kg. For example, a dose may be about any of the following: 0.1 to 100 mg/kg, 0.1 to 50 mglkg, 0.1 to 25 mg/kg, 0.1 to 10 mg/kg, 1 to 500 mg/kg, 100 to 400 mg/kg, 200 to 300 mg/kg, 1 to 100 mg/kg, 100 to 200 mg/kg, 300 to 400 mg/kg, 400 to 500 mg/kg, 500 to 1000 mg/kg, 500 to 5000 mg/kg, or 500 to 10,000 mg/kg. Generally, parenteral routes of administration may require higher doses of polynuceotide compared to more direct application to the nucleotide to diseased tissue, as do polynucleotides of increasing length.

In one embodiment, human unit dose forms of T-cells comprise a suitable dosage range or effective amount that, provides any therapeutic effect. As appreciated by one of ordinary skill in the art, a therapeutic effect depends on a number of factors. Dosages are generally selected by the physician or other health care professional in accordance with a variety of parameters known in the art, such as severity of symptoms, history of the patient and the like. A dose may be about 104 cells to about 106 cells, about 10 6 cells to about 108 cells, about 108 to about 1011 cells, or about 108 to about 5 x 1010 cells.

A dose may also about 106 cellslm 2 to about 1010 cells/m 2 or about 106 cellslm 2 to about 108 cells/m 2 Proteins(s) of the invention, and/or nucleic acids encoding the protein(s), can also be administered via liposomes, which may also serve to: 1) target the proteins(s) to a particular tissue, such as lymphoid tissue; 2) to target selectively to diseases cells; or, 3) to increase the half-life of the peptide composition. Liposomes include emulsions, foams, micelles, insoluble monolayers, liquid crystals, phospholipid dispersions, lamellar layers and the like. In these preparations, the peptide to be delivered is incorporated as part of a liposome, alone or in conjunction with a molecule which binds to a receptor prevalent among lymphoid cells, such as monoclonal antibodies which bind to the CD45 antigen, or with other therapeutic or immunogenic compositions. Thus, liposomes either filled or decorated with a desired peptide of the invention WO 2004/050828 PCT/US2002/038264 can be directed to the site of lymphoid cells, where the liposomes then deliver the peptide compositions. Liposomes for use in accordance with the invention are formed from standard vesicle-forming lipids, which generally include neutral and negatively charged phospholipids and a sterol, such as cholesterol. The selection of lipids is generally guided by consideration of, liposome size, acid lability and stability of the liposomes in the blood stream. A variety of methods are available for preparing liposomes, as described in, Szoka, et Ann. Rev. Biophys. Bioeng. 9:467 (1980), and U.S.

Patent Nos. 4,235,871, 4,501,728, 4,837,028, and 5,019,369.

For targeting cells of the immune system, a ligand to be incorporated into the liposome can include, e.g., antibodies or fragments thereof specific for cell surface determinants of the desired immune system cells. A liposome suspension containing a peptide may be administered intravenously, locally, topically, etc. in a dose which varies according to, inter alia, the manner of administration, the peptide being delivered, and the stage of the disease being treated.

For solid compositions, conventional nontoxic solid carriers may be used which include, for example, pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium saccharin, talcum, cellulose, glucose, sucrose, magnesium carbonate, and the like. For oral administration, a pharmaceutically acceptable nontoxic composition is formed by incorporating any of the normally employed excipients, such as those carriers previously listed, and generally of active ingredient, that is, one or more peptides of the invention, and more preferably at a concentration of 25%-75%.

For aerosol administration, immunogenic peptides are preferably supplied in finely divided form along with a surfactant and propellant. Typical percentages of peptides are about 0.01%-20% by weight, preferably about The surfactant must, of course, be nontoxic, and preferably soluble in the propellant. Representative of such agents are the esters or partial esters of fatty acids containing from about 6 to 22 carbon atoms, such as caproic, octanoic, lauric, palmitic, stearic, linoleic, linolenic, olesteric and oleic acids with an aliphatic polyhydric alcohol or its cyclic anhydride. Mixed esters, such as mixed or natural glycerides may be employed. The surfactant may constitute about 0.1%-20% by weight of the composition, preferably about 0.25-5%. The balance of the composition is ordinarily propellant. A carrier can also be included, as desired, as with, lecithin for intranasal delivery.

XI.) Diagnostic and Prognostic Embodiments of 24P4C12.

As disclosed herein, 24P4C12 polynudeotides, polypeptides, reactive cytotoxic T cells (CTL), reactive helper T cells (HTL) and anti-polypeptide antbodies are used in well known diagnostic, prognostic and therapeutic assays that examine conditions associated with dysregulated cell growth such as cancer, in particular the cancers listed in Table I (see, both its specific pattern of tissue expression as well as its overexpression in certain cancers as described for example in the Example entitled "Expression analysis of 24P4C12 in normal tissues, and patient specimens").

24P4C12 can be analogized to a prostate associated antigen PSA, the archetypal marker that has been used by medical practitioners for years to identify and monitor the presence of prostate cancer (see, Merrill et J. Urol. 163(2): 503-5120 (2000); Polascik et al., J, Urol. Aug; 162(2):293-306 (1999) and Fortier et al, J. Nat. Cancer Inst. 91(19): 1635- 1640(1999)). A variety of other diagnostic markers are also used in similar contexts including p53 and K-ras (see, e.g., Tulchinsky et al., Int J Mol Med 1999 Jul 4(1):99-102 and Minimoto etal., Cancer Detect Prev 2000;24(1):1-12). Therefore, this disclosure of 24P4C12 polynucleotides and polypeptides (as well as 24P4C12 polynucleotide probes and anti-24P4C12 antibodies used to identify the presence of these molecules) and their properties allows skilled artisans to utilize these molecules in methods that are analogous to those used, for example, in a variety of diagnostic assays directed to examining conditions associated with cancer.

Typical embodiments of diagnostic methods which utilize the 24P4C12 polynudeotides, polypeptides, reactive T cells and antibodies are analogous to those methods from well-established diagnostic assays, which employ, PSA polynucleotides, polypeptides, reactive T cells and antibodies. For example, just as PSA polynucleotides are used as probes WO 2004/050828 PCT/US2002/038264 (for example in Northern analysis, see, Sharief et al., Biochem. Mol. Biol. Int. 33(3):567-74(1994)) and primers (for example in PCR analysis, see, Okegawa et a, J. Urol. 163(4): 1189-1190 (2000)) to observe the presence and/or the level of PSA mRNAs in methods of monitoring PSA overexpression or the metastasis of prostate cancers, the 24P4C12 polynucleotides described herein can be utilized in the same way to detect 24P4C12 overexpression or the metastasis of prostate and other cancers expressing this gene. Alternatively, just as PSA polypeptides are used to generate antibodies specific for PSA which can then be used to observe the presence and/or the level of PSA proteins in methods to monitor PSA protein overexpression (see, Stephan et al., Urology 55(4):560-3 (2000)) or the metastasis of prostate cells (see, Alanen etal., Pathol. Res. Pract. 192(3):233-7 (1996)), the 24P4C12 polypeptides described herein can be utilized to generate antibodies for use in detecting 24P4C12 overexpression or the metastasis of prostate cells and cells of other cancers expressing this gene.

Specifically, because metastases involves the movement of cancer cells from an organ of origin (such as the lung or prostate gland etc.) to a different area of the body (such as a lymph node), assays which examine a biological sample for the presence of cells expressing 24P4C12 polynucleotides and/or polypeptides can be used to provide evidence of metastasis. For example, when a biological sample from tissue that does not normally contain 24P4C12-expressing cells (lymph node) is found to contain 24P4C12-expressing cells such as the 24P4C12 expression seen in LAPC4 and LAPC9, xenografts isolated from lymph node and bone metastasis, respectively, this finding is indicative of metastasis.

Alternatively 24P4C12 polynucleotides andfor polypeptides can be used to provide evidence of cancer, for example, when cells in a biological sample that do not normally express 24P4C12 or express 24P4C12 at a different level are found to express 24P4C12 or have an increased expression of 24P4C12 (see, the 24P4C12 expression in the cancers listed in Table I and in patient samples etc. shown in the accompanying Figures). In such assays, artisans may further wish to generate supplementary evidence of metastasis by testing the biological sample for the presence of a second tissue restricted marker(in addition to 24P4C12) such as PSA, PSCA etc. (see, Alanen et al, Pathol. Res. Pract. 192(3): 233- 237 (1996)).

Just as PSA polynucleotide fragments and polynucleotide variants are employed by skilled artisans for use in methods of monitoring PSA, 24P4C12 polynucleotide fragments and polynucleotide variants are used in an analogous manner. In particular, typical PSA polynucleotides used in methods of monitoring PSA are probes or primers which consist of fragments of the PSA cDNA sequence. Illustrating this, primers used to PCR amplify a PSA polynucleotide must include less than the whole PSA sequence to function in the polymerase chain reaction. In the context of such PCR reactions, skilled artisans generally create a variety of different polynucleotide fragments that can be used as primers in order to amplify different portions of a polynucleotide of interest or to optimize amplification reactions (see, Caetano-Anolles, G.

Biotechniques 25(3): 472-476, 478480 (1998); Robertson et al., Methods Mol. Biol. 98:121-154 (1998)). An additional illustration of the use of such fragments is provided in the Example entitled "Expression analysis of 24P4C12 in normal tissues, and patient specimens," where a 24P4C12 polynuleotide fragment is used as a probe to show the expression of 24P4C12 RNAs in cancer cells. In addition, variant polynucleotide sequences are typically used as primers and probes for the corresponding mRNAs in PCR and Northern analyses (see, Sawai et al, Fetal Diagn. Ther. 1996 Nov-Dec 11(6):407-13 and Current Protocols In Molecular Biology, Volume 2, Unit 2, Frederick M. Ausubel etal. eds., 1995)).

Polynucleotide fragments and variants are useful in this context where they are capable of binding to a target polynucleotide sequence a 24P4C12 polynucleotide shown in Figure 2 or variant thereof) under conditions of high stringency.

Furthermore, PSA polypeptides which contain an epitope that can be recognized by an antibody or T cell that specifically binds to that epitope are used in methods of monitoring PSA. 24P4C12 polypeptide fragments and polypeptide analogs or variants can also be used in an analogous manner. This practice of using polypeptide fragments or polypeptide variants to generate antibodies (such as anti-PSA antibodies or T cells) is typical in the art with a wide variety of systems WO 2004/050828 PCT/US2002/038264 such as fusion proteins being used by practitioners (see, Current Protocols In Molecular Biology, Volume 2, Unit 16, Frederick M. Ausubel et a. eds., 1995). In this context, each epitope(s) functions to provide the architecture with which an antibody or T ce!l is reactive. Typically, skilled artisans create a variety of different polypeptide fragments that can be used in order to generate immune responses specific for different portions of a polypeptide of interest (see, U.S. Patent No.

5,840,501 and U.S. Patent No. 5,939,533). For example it may be preferable to utilize a polypeptide comprising one of the 24P4C12 biological motifs discussed herein or a motif-bearing subsequence which is readily identified by one of skill in the art based on motifs available in the art. Polypeptide fragments, variants or analogs are typically useful in this context as long as they comprise an epitope capable of generating an antibody or T cell specific for a target polypeptide sequence a 24P4C12 polypeptide shown in Figure 3).

As shown herein, the 24P4C12 polynucleotides and polypeptides (as well as the 24P4C12 polynucleotide probes and anti-24P4C12 antibodies or T cells used to identify the presence of these molecules) exhibit specific properties that make them useful in diagnosing cancers such as those listed in Table I. Diagnostic assays that measure the presence of 24P4C12 gene products, in order to evaluate the presence or onset of a disease condition described herein, such as prostate cancer, are used to identify patients for preventive measures or further monitoring, as has been done so successfully with PSA. Moreover, these materials satisfy a need in the art for molecules having similar or complementary characteristics to PSA in situations where, for example, a definite diagnosis of metastasis of prostatic origin cannot be made on the basis of a test for PSA alone (see, Alanen et Pathol. Res. Pracl. 192(3): 233-237 (1996)), and consequently, materials such as 24P4C12 polynucleotides and polypeptides (as well as the 24P4C12 polynucleotide probes and anti- 24P4C12 antibodies used to identify the presence of these molecules) need to be employed to confirm a metastases of prostatic origin.

Finally, in addition to their use in diagnostic assays, the 24P4C12 polynucleotides disclosed herein have a number of other utilities such as their use in the identification of oncogenetic associated chromosomal abnormalities in the chromosomal region to which the 24P4C12 gene maps (see the Example entitled "Chromosomal Mapping of 24P4C12" below). Moreover, in addition to their use in diagnostic assays, the 24P4C12-related proteins and polynucleotides disclosed herein have other utilities such as their use in the forensic analysis of tissues of unknown origin (see, Takahama K Forensic Sci Int 1996 Jun 28;80(1-2): 63-9).

Additionally, 24P4C12-related proteins or polynucleolides of the invention can be used to treat a pathologic condition characterized by the over-expression of 24P4C12. For example, the amino acid or nucleic acid sequence of Figure 2 or Figure 3, or fragments of either, can be used to generate an immune response to a 24P4C12 antigen. Antibodies or other molecules that react with 24P4C12 can be used to modulate the function of this molecule, and thereby provide a therapeutic benefit.

XII.) Inhibition of 24P4C12 Protein Function The invention includes various methods and compositions for inhibiting the binding of 24P4C12 to its binding partner or its association with other protein(s) as well as methods for inhibiting 24P4C12 function.

XII.A.) Inhibition of 24P4C12 With Intracellular Antibodies In one approach, a recombinant vector that encodes single chain antibodies that specifically bind to 24P4C12 are introduced into 24P4C12 expressing cells via gene transfer technologies. Accordingly, the encoded single chain anti- 24P4C12 antibody is expressed intracellularly, binds to 24P4C12 protein, and thereby inhibits its function. Methods for engineering such intracellular single chain antibodies are well known. Such intracellular antibodies, also known as "intrabodies", are specifically targeted to a particular compartment within the cell, providing control over where the inhibitory WO 2004/050828 PCT/US2002/038264 activity of the treatment is focused. This technology has been successfully applied in the art (for review, see Richardson and Marasco, 1995, TIBTECH vol. 13). Intrabodies have been shown to virtually eliminate the expression of otherwise abundant cell surface receptors (see, Richardson et al., 1995, Proc. Natl. Acad. Sci USA 92: 3137-3141; Beerli et al, 1994, J.

Biol. Chem. 289: 23931-23936; Deshane et 1994, Gene Ther. 1: 332-337).

Single chain antibodies comprise the variable domains of the heavy and light chain joined by a flexible linker polypeptide, and are expressed as a single polypeptide. Optionally, single chain antibodies are expressed as a single chain variable region fragment joined to the light chain constant region. Well-known intracellular trafficking signals are engineered into recombinant polynucleotide vectors encoding such single chain antibodies in order to target precisely the intrabody to the desired intracellular compartment. For example, intrabodies targeted to the endoplasmic reticulum (ER) are engineered to incorporate a leader peptide and, optionally, a C-terminal ER retention signal, such as the KDEL amino acid motif.

Intrabodies intended to exert activity in the nucleus are engineered to include a nuclear localization signal. Lipid moieties are joined to intrabodies in order to tether the intrabody to the cytosolic side of the plasma membrane. Intrabodies can also be targeted to exert function in the cytosol. For example, cytosolic intrabodies are used to sequester factors within the cytosol, thereby preventing them from being transported to their natural cellular destination.

In one embodiment, intrabodies are used to capture 24P4C12 in the nucleus, thereby preventing its activity within the nucleus. Nuclear targeting signals are engineered into such 24P4C12 intrabodies in order to achieve the desired targeting. Such 24P4C12 intrabodies are designed to bind specifically to a particular 24P4C12 domain. In another embodiment, cytosolic intrabodies that specifically bind to a 24P4C12 protein are used to prevent 24P4C12 from gaining access to the nucleus, thereby preventing it from exerting any biological activity within the nucleus preventing 24P4C12 from forming transcription complexes with other factors).

In order to specifically direct the expression of such intrabodies to particular cells, the transcription of the intrabody is placed under the regulatory control of an appropriate tumor-specific promoter andlor enhancer. In order to target intrabody expression specifically to prostate, for example, the PSA promoter and/or promoterlenhancer can be utilized (See, for example, U.S. Patent No. 5,919,652 issued 6 July 1999).

XII.B.) Inhibition of 24P4C12 with Recombinant Proteins In another approach,'recombinant molecules bind to 24P4C12 and thereby inhibit 24P4C12 function. For example, these recombinant molecules prevent or inhibit 24P4C12 from accessing/binding to its binding partner(s) or associating with other protein(s). Such recombinant molecules can, for example, contain the reactive part(s) of a 24P4C12 specific antibody molecule. In a particular embodiment, the 24P4C12 binding domain of a 24P4C12 binding partner is engineered into a dimeric fusion protein, whereby the fusion protein comprises two 24P4C12 ligand binding domains linked to the Fc portion of a human IgG, such as human IgG1. Such IgG portion can contain, for example, the CH 2 and CH3 domains and the hinge region, but not the CH1 domain. Such dimeric fusion proteins are administered in soluble form to patients suffering from a cancer associated with the expression of 24P4C12, whereby the dimeric fusion protein specifically binds to 24P4C12 and blocks 24P4C12 interaction with a binding partner. Such dimeric fusion proteins are further combined into multimeric proteins using known antibody linking technologies.

XII.C.) Inhibition of 24P4C12 Transcription or Translation The present invention also comprises various methods and compositions for inhibiting the transcription of the 24P4C12 gene. Similarly, the invention also provides methods and compositions for inhibiting the translation of 24P4C12 mRNA into protein.

WO 2004/050828 PCT/US20021038264 In one approach, a method of inhibiting the transcription of the 24P4C12 gene comprises contacting the 24P4C12 gene with a 24P4C12 antisense polynucleotide. In another approach, a method of inhibiting 24P4C12 mRNA translation comprises contacting a 24P4C12 mRNA with an antisense polynucleotide. In another approach, a 24P4C12 specific ribozyme is used to cleave a24P4C12 message, thereby inhibiting translation. Such antisense and ribozyme based methods can also be directed to the regulatory regions of the 24P4C12 gene, such as 24P4C12 promoter and/or enhancer elements. Similarly, proteins capable of inhibiting a 24P4C12 gene transcription factor are used to inhibit 24P4C12 mRNA transcription. The various polynucleotides and compositions useful in the aforementioned methods have been described above. The use of antisense and ribozyme molecules to inhbit transcription and translation is well known in the art.

Other factors that inhibit the transcription of 24P4C12 by interfering with 24P4C12 transcriptional activation are also useful to treat cancers expressing 24P4C12. Similarly, factors that interfere with 24P4C12 processing are useful to treat cancers that express 24P4C12. Cancer treatment methods utilizing such factors are also within the scope of the invention.

XII.D.) General Considerations for Therapeutic Strategies Gene transfer and gene therapy technologies can be used to deliver therapeutic polynucleotide molecules to tumor cells synthesizing 24P4C12 antisense, ribozyme, polynucleotides encoding intrabodies and other 24P4C12 inhibitory molecules).

A number of gene therapy approaches are known in the art. Recombinant vectors encoding 24P4C12 antisense polynucleotides, ribozymes, factors capable of interfering with 24P4C12 transcription, and so forth, can be delivered to target tumor cells using such gene therapy approaches.

The above therapeutic approaches can be combined with any one of a wide variety of surgical, chemotherapy or radiation therapy regimens. The therapeutic approaches of the invention can enable the use of reduced dosages of chemotherapy (or other therapies) andlor less frequent administration, an advantage for all patients and particularly for those that do not tolerate the toxicity of the chemotherapeutic agent well.

The anti-tumor activity of a particular composition antisense, ribozyme, intrabody), or a combination of such compositions, can be evaluated using various in vitro and in vivo assay systems. In vitro assays that evaluate therapeutic activity include cell growth assays, soft agar assays and other assays ndicative of tumor promoting activity, binding assays capable of determining the extent to which a therapeutic composition will inhibit the binding of 24P4C12 to a binding partner, etc.

In vivo, the effect of a 24P4C12 therapeutic composition can be evaluated in a suitable animal model. For example, xenogenic prostate cancer models can be used, wherein human prostate cancer explants or passaged xenograft tissues are introduced into immune compromised animals, such as nude or SCID mice (Klein etal., 1997, Nature Medicine 3:402-408). For example, PCT Patent Application W098116628 and U.S. Patent 6,107,540 describe various xenograft models of human prostate cancer capable of recapitulating the development of primary tumors, micrometastasis, and the formation of osteoblastic metastases characteristic of late stage disease. Efficacy can be predicted using assays that measure inhibition of tumor formation, tumor regression or metastasis, and the like.

In vivo assays that evaluate the promotion of apoptosis are useful in evaluating therapeutic compositions. In one embodiment, xenografts from tumor bearing mice treated with the therapeutic composition can be examined for the presence of apoptotic foci and compared to untreated control xenograft-bearing mice. The extent to which apoptotic foci are found in the tumors of the treated mice provides an indication of the therapeutic efficacy of the composition.

The therapeutic compositions used in the practice of the foregoing methods can be formulated into pharmaceutical compositions comprising a carrier suitable for the desired delivery method. Suitable carriers include any material that when combined with the therapeutic composition retains the anti-tumor function of the therapeutic composition and is generally non-reactive with the patient's immune system. Examples include, but are not limited to, any of a number of standard WO 2004/050828 PCT/US2002/038264 pharmaceutical carriers such as sterile phosphate buffered saline solutions, bacteriostatic water, and the like (see, generally, Remington's Pharmaceutical Sciences 16" Edition, A. Osal., Ed., 1980).

Therapeutic formulations can be solubilized and administered via any route capable of delivering the therapeutic composition to the tumor site. Polentially effective routes of administration include, but are not limited to, intravenous, parenteral, intraperitoneal, intramuscular, intratumor, intradermal, intraorgan, orthotopic, and the like. A preferred formulation for intravenous injection comprises the therapeutic composition in a solution of preserved bacteriostatic water, sterile unpreserved water, and/or diluted in polyvinylchloride or polyethylene bags containing 0.9% sterile Sodium Chloride for Injection, USP. Therapeutic protein preparations can be lyophilized and stored as sterile powders, preferably under vacuum, and then reconstituted in bacteriostatic water (containing for example, benzyl alcohol preservative) or in sterile water prior to injection.

Dosages and administration protocols for the treatment of cancers using the foregoing methods will vary with the method and the target cancer, and will generally depend on a number of other factors appreciated in the art.

XIII. Identification, Characterization and Use of Modulators of 24P4C12 Methods to Identify and Use Modulators In one embodiment, screening is performed to identify modulators that induce or suppress a particular expression profile, suppress or induce specific pathways, preferably generating the associated phenotype thereby. In another embodiment, having identified differentially expressed genes important in a particular state; screens are performed to identify modulators that alter expression of individual genes, either increase or decrease. In another embodiment, screening is performed to identify modulators that alter a biological function of the expression product of a differentially expressed gene.

Again, having identified the importance of a gene in a particular state, screens are performed to identify agents that bind and/or modulate the biological activity of the gene product.

In addition, screens are done for genes that are induced in response to a candidate agent. After identifying a modulator (one that suppresses a cancer expression pattern leading to a normal expression pattern, or a modulator of a cancer gene that leads to expression of the gene as in normal tissue) a screen is performed to identify genes that are specifically modulated in response to the agent. Comparing expression profiles between normal tissue and agent-treated cancer tissue reveals genes that are not expressed in normal tissue or cancer tissue, but are expressed in agent treated tissue, and vice versa. These agent-specific sequences are identified and used by methods described herein for cancer genes or proteins. In particular these sequences and the proteins they encode are used in marking or identifying agenttreated cells. In addition, antibodies are raised against the agent-induced proteins and used to target novel therapeutics to the treated cancer tissue sample.

Modulator-related Identification and Screening Assays: Gene Expression-related Assays Proteins, nucleic acids and antibodies of the invention are used in screening assays. The cancer-associated proteins, antibodies, nucleic acids, modified proteins and cells containing these sequences are used in screening assays, such as evaluating the effect of drug candidates on a 'gene expression profile," expression profile of polypeptides or alteration of biological function. In one embodiment, the expression profiles are used, preferably in conjunction with high throughput screening techniques to allow monitoring for expression profile genes after treatment with a candidate agent Davis, GF, et al, J Biol Screen 7:69 (2002); Zlokarnik, etal., Science 279:84-8 (1998); Held, Genome Res 6:986- 94,1996).

WO 2004/050828 PCT/US2002/038264 The cancer proteins, antibodies, nucleic acids, modified proteins and cells containing the native or modified cancer proteins or genes are used in screening assays. That is, the present invention comprises methods for screening for compositions which modulate the cancer phenotype or a physiological function of a cancer protein of the invention. This is done on a gene itself or by evaluating the effect of drug candidates on a "gene expression profile" or biological function. In one embodiment, expression profiles are used, preferably in conjunction with high throughput screening techniques to allow monitoring after treatment with a candidate agent, see Zlokamik, supra.

A variety of assays are executed directed to the genes and proteins of the invention. Assays are run on an individual nucleic acid or protein level. That is, having identified a particular gene as up regulated in cancer, test compounds are screened for the ability to modulate gene expression or for binding to the cancer protein of the invention. "Modulation" in this context includes an increase or a decrease in gene expression. The preferred amount of modulation will depend on the original change of the gene expression in normal versus tissue undergoing cancer, with changes of at least 10%, preferably more preferably 100-300%, and in some embodiments 300-1000% or greater. Thus, if a gene exhibits a 4-fold increase in cancer tissue compared to normal tissue, a decrease of about four-fold is often desired; similarly, a decrease in cancer tissue compared to normal tissue a target value of a 10-fold increase in expression by the test compound is often desired. Modulators that exacerbate the type of gene expression seen in cancer are also useful, as an upregulated target in further analyses.

The amount of gene expression is monitored using nucleic acid probes and the quantification of gene expression levels, or, alternatively, a gene product itself is monitored, through the use of antibodies to the cancer protein and standard immunoassays. Proteomics and separation techniques also allow for quantification of expression.

Expression Monitoring to Identify Compounds that Modify Gene Expression In one embodiment, gene expression monitoring, an expression profile, is monitored simultaneously for a number of entities. Such profiles will typically involve one or more of the genes of Figure 2. In this embodiment, cancer nucleic acid probes are attached to biochips to detect and quantify cancer sequences in a particular cell. Alternatively, PCR can be used. Thus, a series, wells of a microtiter plate, can be used with dispensed primers in desired wells. A PCR reaction can then be performed and analyzed for each well.

Expression monitoring is performed to identify compounds that modify the expression of one or more cancerassociated sequences, a polynucleotide sequence set out in Figure 2. Generally, a test modulator is added to the cells prior to analysis. Moreover, screens are also provided to identify agents that modulate cancer, modulate cancer proteins of the invention, bind to a cancer protein of the invention, or interfere with the binding of a cancer protein of the invention and an antibody or other binding partner.

In one embodiment, high throughput screening methods involve providing a library containing a large number of potential therapeutic compounds (candidate compounds). Such "combinatorial chemical libraries' are then screened in one or more assays to identify those library members (particular chemical species or subclasses) that display a desired characteristic activity. The compounds thus identified can serve as conventional "lead compounds," as compounds for screening, or as therapeutics.

In certain embodiments, combinatorial libraries of potential modulators are screened for an ability to bind to a cancer polypeptide or to modulate activity. Conventionally, new chemical entities with useful properties are generated by identifying a chemical compound (called a "lead compound") with some desirable property or activity, inhibiting activity, creating variants of the lead compound, and evaluating the property and activity of those variant compounds. Often, high throughput screening (HTS) methods are employed for such an analysis.

WO 2004/050828 PCT/US2002/038264 As noted above, gene expression monitoring is conveniently used to test candidate modulators protein, nucleic acid or small molecule). After the candidate agent has been added and the cells allowed to incubate for a period, the sample containing a target sequence to be analyzed is, added to a biochip.

If required, the target sequence is prepared using known techniques. For example, a sample is treated to lyse the cells, using known lysis buffers, electroporation, etc., with purification and/or amplification such as PCR performed as appropriate. For example, an in vitro transcription with labels covalently attached to the nucleotides is performed. Generally, the nucleic acids are labeled with biotin-FITC or PE, or with cy3 or The target sequence can be labeled with, a fluorescent, a chemiluminescent, a chemical, or a radioactive signal, to provide a means of detecting the target sequence's specific binding to a probe. The label also can be an enzyme, such as alkaline phosphatase or horseradish peroxidase, which when provided with an appropriate substrate produces a product that is detected. Alternatively, the label is a labeled compound or small molecule, such as an enzyme inhibitor, that binds but is not catalyzed or altered by the enzyme. The label also can be a moiety or compound, such as, an epitope tag or biotin which specifically binds to streptavidin. For the example of biotin, the streptavidin is labeled as described above, thereby, providing a detectable signal for the bound target sequence. Unbound labeled streptavidin is typically removed prior to analysis.

As will be appreciated by those in the art, these assays can be direct hybridization assays or can comprise "sandwich assays", which include the use of multiple probes, as is generally outlined in U.S. Patent Nos. 5, 681,702; 5,597,909; 5,545,730; 5,594,117; 5,591,584; 5,571,670; 5,580,731; 5,571,670; 5,591,584; 5,624,802; 5,635,352; 5,594,118; 5,359,100; 5,124, 246; and 5,681,697. In this embodiment, in general, the target nucleic acid is prepared as outlined above, and then added to the biochip comprising a plurality of nucleic acid probes, under conditions that allow the formation of a hybridization complex.

A variety of hybridization conditions are used in the present invention, including high, moderate and low stringency conditions as outlined above. The assays are generally run under stringency conditions which allow formation of the label probe hybridization complex only in the presence of target. Stringency can be controlled by altering a step parameter that is a thermodynamic variable, including, but not limited to, temperature, formamide concentration, salt concentration, chaotropic salt concentration pH, organic solvent concentration, etc. These parameters may also be used to control non-specific binding, as is generally outlined in U.S. Patent No. 5,681,697. Thus, it can be desirable to perform certain steps at higher stringency conditions to reduce non-specific binding.

The reactions outlined herein can be accomplished in a variety of ways. Components of the reaction can be added simultaneously, or sequentially, in different orders, with preferred embodiments outlined below. In addition, the reaction may include a variety of other reagents. These include salts, buffers, neutral proteins, e.g. albumin, detergents, etc. which can be used to facilitate optimal hybridization and detection, and/or reduce nonspecific or background interactions. Reagents that otherwise improve the efficiency of the assay, such as protease inhibitors, nuclease inhibitors, anti-microbial agents, etc., may also be used as appropriate, depending on the sample preparation methods and purity of the target The assay data are analyzed to determine the expression levels of Individual genes, and changes in expression levels as between states, forming a gene expression profile.

Biological Activity-related Assays The invention provides methods identify or screen for a compound that modulates the activity of a cancer-related gene or protein of the invention. The methods comprise adding a test compound, as defined above, to a cell comprising a cancer protein of the invention. The cells contain a recombinant nucleic acid that encodes a cancer protein of the invention.

In another embodiment, a library of candidate agents is tested on a plurality of cells.

WO 2004/050828 PCT/US2002/038264 In one aspect, the assays are evaluated in the presence or absence or previous or subsequent exposure of physiological signals, e.g. hormones, antibodies, peptides, antigens, cytokines, growth factors, action potentials, pharmacological agents including chemotherapeutics, radiation, carcinogenics, or other cells cell-cell contacts). In another example, the determinations are made at different stages of the cell cycle process. In this way, compounds that modulate genes or proteins of the invention are identified. Compounds with pharmacological activity are able to enhance or interfere with the activity of the cancer protein of the invention. Once identified, similar structures are evaluated to identify critical structural features of the compound.

In one embodiment, a method of modulating inhibiting) cancer cell division is provided; the method comprises administration of a cancer modulator. In another embodiment, a method of modulating inhibiting) cancer is provided; the method comprises administration of a cancer modulator. In a further embodiment, methods of treating cells or individuals with cancer are provided; the method comprises administration of a cancer modulator.

In one embodiment, a method for modulating the status of a cell that expresses a gene of the invention is provided.

As used herein status comprises such art-accepted parameters such as growth, proliferation, survival, function, apoptosis, senescence, location, enzymatic activity, signal transduction, etc. of a cell, In one embodiment, a cancer inhibitor is an antibody as discussed above. In another embodiment, the cancer inhibitor is an antisense molecule. A variety of cell growth, proliferation, and metastasis assays are known to those of skill in the art, as described herein.

High Throughput Screening to Identify Modulators The assays to identify suitable modulators are amenable to high throughput screening. Preferred assays thus detect enhancement or inhibition of cancer gene transcription, inhibition or enhancement of polypeptide expression, and inhibition or enhancement of polypeptide acivity.

In one embodiment, modulators evaluated in high throughput screening methods are proteins, often naturally occurring proteins or fragments of naturally occurring proteins. Thus, cellular extracts containing proteins, or random or directed digests of proteinaceous cellular extracts, are used. In this way, libraries of proteins are made for screening in the methods of the invention. Particularly preferred in this embodiment are libraries of bacterial, fungal, viral, and mammalian proteins, with the latter being preferred, and human proteins being especially preferred. Particularly useful test compound will be directed to the class of proteins to which the target belongs, substrates for enzymes, or ligands and receptors.

Use of Soft Aqar Growth and Colony Formation to Identify and Characterize Modulators Normal cells require a solid substrate to attach and grow. When cells are transformed, they lose this phenotype and grow detached from the substrate. For example, transformed cells can grow in stirred suspension culture or suspended in semi-solid media, such as semi-solid or soft agar. The transformed cells, when transfected with tumor suppressor genes, can regenerate normal phenotype and once again require a solid substrate to attach to and grow. Soft agar growth or colony formation in assays are used to identify modulators of cancer sequences, which when expressed in host cells, inhibit abnormal cellular proliferation and transformation. A modulator reduces or eliminates the host cells' ability to grow suspended in solid or semisolid media, such as agar.

Techniques for soft agar growth or colony formation in suspension assays are described in Freshney, Culture of Animal Cells a Manual of Basic Technique (3rd ed., 1994). See also, the methods section of Garkavtsev et al. (1996), supra.

Evaluation of Contact Inhibition and Growth Density Limitation to Identify and Characterize Modulators Normal cells typically grow in a flat and organized pattern in cell culture until they touch other cells. When the cells touch one another, they are contact inhibited and stop growing. Transformed cells, however, are not contact inhibited and continue to grow to high densities in disorganized foci. Thus, transformed cells grow to a higher saturation density than corresponding normal cells. This is detected morphologically by the formation of a disoriented monolayer of cells or cells in WO 2004/050828 PCT/US2002/038264 foci. Alternatively, labeling index with 3 H)-thymidine at saturation density is used to measure density limitation of growth, similarly an MTT or Alamar blue assay will reveal proliferation capacity of cells and the the ability of modulators to affect same. See Freshney (1994), supra. Transformed cells, when transfected with tumor suppressor genes, can regenerate a normal phenotype and become contact inhibited and would grow to a lower density.

In this assay, labeling index with 3 H)-thymidine at saturation density is a preferred method of measuring density limitation of growth. Transformed host cells are transfected with a cancer-associated sequence and are grown for 24 hours at saturation density in non-limiting medium conditions. The percentage of cells labeling with (1H)-thymidine is determined by incorporated cpm.

Contact independent growth is used to identify modulators of cancer sequences, which had led to abnormal cellular proliferation and transformation. A modulator reduces or eliminates contact independent growth, and returns the cells to a normal phenotype.

Evaluation of Growth Factor or Serum Dependence to Identify and Characterize Modulators Transformed cells have lower serum dependence than their normal counterparts (see, Temin, J. Natl. Cancer Inst. 37:167-175 (1966); Eagle et al., J. Exp. Med 131:836-879 (1970)); Freshney, supra. This is in part due to release of various growth factors by the transformed cells. The degree of growth factor or serum dependence of transformed host cells can be compared with that of control. For example, growth factor or serum dependence of a cell is monitored in methods to identify and characterize compounds that modulate cancer-associated sequences of the invention.

Use of Tumor-specific Marker Levels to Identify and Characterize Modulators Tumor cells release an increased amount of certain factors (hereinafter "tumor specific markers") than their normal counterparts. For example, plasminogen activator (PA) is released from human glioma at a higher level than from normal brain cells (see, Gullino, Angiogenesis, Tumor Vascularization, and Potential Interference with Tumor Growth, in Biological Responses in Cancer, pp. 178-184 (Mihich 1985)). Similarly, Tumor Angiogenesis Factor (TAF) is released at a higher level in tumor cells than their normal counterparts. See, Folkman, Angiogenesis and Cancer, Sem Cancer Biol. (1992)), while bFGF is released from endothelial tumors (Ensoli, B et al).

Various techniques which measure the release of these factors are described in Freshney (1994), supra. Also, see, Unkless etal., J. Biol. Chem. 249:4295-4305 (1974); Strickland Beers, J. Biol. Chem. 251:5694-5702 (1976); Whur et al., Br. J. Cancer 42:305 312 (1980); Gullino, Angiogenesis, Tumor Vascularization, and Potential Interference with Tumor Growth, in Biological Responses in Cancer, pp. 178-184 (Mihich 1985); Freshney, Anticancer Res. 5:111-130 (1985).

For example, tumor specific marker levels are monitored in methods to identify and characterize compounds that modulate cancer-associated sequences of the invention.

Invasiveness into Matriael to Identify and Characterize Modulators The degree of invasiveness into Matrigel or an extracellular matrix constituent can be used as an assay to identify and characterize compounds that modulate cancer associated sequences. Tumor cells exhibit a positive correlation between malignancy and invasiveness of cells into Matrigel or some other extracellular matrix constituent. In this assay, tumorigenic cells are typically used as host cells. Expression of a tumor suppressor gene in these host cells would decrease invasiveness of the host cells. Techniques described in Cancer Res. 1999; 59:6010; Freshney (1994), supra, can be used.

Brief y, the level of invasion of host cells is measured by using filters coated with Matrigel or some other extracellular matrix constituent Penetration into the gel, or through to the distal side of the filter, is rated as invasiveness, and rated histologically by number of cells and distance moved, or by prelabeling the cells with 1251 and counting the radioactivity on the distal side of the filter or bottom of the dish. See, Freshney (1984), supra.

Evaluation of Tumor Growth In Vivo to Identify and Characterize Modulators WO 2004/050828 PCT/US2002/038264 Effects of cancer-associated sequences on cell growth are tested in transgenic or immune-suppressed organisms.

Transgenic organisms are prepared in a variety of art-accepted ways. For example, knock-out transgenic organisms, e.g., mammals such as mice, are made, in which a cancer gene is disrupted or in which a cancer gene is inserted. Knock-out transgenic mice are made by insertion of a marker gene or other heterologous gene into the endogenous cancer gene site in the mouse genome via homologous recombination. Such mice can also be made by substituting the endogenous cancer gene with a mutated version of the cancer gene, or by mutating the endogenous cancer gene, by exposure to carcinogens.

To prepare transgenic chimeric animals, mice a DNA construct is introduced into the nuclei of embryonic stem cells. Cells containing the newly engineered genetic lesion are injected into a host mouse embryo, which is reimplanted into a recipient female. Some of these embryos develop into chimeric mice that possess germ cells some of which are derived from the mutant cell line. Therefore, by breeding the chimeric mice it is possible to obtain a new line of mice containing the introduced genetic lesion (see, Capecchi etal., Science 244:1288 (1989)). Chimeric mice can be derived according to US Patent 6,365,797, issued 2 April 2002; US Patent 6,107,540 issued 22 August 2000; Hogan et al., Manipulating the Mouse Embryo: A laboratory Manual, Cold Spring Harbor Laboratory (1988) and Teratocarcinomas and Embryonic Stem Cells: A Practical Approach, Robertson, ed., IRL Press, Washington, (1987).

Alternatively, various immune-suppressed or immune-deficient host animals can be used. For example, a genetically athymic "nude" mouse (see, Giovanella et al., J. Natl. Cancer Inst. 52:921 (1974)), a SCID mouse, a thymectomized mouse, or an irradiated mouse (see, Bradley et al., Br. J. Cancer 38:263 (1978); Selby et al., Br. J.

Cancer 41:52 (1980)) can be used as a host. Transplantable tumor cells (typically about 106 cells) injected into isogenic hosts produce invasive tumors in a high proportion of cases, while normal cells of similar origin will not. In hosts which developed invasive tumors, cells expressing cancer-associated sequences are injected subcutaneously or orthotopically.

Mice are then separated into groups including control groups and treated experimental groups) e.g. treated with a modulator). After a suitable length of time, preferably 4-8 weeks, lumor growth is measured by volume or by its two largest dimensions, or weight) and compared to the control. Tumors that have statistically significant reduction (using, e.g., Student's T test) are said to have inhibited growth.

In Vitro Assays to Identify and Characterize Modulators Assays to identify compounds with modulating activity can be performed in vitro. For example, a cancer polypeptide is first contacted with a potential modulator and incubated for a suitable amount of time, from 0.5 to 48 hours. In one embodiment, the cancer polypeptide levels are determined in vitro by measuring the level of protein or mRNA.

The level of protein is measured using immunoassays such as Western blotting, ELISA and the like with an antibody that selectively binds to the cancer polypeptide or a fragment thereof. For measurement of mRNA, amplification, using PCR, LCR, or hybridization assays, e. g, Northern hybridization, RNAse protection, dot blotting, are preferred. The level of protein or mRNA is detected using drectly or indirectly labeled detection agents, fluorescently or radioactively labeled nucleic acids, radioactively or enzymatically labeled antibodies, and the like, as described herein.

Alternatively, a reporter gene system can be devised using a cancer protein promoter operably linked to a reporter gene such as luciferase, green fluorescent protein, CAT, or P-gal. The reporter construct is typically transfected into a cell.

After treatment with a potential modulator, the amount of reporter gene transcription, translation, or activity is measured according to standard techniques known to those of skill in the art (Davis GF, supra; Gonzalez, J. Negulescu, P. Curr.

Opin. Biotechnol. 1998: 9:624).

WO 2004/050828 PCT/US20021038264 As outlined above, in vitro screens are done on individual genes and gene products. That is, having identified a particular differentially expressed gene as important in a partcular state, screening of modulators of the expression of the gene or the gene product itself is performed.

In one embodiment, screening for modulators of expression of specific gene(s) is performed. Typically, the expression of only one or a few genes is evaluated. In another embodiment, screens are designed to first find compounds that bind to differentially expressed proteins. These compounds are then evaluated for the ability to modulate differentially expressed activity. Moreover, once initial candidate compounds are identified, variants can be further screened to better evaluate structure activity relationships.

Binding Assays to Identify and Characterize Modulators In binding assays in accordance with the invention, a purified or isolated gene product of the invention is generally used. For example, antibodies are generated to a protein of the invention, and immunoassays are run to determine the amount and/or location of protein. Alternatively, cells comprising the cancer proteins are used in the assays.

Thus, the methods comprise combining a cancer protein of the invention and a candidate compound such as a ligand, and determining the binding of the compound to the cancer protein of the invention. Preferred embodiments utilize the human cancer protein; animal models of human disease of can also be developed and used. Also, other analogous mammalian proteins also can be used as appreciated by those of skill in the art. Moreover, in some embodiments variant or derivative cancer proteins are used.

Generally, the cancer protein of the invention, or the ligand, is non-diffusibly bound to an insoluble support. The support can, be one having isolated sample receiving areas (a microtiter plate, an array, etc.). The insoluble supports can be made of any composition to which the compositions can be bound, is readily separated from soluble material, and is otherwise compatible with the overall method of screening. The surface of such supports can be solid or porous and of any convenient shape.

Examples of suitable insoluble supports include microtiter plates, arrays, membranes and beads. These are typically made of glass, plastic polystyrene), polysaccharide, nylon, nitrocellulose, or Teflon

T

etc. Microtiter plates and arrays are especially convenient because a large number of assays can be carried out simultaneously, using small amounts of reagents and samples. The particular manner of binding of the composition to the support is not crucial so long as it is compatible with the reagents and overall methods of the invention, maintains the activity of the composition and is nondiffusable. Preferred methods of binding include the use of antibodies which do not sterically block either the ligand binding site or activation sequence when attaching the protein to the support, direct binding to "sticky" or ionic supports, chemical crosslinking, the synthesis of the protein or agent on the surface, etc. Following binding of the protein or ligand/binding agent to the support, excess unbound material is removed by washing. The sample receiving areas may then be blocked through incubation with bovine serum albumin (BSA), casein or other innocuous protein or other moiety.

Once a cancer protein of the invention is bound to the support, and a test compound is added to the assay.

Alternatively, the candidate binding agent is bound to the support and the cancer protein of the invention is then added.

Binding agents include specific antibodies, non-natural binding agents identified in screens of chemical libraries, peptide analogs, etc.

Of particular interest are assays to identify agents that have a low toxicity for human cells. A wide variety of assays can be used for this purpose, including proliferation assays, cAMP assays, labeled in vitro protein-protein binding assays, electrophoretic mobility shift assays, immunoassays for protein binding, functional assays (phosphorylation assays, etc.) and the like.

WO 2004/050828 PCT/US2002/038264 A determination of binding of the test compound (ligand, binding agent, modulator, etc.) to a cancer protein of the invention can be done in a number of ways. The test compound can be labeled, and binding determined directly, by attaching all or a portion of the cancer protein of the invention to a solid support, adding a labeled candidate compound a fluorescent label), washing off excess reagent, and determining whether the label is present on the solid support. Various blocking and washing steps can be utilized as appropriate.

In certain embodiments, only one of the components is labeled, a protein of the invention or ligands labeled.

Alternatively, more than one component is labeled with different labels, I125, for the proteins and a fluorophor for the compound. Proximity reagents, quenching or energy transfer reagents are also useful.

Competitive Binding to Identify and Characterize Modulators In one embodiment, the binding of the "test compound" is determined by competitive binding assay with a "competitor." The competitor is a binding moiety that binds to the target molecule a cancer protein of the invention).

Competitors include compounds such as antibodies, peptides, binding partners, ligands, etc. Under certain circumstances, the competitive binding between the test compound and the competitor displaces the test compound. In one embodiment, the test compound is labeled. Either the test compound, the competitor, or both, is added to the protein for a time sufficient to allow binding. Incubations are performed at a temperature that facilitates optimal activity, typically between four and Incubation periods are typically optimized, to facilitate rapid high throughput screening; typically between zero and one hour will be sufficient. Excess reagent is generally removed or washed away. The second component is then added, and the presence or absence of the labeled component is followed, to indicate binding.

In ore embodiment, the competitor is added first, followed by the test compound. Displacement of the competitor is an indicatior that the test compound is binding to the cancer protein and thus is capable of binding to, and potentially modulating, the activity of the cancer protein. In this embodiment, either component can be labeled. Thus, if the competitor is labeled, the presence of label in the post-test compound wash solution indicates displacement by the test compound. Alternatively, if the test compound is labeled, the presence of the label on the support indicates displacement.

In an alternative embodiment the test compound is added first, with incubation and washing, followed by the competitor. The absence of binding by the competitor indicates that the test compound binds to the cancer protein with higher affinity than the competitor. Thus, if the test compound is labeled, the presence of the label on the support, coupled with a lack of competitor binding, indicates that the test compound binds to and thus potentially modulates the cancer protein of the invention.

Accordingly, the competitive binding methods comprise differential screening to identity agents that are capable of modulating the activity of the cancer proteins of the invention. In this embodiment, the methods comprise combining a cancer protein and a competitor in a first sample. A second sample comprises a test compound, the cancer protein, and a competitor. The binding of the competitor is determined for both samples, and a change, or difference in binding between the two samples indicates the presence of an agent capable of binding to the cancer protein and potentially modulating its activity. That is, if the binding of the competitor is different in the second sample relative to the first sample, the agent is capable of binding to the cancer protein.

Alternatively, differential screening is used to identify drug candidates that bind to the native cancer protein, but cannot bind to modified cancer proteins. For example the structure of the cancer protein is modeled and used in rational drug design to synthesize agents that interact with that site, agents which generally do not bind to site-modified proteins.

Moreover, such drug candidates that affect the activity of a native cancer protein are also identified by screening drugs for the ability to either enhance or reduce the activity of such proteins.

WO 2004/050828 PCT/US2002/038264 Positive controls and negative controls can be used in the assays. Preferably control and test samples are performed in at least triplicate to obtain statistically significant results. Incubation of all samples occurs for a time sufficient to allow for the binding of the agent to the protein. Following incubation, samples are washed free of non-specifically bound material and the amount of bound, generally labeled agent determined. For example, where a radiolabel is employed, the samples can be counted in a scintillation counter to determine tie amount of bound compound.

A variety of other reagents can be included in the screening assays. These include reagents like salts, neutral proteins, e.g. albumin, detergents, etc. which are used to facilitate optimal protein-protein binding and/or reduce non-specific or background interactions. Also reagents that otherwise improve the efficiency of the assay, such as protease inhibitors, nuclease inhibitors, anti-microbial agents, etc., can be used. The mixture of components is added in an order that provides for the requisite binding.

Use of Polynucleotides to Down-regulate or Inhibit a Protein of the Invention.

Polynucleotide modulators of cancer can be introduced into a cell containing the target nucleotide sequence by formation of a conjugate with a ligand-binding molecule, as described in WO 91/04753. Suitable ligand-binding molecules include, but are not limited to, cell surface receptors, growth factors, other cytokines, or other ligands that bind to cell surface receptors. Preferably, conjugation of the ligand binding molecule does not substantially interfere with the ability of the ligand binding molecule to bind to its corresponding molecule or receptor, or block entry of the sense or antisense oligonucleotide or its conjugated version into the cell. Alternatively, a polynucleotide modulator of cancer can be introduced into a cell containing the target nucleic acid sequence, by formation of a polynucleotide-lipid complex, as described in WO 90/10448. It is understood that the use of antisense molecules or knock out and knock in models may also be used in screening assays as discussed above, in addition to methods of treatment.

Inhibitory and Antisense Nucleotides In certain embodiments, the activity of a cancer-associated protein is down-regulated, or entirely inhibited, by the use of antisense polynucleotide or inhibitory small nuclear RNA (snRNA), a nucleic acid complementary to, and which can preferably hybridize specifically to, a coding mRNA nucleic acid sequence, a cancer protein of the invention, mRNA, or a subsequence thereof. Binding of the antisense polynucleotide to the mRNA reduces the translation and/or stability of the mRNA.

In the context of this invention, antisense polynucleotides can comprise naturally occurring nucleotides, or synthetic species formed from naturally occurring subunits or their close homologs. Antisense polynucleotides may also have altered sugar moieties or inter-sugar linkages. Exemplary among these are the phosphorothioate and other sulfur containing species which are known for use in the art. Analogs are comprised by this invention so long as they function effectively to hybridize with nucleotides of the invention. See, Isis Pharmaceuticals, Carlsbad, CA; Sequitor, Inc., Natick, MA.

Such antisense polynucleotides can readily be synthesized using recombinant means, or can be synthesized in vitro. Equipment for such synthesis is sold by several vendors, including Applied Biosystems. The preparation of other oligonuceotides such as phosphorothioates and alkylated derivatives is also well known to those of skill in the art Antisense molecules as used herein include antisense or sense oligonucleotides. Sense oligonucleotides can, be employed to block transcription by binding to the anti-sense strand. The antisense and sense oligonucleotide comprise a single stranded nucleic acid sequence (either RNA or DNA) capable of binding to target mRNA (sense) or DNA (antisense) sequences for cancer molecules. Antisense or sense oligonucleotides, according to the present invention, comprise a fragment generally at least about 12 nucleotides, preferably from about 12 to 30 nucleotides. The ability to derive WO 2004/050828 PCT/US2002/038264 an antisense or a sense oligonucleotide, based upon a cDNA sequence encoding a given protein is described in, Stein &Cohen (Cancer Res. 48:2659 (1988 and van der Krol et al. (BioTechniques 6:958 (1988)).

Ribozvymes In addition to antisense polynucleotides, ribozymes can be used to target and inhibit transcription of cancerassociated nucleotide sequences. A ribozyme is an RNA molecule that catalytically cleaves other RNA molecules. Different kinds of ribozymes have been described, including group I ribozymes, hammerhead ribozymes, hairpin ribozymes, RNase P, and axhead ribozymes (see, Castanotto et al., Adv. in Pharmacology 25: 289-317 (1994) for a general review of the properties of different ribozymes).

The general features of hairpin ribozymes are described, in Hampel et al., Nucl. Acids Res. 18:299-304 (1990); European Patent Publication No. 0360257; U.S. Patent No. 5,254,678. Methods of preparing are well known to those of skill in the art (see, WO 94/26877; Ojwang et al., Proc. Natl. Acad. Sci. USA 90:6340-6344 (1993); Yamada et al., Human Gene Therapy 1:39-45 (1994); Leavittet al., Proc. Natl. Acad Sci. USA 92:699- 703 (1995); Leavitt et al., Human Gene Therapy 5:1151-120 (1994); and Yamada et al., Virology 205:121-126 (1994)).

Use of Modulators in Phenotypic Screening In one embodiment, a test compound is administered to a population of cancer cells, which have an associated cancer expression prof le. By "administration" or "contacting' herein is meant that the modulator is added to the cells in such a manner as to allow the modulator to act upon the cell, whether by uptake and intracellular action, or by action at the cell surface. In some embodiments, a nucleic acid encoding a proteinaceous agent a peptide) is put into a viral construct such as an adenoviral or retroviral construct, and added to the cell, such that expression of the peptide agent is accomplished, PCT US97/01019. Regulatable gene therapy systems can also be used. Once the modulator has been administered to the cells, the cells are washed if desired and are allowed to incubate under preferably physiological conditions for some period. The cells are then harvested and a new gene expression profile is generated. Thus, e.g., cancer tissue is screened for agents that modulate, induce or suppress, the cancer phenotype. A change in at least one gene, preferably many, of the expression profile indicates that the agent has an effect on cancer activity. Similarly, altering a biological function or a signaling pathway is indicative of modulator activity. By defining such a signature for the cancer phenotype, screens for new drugs that alter the phenotype are devised. With this approach, the drug target need not be known and need not be represented in the original genelprotein expression screening platform, nor does the level of transcript for the target protein need to change. The modulator inhibiting function will serve as a surrogate marker As outlined above, screens are done to assess genes or gene products. That is, having identified a particular differentially expressed gene as important in a particular state, screening of modulators of either the expression of the gene or the gene product itself is performed.

Use of Modulators to Affect Peptides of the Invention Measurements of cancer polypeptide activity, or of the cancer phenotype are performed using a variety of assays.

For example, the effects of modulators upon the function of a cancer polypeptide(s) are measured by examining parameters described above. A physiological change that affects activity is used to assess the influence of a test compound on the polypeptides of this invention. When the functional outcomes are determined using intact cells or animals, a variety of effects can be assesses such as, in the case of a cancer associated with solid tumors, tumor growth, tumor metastasis, neovascularization, hormone release, transcriptional changes to both known and uncharacterized genetic markers by Northern blots), changes in cell metabolism such as cell growth or pH changes, and changes in intracellular second messengers such as cGNIP.

WO 2004/050828 PCT/US2002/038264 Methods of Identifying Characterizing Cancer-associated Sequences Expression of various gene sequences is correlated with cancer. Accordingly, disorders based on mutant or variant cancer genes are determined. In one embodiment, the invention provides methods for identifying cells containing variant cancer genes, determining the presence of, all or part, the sequence of at least one endogenous cancer gene in a cell. This is accomplished using any number of sequencing techniques. The invention comprises methods of identifying the cancer genotype of an individual, determining all or part of the sequence of at least one gene of the invention in the individual. This is generally done in at least one tissue of the individual, a tissue set forth in Table I, and may include the evaluation of a number of tissues or different samples of the same tissue. The method may include comparing the sequence of the sequenced gene to a known cancer gene, a wild-type gene to determine the presence of family members, homologies, mutations or variants. The sequence of all or part of the gene can then be compared to the sequence of a known cancer gene to determine if any differences exist. This is done using any number of known homology programs, such as BLAST, Bestfit, etc. The presence of a difference in the sequence between the cancer gene of the patient and the known cancer gene correlates with a disease state or a propensity for a disease state, as outlined herein.

In a preferred embodiment, the cancer genes are used as probes to determine the number of copies of the cancer gene in the genome. The cancer genes are used as probes to determine the chromosomal localization of the cancer genes.

Information such as chromosomal localization finds use in providing a diagnosis or prognosis in particular when chromosomal abnormalities such as translocations, and the like are identified in the cancer gene locus.

XIV.) Kits/Articles of Manufacture For use in the diagnostic and therapeutic applications described herein, kits are also within the scope of the invention. Such kits can comprise a carrier, package or container that is compartmentalized to receive one or more containers such as vials, tubes, and the like, each of the container(s) comprising one of the separate elements to be used in the method. For example, the container(s) can comprise a probe that is or can be detectably labeled. Such probe can be an antibody or polynucleotide specific for a Figure 2-related protein or a Figure 2 gene or message, respectively. Where the method utilizes nucleic acid hybridization to detect the target nucleic acid, the kit can also have containers containing nucleotide(s) for amplification of the target nucleic acid sequence and/or a container comprising a reporter-means, such as a biotin-binding protein, such as avidin or streptavidin, bound to a reporter molecule, such as an enzymatic, florescent, or radioisotope label. The kit can include all or part of the amino acid sequences in Figure 2 or Figure 3 or analogs thereof, or a nucleic acid molecules that encodes such amino acid sequences.

The kit of the invention will typically comprise the container described above and one or more other containers comprising materials desirable from a commercial and user standpoint, including buffers, diluents, filters, needles, syringes; carrier, package, container, vial and/or tube labels listing contents and/or instructions for use, and package inserts with instructions for use.

A label can be present on the container to indicate that the composition is used for a specific therapy or non-therapeutic application, such as a diagnostic or laboratory application, and can also indicate directions for either in vivoor in vitro use, such as those described herein. Directions and or other information can also be included on an insert(s) or label(s) which is included with or on the kit.

The terms "kit" and "article of manufacture" can be used as synonyms.

In another embodiment of the invention, an article(s) of manufacture containing compositions, such as amino acid sequence(s), small molecule(s), nucleic add sequence(s), and/or antibody(s), materials useful for the diagnosis, prognosis, prophylaxis and/or treatment of neoplasias of tissues such as those set forth in Table I is provided. The article of WO 2004/050828 PCT/US2002/038264 manufacture typically comprises at least one container and at least one label. Suitable containers include, for example, bottles, vials, syringes, and test tubes. The containers can be formed from a variety of materials such as glass or plastic.

The container can hold amino acid sequence(s), small molecule(s), nucleic acid sequence(s), and/or antibody(s), in one embodiment the container holds a polynucleotide for use in examining the mRNA expression profile of a cell,, together with reagents used for this purpose.

The container can alternatively hold a composition which is effective for treating, diagnosis, prognosing or prophylaxing a condition and can have a sterile access port (for example the container can be an intravenous solution bag or a vial having a stopper pierceable by a hypodermic injection needle). The active agents in the composition can be an antibody capable of specifically binding 24P4C12 and modulating the function of 24P4C12.

The label can be on or associated with the container. A label a can be on a container when letters, numbers or other characters forming the label are molded or etched into the container itself; a label can be associated with a container when it is present within a receptacle or carrier that also holds the container, as a package insert. The label can indicate that the composition is used for diagnosing, treating, prophylaxing or prognosing a condition, such as a neoplasia of a tissue set forth in Table I. The article of manufacture can further comprise a second container comprising a pharmaceutically-acceptable buffer, such as phosphate-buffered saline, Ringer's solution and/ordextrose solution. It can further include other materials desirable from a commercial and user standpoint, including other buffers, diluents, filters, stirrers, needles, syringes, and/or package inserts with indications and/or instructions for use.

EXAMPLES:

Various aspects of the invention are further described and illustrated by way of the several examples that follow, none of which are intended to limit the scope of the invention.

Example 1: SSH-Generated Isolation of cDNA Fragment of the 24P4C12 Gene Suppression Subtractive Hybridization (SSH) was used to identify cDNAs corresponding to genes that may be differentially expressed in prostate cancer. The SSH reaction utilized cDNA from the LAPC-9 AD prostate cancer xenograft. The gene 24P4C12 was derived from an LAPC-9 AD minus benign prostatic hyperplasia experiment.

The 24P4C12 SSH cDNA of 160 bp is listed in Figure 1. The full length 24P4C12 cDNAs and ORFs are described in.Figure 2 with the protein sequences listed in Figure 3.

Materials and Methods Human Tissues: The patient cancer and normal tissues were purchased from different sources such as the NDRI (Philadelphia, PA).

mRNA for some normal tissues were purchased from Clontech, Palo Alto, CA.

RNA Isolation: Tissues were homogenized in Trizol reagent (Life Technologies, Gibco BRL) using 10 ml/ g tissue isolate total RNA. Poly A RNA was purified from total RNA using Qiagen's Oligotex mRNA Mini and Midi kits. Total and mRNA were quantified by spectrophotometric analysis 2601280 nm) and analyzed by gel electrophoresis.

Oligonucleotides: The following HPLC purified oligonucleotides were used.

DPNCDN (cDNA synthesis primer): 5'TTTTGATCAAGCTT3o3' (SEQ ID NO: 33) WO 2004/050828 PCT/US20021038264 Adaptor 1: 5'CTAATACGACTCACTATAGGGCTCGAGCGGCCGCCCGGGCAG3' (SEQ ID NO: 34) (SEQ ID NO: Adaptor 2: 5'GTAATACGACTCACTATAGGGCAGCGTGGTCGCGGCCGAG3' (SEQ ID NO: 36) (SEQ ID NO: 37) PCR primer 1: 5'CTAATACGACTCACTATAGGGC3' (SEQ ID NO: 38) Nested primer (NP)1: 5'TCGAGCGGCCGCCCGGGCAGGA3' (SEQ ID NO: 39) Nested primer (NP)2: 5'AGCGTGGTCGCGGCCGAGGA3' (SEQ ID NO: Suppression Subtractive Hybridization: Suppression Subtractive Hybridization (SSH) was used to identify cDNAs corresponding to genes that may be differentially expressed in prostate cancer. The SSH reaction utilized cDNA from prostate cancer and normal tissues.

The gene 24P4C12 sequence was derived from LAPC-4AD prostate cancer xenograft minus begnin prostatic hyperplasia cDNA subtraction. The SSH DNA sequence (Figure 1) was identified.

The cDNA derived from a pool of normal tissues and benign prostatic hyperplasia was used as the source of the "driver" cDNA, while the cDNA from LAPC-4AD xenograft was used as the source of the "tester' cDNA. Double stranded cDNAs corresponding to tester and driver cDNAs were synthesized from 2 pg of poly(A)* RNA isolated from the relevant xenograft tissue, as described above, using CLONTECH's PCR-Select cDNA Subtraction Kit and 1 ng of oligonucleotide DPNCDN as primer. Firstand second-strand synthesis were carried out as described in the Kit's user manual protocol (CLONTECH Protocol No. PT1117-1, Catalog No. K1804-1). The resulting cDNA was digested with Dpn II for 3 hrs at 370C. Digested cDNA was extracted with phenol/chloroform and ethanol precipitated.

Driver cDNA was generated by combining In a 1:1 ratio Dpn II digested cDNA from the relevant tissue source (see above) with a mix of digested cDNAs derived from the nine normal tissues: stomach, skeletal muscle, lung, brain, liver, kidney, pancreas, small intestine, and heart.

Tester cDNA was generated by diluting 1 il of Dpn II digested cDNA from the relevant tissue source (see above) (400 ng) in 5 pl of water. The diluted cDNA (2 p1, 160 ng) was then ligated to 2 pi of Adaptor 1 and Adaptor 2 (10 pM), in separate ligation reactions, in a total volume of 10 pl at 16oC overnight, using 400 u of T4 DNA ligase (CLONTECH). Ligation was terminated with 1 pl of 0.2 M EDTA and heating at 720C for 5 min.

The first hybridization was performed by adding 1.5 pl (600 ng) of driver cDNA to each of two tubes containing 1.5 pl ng) Adaptor 1- and Adaptor 2- ligated tester cDNA. In a final volume of 4 pi, the samples were overlaid with mineral oil, denatured in an MJ Research thermal cyder at 98oC for 1.5 minutes, and then were allowed to hybridize for 8 hrs at 683C. The two hybridizations were then mixed together with an additional 1 pl of fresh denatured driver cDNA and were allowed to hybridize WO 2004/050828 PCT/US2002/038264 overnight at 680C. The second hybridization was then diluted in 200 pl of 20 mM Hepes, pH 8.3, 50 mM NaCI, 0.2 mM EDTA, heated at 70oC for 7 min. and stored at -200C.

PCR Amplification. Cloning and Sequencing of Gene Fragments Generated from SSH: To amplify gene fragments resulting from SSH reactions, two PCR amplifications were performed. In the primary PCR reaction 1 p of the diluted final hybridization mix was added to 1 pl of PCR primer 1 (10 pM), 0.5 pJ dNTP mix (10 pM), 2.5 pl x reaction buffer (CLONTECH) and 0.5 pJ 50 x Advantage cDNA polymerase Mix (CLONTECH) in a final volume of 25 pl. PCR 1 was conducted using the following conditions: 75oC for 5 min., 94oC for 25 sec., then 27 cycles of 94oC for 10 sec, 660C for 30 sec, 72°C for 1.5 min. Five separate primary PCR reactions were performed for each experiment. The products were pooled and diluted 1:10 with water. For the secondary PCR reaction, 1 pi from the pooled and diluted primary PCR reaction was added to the same reaction mix as used for PCR 1, except that primers NP1 and NP2 (10 pM) were used instead of PCR primer 1. PCR 2 was performed using 10-12 cycles of 94oC for 10 sec, 68oC for 30 sec, and 72oC for 1.5 minutes. The PCR products were analyzed using 2% agarose gel electrophoresis.

The PCR products were inserted into pCR2.1 using the T/A vector cloning kit (Invitrogen). Transformed E coliwere subjected to blue/white and ampicillin selection. White colonies were picked and arrayed into 96 well plates and were grown in liquid culture overnight. To identify inserts, PCR amplification was performed on 1 ul of bacterial culture using the conditions of PCR1 and NP1 and NP2 as primers. PCR products were analyzed using 2% agarose gel electrophoresis.

Bacterial clones were stored in 20% glycerol in a 96 well format. Plasmid DNA was prepared, sequenced, and subjected to nucleic acid homology searches of the Gen Bank, dBest, and NCI-CGAP databases.

RT-PCR Expression Analysis: First strand cDNAs can be generated from 1 pg of mRNA with oligo (dT)12-18 priming using the Gibco-BRL Superscript Preamplification system. The manufacturer's protocol was used which included an incubation for 50 min at 42oC with reverse transcriptase followed by RNAse H treatment at 370C for 20 min. After completing the reaction, the volume can be increased to 200 p with water prior to normalization. First strand cDNAs from 16 different normal human tissues can be obtained from Clontech.

Normalization of the first strand cDNAs from multiple tissues was performed by using the primers 5'atatcgccgcgctcgtcgtcgacaa3' (SEQ ID NO: 41) and 5'agccacacgcagctcattgtagaagg 3' (SEQ ID NO: 42) to amplify p-actin. First strand cDNA (5 pi) were amplified in a total volume of 50 pi containing 0.4 pM primers, 0.2 pM each dNTPs, 1 XPCR buffer (Clontech, 10 mM Tris-HCL, 1.5 mM MgCl2, 50 mM KCI, pH8.3) and 1X Klentaq DNA polymerase (Clontech). Five pl of the PCR reaction can be removed at 18, 20, and 22 cycles and used for agarose gel electrophoresis. PCR was performed using an MJ Research thermal cycler under the following conditions: Initial denaturation can be at 940C for 15 sec, followed by a 18, 20, and 22 cycles of 94C for 15, 65oC for 2 min, 72oC for 5 sec. A final extension at 72oC was carried out for 2 min. After agarose gel electrophoresis, the band intensities of the 283 b.p. p-actin bands from multiple tissues were compared by visual inspection.

Dilution factors for the first strand cDNAs were calculated to result in equal p-actin band intensities in all tissues after 22 cycles of PCR. Three rounds of normalization can be required to achieve equal band intensities in all tissues after 22 cycles of PCR.

To determine expression levels of the 24P4C12 gene, 5 p1 of normalized first strand cDNA were analyzed by PCR using 26, and 30 cycles of amplification. Semi-quantitative expression analysis can be achieved by comparing the PCR products at cycle numbers that give light band intensities. The primers used for RT-PCR were designed using the 24P4C12 SSH sequence and are listed below: 24P4C12.1 AGATGAGGAGGAGGACAAAGGTG 3' (SEQ ID NO: 43) WO 2004/050828 PCT/US2002/038264 24P4C12.2 ACTGCTGGGAGGAGTACCGAGTG 3' (SEQ ID NO: 44) Example 2: Isolation of Full Length 24P4C12 Encoding cDNA The 24P4C12 SSH cDNA sequence was derived from a substraction consisting of LAPC-4AD xenograft minus benign prostatic hyperplasia. The SSH cDNA sequence (Figure 1) was designated 24P4C12.

The isolated gene fragment of 150 bp encodes a putative open reading frame (ORF) of 53 amino adds and exhibits significant homology to an EST derived from a colon tumor library. Two larger cDNA clones were obtained by gene trapper experiments, GTE9 and GTF8. The ORF revealed a significant homology to the mouse gene NG22 and the C.elegans gene CEESB82F. NG22 was recently identified as one of many ORFs within a genomic BAC clone that encompasses the MHC class III in the mouse genome. Both NG22 and CEESB82F appear to be genes that contain 12 transmembrane domains. This suggests that the gene encoding 24P4C12 contains 12 transmembrane domains and is the human homologue of mouse NG22 and C.

elegans CEESB82F. Functional studies in Ce. elegans may reveal the biological role of these homologs. If 24P4C12 is a cell surface marker, then it may have an application as a potential imaging reagent andlor therapeutic target in prostate cancer.

The 24P4C12 v.1 of 2587 bp codes for a protein of 710 amino acids (Figure 2 and Figure Other variants of 24P4C12 were also identified and these are listed in Figures 2 and 3.24P4C12 v.1, v.3, v.5 and v.6 proteins are 710 amino acids in length and differ from each other by one amino acid as shown in Figure 11. 24P4C12 v.2 and v.4 code for the same protein as 24P4C12 v.1. 24P4C12 v.7, v.8 and v.9 are alternatve splice variants and code for proteins of 598, 722 and 712 amino acids in length, respectively.

Example 3: Chromosomal Mapping of 24P4C12 Chromosomal localization can implicate genes in disease pathogenesis. Several chromosome mapping approaches are available including fluorescent in situ hybridization (FISH), human/hamster radiation hybrid (RH) panels (Walter et al., 1994; Nature Genetics 7:22; Research Genetics, Huntsville Al), human-rodent somatic cell hybrid panels such as is available from the Coriell Institute (Camden, New Jersey), and genomic viewers utilizing BLAST homologies to sequenced and mapped genomic clones (NCBI, Bethesda, Maryland). 24P4C12 maps to chromosome 6p21.3 using 24P4C12 sequence and the NCBI BLAST tool located on the World Wide Web at (.ncbi.nlm.nih.gov/genome/seqfpage.cgi?F=HsBlast.html&&ORG=Hs).

Example 4: Expression Analysis of 24P4C12 Expression analysis by RT-PCR demonstrated that 24P4C12 is strongly expressed in prostate and ovary cancer patient specimens (Figure 14). First strand cDNA was generated from vital pool 1 (kidney, liver and lung), vital pool 2 (colon, pancreas and stomach), a pool of prostate cancer xenografts (LAPC-4AD, LAPC-4AI, LAPC-9AD and LAPC-9AI), prostate cancer pool, bladder cancer pool, kidney cancer pool, colon cancer pool, ovary cancer pool, breast cancer pool, and cancer metastasis pool.

Normalization was performed by PCR using primers to actin. Semi-quantitative PCR, using primers to 24P4C12, was performed at 26 and 30 cycles of amplification. Results show strong expression of 24P4C12 in prostate cancer pool and ovary cancer pool.

Expression was also detected in prostate cancer xenografts, bladder cancer pool, kidney cancer pool, colon cancer pool, breast cancer pool, cancer metastasis pool, vital pool 1, and vital pool 2.

Extensive northern blot analysis of 24P4C12 in multiple human normal tissues is shown in Figure 15. Two multiple tissue northern blots (Clontech) both with 2 pg of mRNA/lane were probed with the 24P4C12 SSH sequence. Expression of 24P4C12 was detected in prostate, kidney and colon. Lower expression is detected in pancreas, lung and placenta amongst all 16 normal tissues tested.

WO 2004/050828 PCT/US2002/038264 Expression of 24P4C12 was tested in prostate cancer xenografts and cell lines. RNA was extracted from a panel of cell lines and prostate cancer xenografts (PrEC, LAPC-4AD, LAPC-4AI, LAPC-9AD, LAPC-9AI, LNCaP, PC-3, DU145, TsuPr, and LAPC- 4CL). Northern blot with 10 pg of total RNNlane was probed with 24P4C12 SSH sequence. Size standards in kilobases (kb) are indicated on the side. The 24P4C12 transcript was detected in LAPC-4AD, LAPC-4AI, LAPC-9AD, LAPC-9AI, LNCaP, and LAPC-4

CL

Expression of 24P4C12 in patient cancer specimens and human normal tissues is shown in Figure 16. RNA was extracted from a pool of prostate cancer specimens, bladder cancer specimens, colon cancer specimens, ovary cancer specimens, breast cancer specimens and cancer metastasis specimens, as well as from normal prostate normal bladder normal kidney and normal colon Northern blot with 10 pg of total RNAlane was probed with 24P4C12 SSH sequence. Size standards in kilobases (kb) are indicated on the side. Strong expression of 24P4C12 transcript was detected in the patient cancer pool specimens, and in normal prostate but not in the other normal tissues tested.

Expression of 24P4C12 was also detected in individual prostate cancer patient specimens (Figure 17). RNA was extracted from normal prostate prostate cancer patient tumors and their matched normal adjacent tissues (Nat).

Northern blots with 10 pg of total RNA were probed with the 24P4C12 SSH fragment. Size standards in kilobases are on the side. Results show expression of 24P4C12 in normal prostate and all prostate patient tumors tested.

Expression of 24P4C12 in colon cancer patient specimens is shown in figure 18. RNA was extracted from colon cancer cell lines (CL: Colo 205, LoVo, and SK-CO-), normal colon colon cancer patient tumors and their matched normal adjacent tissues (Nat). Northern blots with 10 pg of total RNA were probed with the 24P4C12 SSH fragment. Size standards in kilobases are on the side. Results show expression of 24P4C12 in normal colon and all colon patient tumors tested. Expression was detected in the cell lines Colo 205 and SK-CO-, but not in LoVo.

Figure 20 displays expression results of 24P4C12 in lung cancer patient specimens. Ma was extracted from lung cancer cell lines (CL: CALU-1, A427, NCI-H82, NCI-H146), normal lung lung cancer patient tumors and their matched normal adjacent tissues (Nat). Northern blots with 10 pg of total RNA were probed with the 24P4C12 SSH fragment. Size standards in kilobases are on the side. Results show expression of 24P4C12 in lung patient tumors tested, but not in normal lung. Expression was also detected in CALU-1, but not in the other cell lines A427, NCI-H82, and NCI- H146.

24P4C12 was assayed in a panel of human stomach and breast cancers and their respective matched normal tissues on RNA dot blots. 24P4C12 expression was seen in both stomach and breast cancers. The expression detected in normal adjacent tissues (isolated from diseased tissues) but not in normal tissues (isolated from healthy donors) may indicate that these tissues are not fully normal and that 24P4C12 may be expressed in early stage tumors.

The level of expression of 24P4C12 was analyzed and quantitated in a panel of patient cancer tissues. First strand cDNA was prepared from a panel of ovary patient cancer specimens uterus patient cancer specimens prostate cancer specimens bladder cancer patient specimens lung cancer patient specimens pancreas cancer patient specimens colon cancer specimens and kidney cancer specimens Normalization was performed by PCR using primers to actin. Semi-quantitative PCR, using primers to 24P4C12, was performed at 26 and 30 cycles of amplification.

Samples were run on an agarose gel, and PCR products were quantitated using the Alphalmager software. Expression was recorded as absent, low, medium or strong. Results show expression of 24P4C12 in the majority of patient cancer specimens tested, 73.3% of ovary patient cancer specimens, 83.3% of uterus patient cancer specimens, 95.0% of prostate cancer specimens, 61.1% of bladder cancer patient specimens, 80.6% of lung cancer patient specimens, 87.5% of pancreas cancer patient specimens, 87.5% of colon cancer specimens, 68.4% of clear cell renal carcinoma, 100% of papillary renal cell carcinoma.

WO 2004/050828 PCT/US2002/038264 The restricted expression of 24P4C12 in normal tissues and the expression detected in prostate cancer, ovary cancer, bladder cancer, colon cancer, lung cancer pancreas cancer, uterus cancer, kidney cancer, stomach cancer and breast cancer suggest that 24P4C12 is a potential therapeutic target and a diagnostic marker for human cancers.

Example 5: Transcript Variants of 24P4C12 Transcript variants are variants of mature mRNA from the same gene which arise by alternative transcription or alternative splicing. Alternative transcripts are transcripts from the same gene but start transcription at different points.

Splice variants are mRNA variants spliced differently from the same transcript. In eukaryotes, when a multi-exon gene is transcribed from genomic DNA, the initial RNA is spliced to produce functional mRNA, which has only exons and is used for translation into an amino acid sequence. Accordingly, a given gene can have zero to many alternative transcripts and each transcript can have zero to many splice variants. Each transcript variant has a unique exon makeup, and can have different coding andlor non-coding or 3' end) portions, from the original transcript. Transcript variants can code for similar or different proteins with the same or a similar function or can encode proteins with different functions, and can be expressed in the same tissue at the same time, or in different tissues at the same time, or in the same tissue at different times, or in different tissues at different times. Proteins encoded by transcript variants can have similar or different cellular or extracellular localizations, secreted versus intracellular.

Transcript variants are identified by a variety of art-accepted methods. For example, alternative transcripts and splice variants are identified by full-length cloning experiment, or by use of full-length transcript and EST sequences. First, all human ESTs were grouped into clusters which show direct or indirect identity with each other. Second, ESTs in the same cluster were further grouped into sub-clusters and assembled into a consensus sequence. The original gene sequence is compared to the consensus sequence(s) or other full-length sequences. Each consensus sequence is a potential splice variant for that gene. Even when a vanant is identified that is not a full-length clone, that portion of the variant is very useful for antigen generation and for further cloning of the full-length splice variant, using techniques known in the art.

Moreover, computer programs are available in the art that identify transcript variants based on genomic sequences. Genomic-based transcript variant identification programs include FgenesH Salamov and V. Solovyev, "Ab initio gene finding in Drosophila genomic DNA,' Genome Research. 2000 April; 10(4):516-22); Grail (URL at compbio.oml.gov/Grail-bin/EmptyGrailForm) and GenScan (URL at genes.mit.edu/GENSCAN.html). For a general discussion of splice variant identification protocols see., Southan, A genomic perspective on human proteases, FEBS Lett. 2001 Jun 8; 498(2-3):214-8; de Souza, et aL., Identification of human chromosome 22 transcribed sequences with ORF expressed sequence tags, Proc. Nal Acad Sci U S A. 2000 Nov 7; 97(23):12690-3.

To further confirm the parameters of a transcript variant, a variety of techniques are available in the art, such as full-length cloning, proteomic validaton, PCR-based validation, and 5' RACE validation, etc. (see Proteomic Validation: Brennan, et al., Albumin banks peninsula: a new termination variant characterized by electrospray mass spectrometry, Biochem Biophys Acta. 1999 Aug 17;1433(1-2):321-6; Ferranti P, et al., Differential splicing of pre-messenger RNA produces multiple forms of mature caprine alpha(s1)-casein, Eur J Biochem. 1997 Oct 1;249(1):1-7. For PCR-based Validation: Wellmann S, et al., Specific reverse transcription-PCR quantification of vascular endothelial growth factor (VEGF) splice variants by LightCycler technology, Clin Chem. 2001 Apr;47(4):654-60; Jia, et al., Discovery of new human betadefensins using a genomics-based approach, Gene. 2001 Jan 24; 263(1-2):211-8. For PCR-based and 5' RACE Validation: Brigle, et al., Organization of the murine reduced folate carrier gene and identification of variant splice forms, Biochem Biophys Acta. 1997 Aug 7; 1353(2): 191-8).

It is known in the art that genomic regions are modulated in cancers. When the genomic region to which a gene maps is modulated in a particular cancer, the alternative transcripts or splice variants of the gene are modulated as well.

WO 2004/050828 PCT/US2002/038264 Disclosed herein is that 24P4C12 has a particular expression profile related to cancer. Alternative transcripts and splice variants of 24P4C12 may also be involved in cancers in the same or different tissues, thus serving as tumor-associated markers/antigens.

The exon composition of the original transcript, designated as 24P4C12 v.1, is shown in Table LI. Using the fulllength gene and EST sequences, three transcript variants were identified, designated as 24P4C12 v.7, v.8 and v.9.

Compared with 24P4C12 v.1, transcript variant 24P4C12 v.7 has spliced out exons 10 and 11 from variant 24P4C12 v.1, as shown in Figure 12. Variant 24P4C12 v.8 inserted 36 bp in between 1931 and 1932 of variant 24P4C12 v.1 and variant 24P4C12 v.9 replaced with 36 bp the segment 1136-1163 of variant 24P4C12 v.1. Theoretically, each different combination of exons in spatial order, e.g. exons 2 and 3, is a potential splice variant. Figure 12 shows the schematic alignment of exons of the four transcript variants.

Tables LII through LXIII are set forth on a variant by variant basis. Tables LII, LVI, and LX show nucleotide sequences of the transcript variant. Tables LIII, LVII, and LXI show the alignment of the transcript variant with the nucleic acid sequence of 24P4C12 v.1. Tables LIV, LVIII, and LXII lay out the amino acid translation of the transcript variant for the identified reading frame orientation. Tables LV, LIX, and LXIII display alignments of the amino acid sequence encoded by the splice variant with that of 24P4C12 v, 1.

Example 6: Single Nucleotide Polymorphisms of 24P4C12 A Single Nucleotide Polymorphism (SNP) is a single base pair variation in a nucleotide sequence at a specific location. At any given point of the genome, there are four possible nucleotide base pairs: A/T, C/G, GIC and T/A. Genotype refers to the specific base pair sequence of one or more locations in the genome of an individual. Haplotype refers to the base pair sequence of more than one location on the same DNA molecule (or the same chromosome in higher organisms), often in the context of one gene or in the context of several lightly linked genes. SNPs that occur on a cDNA are called cSNPs. These cSNPs may change amino acids of the protein encoded by the gene and thus change the functons of the protein. Some SNPs cause inherited diseases; others contribute to quantitative variations in phenotype and reactions to environmental factors including diet and drugs among individuals. Therefore, SNPs and/or combinations of alleles (called haplotypes) have many applications, including diagnosis of inherited diseases, determination of drug reactions and dosage, identification of genes responsible for diseases, and analysis of the genetic relationship between individuals Nowotny, J.

M. Kwon and A. M. Goate," SNP analysis to dissect human traits," Curr. Opin. Neurobiol. 2001 Oct; 11(5):637-641; M.

Pirmohamed and B. K. Park, "Genetic susceptibility to adverse drug reactions," Trends Pharmacol. Sci. 2001 Jun; 22(6):298- 305; J. H. Riley, C. J. Allan, E. Lai and A. Roses, The use of single nucleotide polymorphisms in the isolation of common disease genes," Pharmacogenomics. 2000 Feb; 1(1):39-47; R. Judson, J. C. Stephens and A. Windemuth, "The predictive power of haplotypes in clinical response," Pharmacogenomics. 2000 feb; 1(1):15-26).

SNPs are identified by a variety of art-accepted methods Bean, "The promising voyage of SNP target discovery," Am. Clin. Lab. 2001 Oct-Nov; 20(9):18-20; K. M. Weiss, "In search of human variation," Genome Res. 1998 Jul; 8(7):691-697; M. M. She, "Enabling large-scale pharmacogenetic studies by high-throughput mutation detection and genotyping technologies," Clin. Chem. 2001 Feb; 47(2):164-172). For example, SNPs are identified by sequencing DNA fragments that show polymorphism by gel-based methods such as restriction fragment length polymorphism (RFLP) and denaturing gradient gel electrophoresis (DGGE). They can also be discovered by direct sequencing of DNA samples pooled from different individuals or by comparing sequences from different DNA samples. With the rapid accumulation of sequence data in public and private databases, one can discover SNPs by comparing sequences using computer programs Gu, L.

Hillier and P. Y. Kwok, "Single nucleotide polymorphism hunting in cyberspace," Hum. Mutat. 1993; 12(4):221-225). SNPs can be verified and genotype or haplotype of an individual can be determined by a variety of methods including direct WO 2004/050828 PCT/US2002/038264 sequencing and high throughput microarrays Y. Kwok, "Methods for genotyping single nucleotide polymorphisms," Annu.

Rev. Genomics Hum. Genet. 2001; 2:235-258; M. Kokoris, K. Dix, K. Moynihan, J. Mathis, B. Erwin, P. Grass, B. Hines and A. Duesterhoeft, "High-throughput SNP genotyping with the Masscode system," Mol. Diagn. 2000 Dec; 5(4):329-340).

Using the methods described above, five SNPs were identified in the original transcript, 24P4C12 v.1, at positions 542 (GIA), 564 818 981(A/G) and 1312 The transcripts or proteins with alternative alleles were designated as variants 24P4C12 v.2, v.3, v.4, v.5 and v.6, respectively. Figure 10 shows the schematic alignment of the SNP variants.

Figure 11 shows the schematic alignment of protein variants, corresponding to nudeotide variants. Nucleotide variants that code for the same amino acid sequence as variant 1 are not shown in Figure 11. These alleles of the SNPs, though shown separately here, can occur in different combinations (haplotypes) and in any one of the transcript variants (such as 24P4C12 v.7) that contains the sequence context of the SNPs.

Example 7: Production of Recombinant 24P4C12 in Prokarvotic Systems To express recombinant 24P4C12 and 24P4C12 variants in prokaryotic cells, the full or partial length 24P4C12 and 24P4C 12 variant cDNA sequences are cloned into any one of a variety of expression vectors known in the art. The full length cDNA, or any 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30 or more contiguous amino acids from 24P4C12, variants, or analogs thereof are used.

A. In vitro transcription and translation constructs: pCRII To generate 24P4C12 sense and anti-sense RNA probes for RNA in situ investigations, pCRII constructs (Invitrogen, Carlsbad CA) are generated encoding either all or fragments of the 24P4C12 cDNA. The pCRII vector has Sp6 and T7 promoters flanking the insert to drive the transcription of 24P4C12 RNA for use as probes in RNA in situ hybridization experiments. These probes are used to analyze the cell and tissue expression of 24P4C12 at the RNA level. Transcribed 24P4C12 RNA representing the cDNA amino acid coding region of the 24P4C12 gene is used in in vitro translation systems such as the TnTTM Coupled Reticulolysate System (Promega, Corp., Madison, WI) to synthesize 24P4C12 protein.

B. Bacterial Constructs: pGEX Constructs: To generate recombinant 24P4C12 proteins in bacteria that are fused to the Glutathione Stransferase (GST) protein, all or parts of the 24P4C12 cDNA or variants are cloned into the GST- fusion vector of the pGEX family (Amersham Pharmacia Biotech, Piscataway, NJ). These constructs allow controlled expression of recombinant 24P4C12 protein sequences with GST fused at the amino-terminus and a six histidine epitope (6X His) at the carboxylterminus. The GST and 6X His tags permit purification of the recombinant fusion protein from induced bacteria with the appropriate affinity matrix and allow recognition of the fusion protein with anti-GST and anti-His antibodies. The 6X His tag is generated by adding 6 histidine codons to the cloning primer at the 3' end, of the open reading frame (ORF). A proteolytic cleavage site, such as the PreScissionM recognition site in pGEX-6P-1, may be employed such that it permits cleavage of the GST tag from 24P4C12-related protein. The ampicillin resistance gene and pBR322 origin permits selection and maintenance of the pGEX plasmids in E. coli.

pMAL Constructs: To generate, in bacteria, recombinant 24P4C12 proteins that are fused to maltose-binding protein (MBP), all or parts of the 24P4C12 cDNA protein coding sequence are fused to the MBP gene by cloning into the pMAL-c2X and pMAL-p2X vectors (New England Biolabs, Beverly, MA). These constructs allow controlled expression of recombinant 24P4C12 protein sequences with MBP fused at the amino-terminus and a 6X His epitope tag at the carboxyl, terminus. The MBP and 6X His tags permit purification of the recombinant protein from induced bacteria with the appropriate affinity matrix and allow recognition of the fusion protein with anti-MBP and anti-His antibodies. The 6X His epitope tag is generated by adding 6 histidine codons to the 3' cloning primer. A Factor Xa recognition site permits cleavage of the pMAL WO 2004/050828 PCT/US2002/038264 tag from 24P4C12. The pMAL-c2X and pMAL-p2X vectors are optimized to express the recombinant protein in the cytoplasm or periplasm respectively. Periplasm expression enhances folding of proteins with disulfide bonds.

pET Constructs: To express 24P4C12 in bacterial cells, all or parts of the 24P4C12 cDNA protein coding sequence are cloned into the pET family of vectors (Novagen, Madison, WI). These vectors allow tightly controlled expression of recombinant 24P4C12 protein in bacteria with and without fusion to proteins that enhance solubility, such as NusA and thioredoxin (Trx), and epitope tags, such as 6X His and S-Tag T that aid purification and detection of the recombinant protein. For example, constructs are made utilizing pET NusA fusion system 43.1 such that regions of the 24P4C12 protein are expressed as amino-terminal fusions to NusA.

C. Yeast Constructs: pESC Constructs: To express 24P4C12 in the yeast species Saccharomyces cerevisiae for generation of recombinant protein and functional studies, all or parts of the 24P4C12 cDNA protein coding sequence are cloned into the pESC family of vectors each of which contain 1 of 4 selectable markers, HIS3, TRP1, LEU2, and URA3 (Stratagene, La Jolla, CA). These vectors allow controlled expression from the same plasmid of up to 2 different genes or cloned sequences containing either FlagTM or Myc epitope tags in the same yeast cell. This system is useful to confirm protein-protein interactions of 24P4C12. In addition, expression in yeast yields similar post-translational modifications, such as glycosylatons and phosphorylations, that are found when expressed in eukaryotic cells.

pESP Constructs: To express 24P4C12 in the yeast species Saccharomyces pombe, all or parts of the 24P4C12 cDNA protein coding sequence are cloned into the pESP family of vectors. These vectors allow controlled high level of expression of a 24P4C12 protein sequence that is fused at either the amino terminus or at the carboxyl terminus to GST which aids purification of the recombinant protein. A FlagTM epitope tag allows detection of the recombinant protein with anti- FlagTM antibody.

Example 8: Production of Recombinant 24P4C12 in Higher Eukarvotic Systems A. Mammalian Constructs: To express recombinant 24P4C12 in eukaryotic cells, the full or partial length 24P4C12 cDNA sequences can be cloned into any one of a variety of expression vectors known in the art. One or more of the following regions of 24P4C12 are expressed in these constructs, amino acids 1 to 710, or any 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21,22, 23, 24, 26, 27, 28, 29, 30 or more contiguous amino acids from 24P4C12 v.1 through v.6; amino acids 1 to 598, or any 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30 or more contiguous amino acids from 24P4C12 v.7; amino acids 1 to 722, or any 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30 or more contiguous amino acids from 24P4C12 v.8, amino acids 1 to712, or any 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18,19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30 or more contiguous amino acids from 24P4C12 v.9, variants, or analogs thereof.

The constructs can be transfected into any one of a wide variety of mammalian cells such as 293T cells.

Transfected 293T cell lysates can be probed with the anti-24P4C12 polyclonal serum, described herein.

pcDNA3.11MycHis Constructs: To express 24P4C12 in mammalian cells, a 24P4C12 ORF, or portions thereof, of 24P4C12 with a consensus Kozak translation initiation site was cloned into pcDNA3.11MycHis Version A (Invitrogen, Carlsbad, CA). Protein expression is driven from the cytomegalovirus (CMV) promoter. The recombinant proteins have the myc epitope and 6X His epitope fused to the carboxyl-terminus. The pcDNA3.1/MycHis vector also contains the bovine growth hormone (BGH) polyadenylation signal and transcription termination sequence to enhance mRNA stability, along with the SV40 origin for episomal replication and simple vector rescue in cell lines expressing the large T antigen. The Neomycin resistance gene can be used, as it allows for selection of mammalian cells expressing the protein and the ampicillin WO 2004/050828 PCT/US2002/038264 resistance gene and ColE1 origin permits selection and maintenance of the plasmid in E. coll. Figure 24 demonstrates expression of 24P4C12 from the pcDNA3.1/MycHis construct in transiently transfected 293T cells.

pcDNA41HisMax Constructs: To express 24P4C12 in mammalian cells, a 24P4C12 ORF, or portions thereof, of 24P4C12 are cloned into pcDNA4/HisMax Version A (Invitrogen, Carlsbad, CA). Protein expression is driven from the cytomegalovirus (CMV) promoter and the SP16 translational enhancer. The recombinant protein has XpressTM and six histidine (6X His) epitopes fused to the amino-terminus. The pcDNA4/HisMax vector also contains the bovine growth hormone (BGH) polyadenylation signal and transcription termination sequence to enhance mRNA stability along with the origin for episomal replication and simple vector rescue in cell lines expressing the large T antigen. The Zeocin resistance gene allows for selection of mammalian cells expressing the protein and the ampicillin resistance gene and ColE1 origin permits selection and maintenance of the plasmid in E. coli, pcDNA3.1ICT-GFP-TOPO Construct: To express 24P4C12 in mammalian cells and to alow detection of the recombinant proteins using fluorescence, a 24P4C12 ORF, or portions thereof, with a consensus Kozak translation initiation site are cloned into pcDNA3.1/CT-GFP-TOPO (Invitrogen, CA) Protein expression is driven from the cytomegalovirus (CMV) promoter. The recombinant proteins have the Green Fluorescent Protein (GFP) fused to the carboxyl-terminus facilitating non-invasive, in vivo detection and cell biology studies. The pcDNA3.1CT-GFP-TOPO vector also contains the bovine growth hormone (BGH) polyadenylation signal and transcription termination sequence to enhance mRNA stability along with the SV40 origin for episomal replication and simple vector rescue in cell lines expressing the large T antigen. The Neomycin resistance gene allows for selection of mammalian cells that express the protein and the ampicillin resistance gene and ColE1 origin permits selection and maintenance of the plasmid in E coil. Additional constructs with an aminoterminal GFP fusion are made in pcDNA3.1/NT-GFP-TOPO spanning the entire length of a 24P4C12 protein.

A 24P4C12 ORF, or portions thereof, were cloned into plag-5. This vector is similar to pAPtag but without the alkaline phosphatase fusion. This construct generates 24P4C12 protein with an amino-terminal IgGK signal sequence and myc and 6X His epitope tags at the carboxyl-terminus that facilitate detection and affinity purification. The resulting recombinant 24P4C12 protein were optimized for secretion into the media of transfected mammalian cells, and is used as immunogen or ligand to identify proteins such as ligands or receptors that interact with the 24P4C12 proteins.

Protein expression is driven from the CMV promoter. The Zeocin resistance gene present in the vector allows for selection of mammalian cells expressing the protein, and the ampicillin resistance gene permits selection of the plasmid in E. coli.

Figure 26 shows expression of 24P4C12 from two different pTag5 constructs.

PAPtag: A 24P4C12 ORF, or portions thereof, is cloned into pAPtag-5 (GenHunter Corp. Nashville, TN). This construct generates an alkaline phosphatase fusion at the carboxyl-terminus of a 24P4C12 protein while fusing the IgGK signal sequence to the amino-terminus. Constructs are also generated in which alkaline phosphatase with an aminoterminal IgGK signal sequence is fused to the amino-terminus of a 24P4C12 protein. The resulting recombinant 24P4C12 proteins are optimized for secretion into the media of transfected mammalian cells and can be used to identify proteins such as ligands or receptors that interact with 24P4C12 proteins. Protein expression is driven from the CMV promoter and the recombinant proteins also contain myc and 6X His epitopes fused at the carboxyl-terminus that facilitates detection and purification. The Zeocin resistance gene present in the vector allows for selection of mammalian cells expressing the recombinant protein and the ampicillin resistance gene permits selection of the plasmid in E. coli.

PsecFc: A 24P4C12 ORF, or portions thereof, is also cloned into psecFc. The psecFc vector was assembled by cloning the human immunoglobulin G1 (IgG) Fc (hinge, CH2, CH3 regions) into pSecTag2 (Invitrogen, California). This construct generates an IgG1 Fc fusion at the carboxyl-terminus of the 24P4C12 proteins, while fusing the IgGK signal sequence to N-terminus. 24P4C12 fusions utilizing the murine IgG1 Fc region are also used. The resulting recombinant 24P4C12 proteins are optimized for secretion into the media of transfected mammalian cells, and can be used as WO 2004/050828 PCT/US2002/038264 immunogens or to identify proteins such as ligands or receptors that interact with 24P4C12 protein. Protein expression is driven from the CMV promoter. The hygromycin resistance gene present in the vector allows for selection of mammalian cells that express the recombinant protein, and the ampicillin resistance gene permits selection of the plasmid in E coli.

pSRa Constructs: To generate mammalian cell lines that express 24P4C12 constitutively, 24P4C12 ORF, or portions thereof, of 24P4C12 were cloned into pSRa constructs. Amphotropic and ecotropic retroviruses were generated by transfection of pSRa constructs into the 293T-10A1 packaging line or co-transfection of pSRa and a helper plasmid (containing deleted packaging sequences) into the 293 cells, respectively. Theretrovirus is used to infect a variety of mammalian cell lines, resulting in the integration of the cloned gene, 24P4C12, into the host cell-lines. Protein expression is driven from a long terminal repeat (LTR). The Neomycin resistance gene presentin the vector allows for selection of mammalian cells that express the protein, and the ampicillin resistance gene and ColE1 origin permit selection and maintenance of the plasmid in E. coli. The retroviral vectors can thereafter be used for infection and generation of various cell lines using, for example, PC3, NIH 3T3, TsuPrl, 293 or rat-1 cells. Figure 23 shows RNA expression of 24P4C12 driven from the 24P4C12.pSRa construct in stably transduced PC3, 3T3 and 300.19 cells. Figure 25 shows 24P4C12 protein expression in PC3 cells stably transduced with 24P4C12.pSRa construct.

Additional pSRa constructs are made that fuse an epitope tag such as the FLAGTM tag to the carboxyl-terminus of 24P4C12 sequences to allow detection using anti-Flag antibodies. For example, the FLAGTM sequence 5' gat tac aag gat gac gac gat aag 3' (SEQ ID NO: 45) is added to cloning primer at the 3'end of the ORF. Additional pSRa constructs are made to produce both amino-terminal and carboxyl-terminal GFP and myc6X His fusion proteins of the full-length 24P4C12 proteins.

Additional Viral Vectors: Additional constructs are made for viral-mediated delivery and expression of 24P4C12.

High virus titer leading to high level expression of 24P4C12 is achieved in viral delivery systems such as adenoviral vectors and herpes amplicon vectors. A 24P4C12 coding sequences or fragments thereof are amplified by PCR and subcloned into the AdEasy shuttle vector (Stratagene). Recombination and virus packaging are performed according to the manufacturer's instructions to generate adenoviral vectors. Alternatively, 24P4C12 coding sequences or fragments thereof are cloned into the HSV-1 vector (Imgenex) to generate herpes viral vectors. The viral vectors are thereafter used for infection of various cell lines such as PC3, NIH 3T3, 293 or rat-1 cells.

Regulated Expression Systems: To control expression of 24P4C12 in mammalian cells, coding sequences of 24P4C12, or portions thereof, are cloned into regulated mammalian expression systems such as the T-Rex System (Invitrogen), the GeneSwitch System (Invitrogen) and the tightly-regulated Ecdysone System (Sratagene). These systems allow the study of the temporal and concentration dependent effects of recombinant 24P4C12. These vectors are thereafter used to control expression of 24P4C12 in various cell lines such as PC3, NIH 3T3, 293 or rat-1 cells.

B. Baculovirus Expression Systems To generate recombinant 24P4C12 proteins in a baculovirus expression system, 24P4C12 ORF, or portions thereof, are cloned into the baculovirus transfer vector pBlueBac 4.5 (Invitrogen), which provides a His-tag at the N-terminus.

Specifically, pBlueBac-24P4C12 is co-transfected with helper plasmid pBac-N-Blue (Invitrogen) into SF9 (Spodoptera frugiperda) insect cells to generate recombinant baculovirus (see Invitrogen instruction manual for details). Baculovirus is then collected from cell supernatant and purified by plaque assay.

Recombinant 24P4C12 protein is then generated by infection of HighFive insect cells (Invitrogen) with purified baculovirus. Recombinant 24P4C12 protein can be detected using anti-24P4C12 or anti-His-tag antibody. 24P4C12 protein can be purified and used in various cell-based assays or as immunogen to generate polyclonal and monoclonal antibodies specific for 24P4C12.

WO 2004/050828 PCT/US2002/038264 Example 9: Antigenicity Profiles and Secondary Structure Figures 5-9 depict graphically five amino acid profiles of the 24P4C12 variant 1, assessment available by accessing the ProtScale website located on the World Wide Web at (.expasy.ch/cgi-bin/protscale.pl) on the ExPasy molecular biology server.

These profiles: Figure 5, Hydrophilicity, (Hopp Woods 1981. Proc. Natl. Acad. Sci. U.S.A. 78:3824- 3828); Figure 6, Hydropathicity, (Kyte Doolittle 1982. J. Mol. Biol. 157:105-132); Figure 7, Percentage Accessible Residues (Janin 1979 Nature 277:491-492); Figure 8, Average Flexibility, (Bhaskaran and Ponnuswamy 1988.

Int. J. Pept. Protein Res. 32:242-255); Figure 9, Beta-turn (Deleage, Roux B. 1987 Protein Engineering 1:289-294); and optionally others available in the art, such as on the ProtScale website, were used to identify antigenic regions of the 24P4C12 protein. Each of the above amino add profiles of 24P4C12 were generated using the following ProtScale parameters for analysis: 1) A window size of 9; 2) 100% weight of the window edges compared to the window center; and, 3) amino acid profile values normalized to lie between 0 and 1.

Hydrophilicity (Figure Hydropathicity (Figure 6) and Percentage Accessible Residues (Figure 7) profiles were used to determine stretches of hydrophilic amino acids values greater than 0.5 on the Hydrophilicity and Percentage Accessible Residues profile, and values less than 0.5 on the Hydropathicity profile). Such regions are likely to be exposed to the aqueous environment, be present on the surface of the protein, and thus available for immune recognition, such as by antibodies.

Average Flexibility (Figure 8) and Beta-turn (Figure 9) profiles determine stretches of amino acids values greater than 0.5 on the Beta-turn profile and the Average Flexibility profile) that are not constrained in secondary structures such as beta sheets and alpha helices. Such regions are also more likely to be exposed on the protein and thus accessible to immune recognition, such as by antibodies.

Antigenic sequences of the 24P4C12 protein and of the variant proteins indicated, by the profiles set forth in Figure 5, Figure 6, Figure 7, Figure 8, and/or Figure 9 are used to prepare immunogens, either peptides or nucleic acids that encode them, to generate therapeutic and diagnostic anti-24P4C12 antibodies. The immunogen can be any 5, 6, 7, 8, 9, 11, 12, 13, 14,15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 30, 35, 40, 45, 50 or more than 50 contiguous amino acids, or the corresponding nucleic acids that encode them, from the 24P4C12 protein variants listed in Figures 2 and 3. In particular, peptide immunogens of the invention can comprise, a peptide region of at least 5 amino acids of Figures 2 and 3 in any whole number increment that includes an amino acid position having a value greater than 0.5 in the Hydrophilicity profile of Figure 5; a peptide region of at least 5 amino acids of Figures 2 and 3 in any whole number ncrement that includes an amino acid position having a value less than 0.5 in the Hydropathicity profile of Figure 6; a peptide region of at least 5 amino acids of Figures 2 and 3 in any whole number increment that includes an amino acid position having a value greater than 0.5 in the Percent Accessible Residues profile of Figure 7; a peptide region of at least 5 amino acids of Figures 2 and 3 in any whole number increment that includes an amino acid position having a value greater than 0.5 in the Average Flexibility profile on Figure 8; and, a peptide region of at least 5 amino acids of Figures 2 and 3 in any whole number increment that includes an amino acid position having a value greater than 0.5 in the Beta-turn profile of Figure 9. Peptide immunogens of the invention can also comprise nucleic acids that encode any of the forgoing.

All immunogens of the invention, peptide or nucleic acid, can be embodied in human unit dose form, or comprised by a composition that includes a pharmaceutical excipient compatible with human physiology.

The secondary structure of 24P4C12 variant 1, namely the predicted presence and location of alpha helices, extended strands, and random coils, are predicted from the respective primary amino acid sequences using the HNN Hierarchical Neural Network method (Guermeur, 1997, http://pbil.ibcp.fr/cgi-bin/npsaautomat.pl?page=npsann.html), WO 2004/050828 PCT/US2002/038264 accessed from the ExPasy molecular biology server (http:l/www.expasy.chltools/). The analysis indicates that 24P4C12 variant 1 is composed of 53.94% alpha helix, 9.44% extended strand, and 36.62% random coil (Figure 13a).

Analysis for the potential presence of transmembrane domains in 24P4C12 variants were carried out using a variety of transmembrane prediction algorithms accessed from the ExPasy molecular biology server (http://www.expasy.chltoolsl).

Shown graphically are the results of analysis of variant 1 depicting the presence and location of 10 transmembrane domains using the TMpred program (Figure 13b) and TMHMM program (Figure 13c). The results of each program, namely the amino acids encoding the transmembrane domains are summarized in Table L.

Example 10: Generation of 24P4C12 Polvclonal Antibodies Polyclonal antibodies can be raised in a mammal, for example, by one or more injections of an immunizing agent and, if desired, an adjuvant. Typically, the immunizing agent and/or adjuvant will be injected in the mammal by multiple subcutaneous or intraperitoneal injections. In addition to immunizing with the full length 24P4C12 protein, computer algorithms are employed n design of immunogens that, based on amino acid sequence analysis contain characteristics of being antigenic and available for recognition by the immune system of the immunized host (see the Example entitled "Antigenicity Profiles"). Such regions would be predicted to be hydrophilic, flexible, in beta-turn conformations, and be exposed on the surface of the protein (see, Figure 5, Figure 6, Figure 7, Figure 8, or Figure 9 for amino acid profiles that indicate such regions of 24P4C12 and variants).

For example, 24P4C12 recombinant bacterial fusion proteins or peptides containing hydrophilic, flexible, beta-turn regions of 24P4C12 variant proteins are used as antigens to generate polyclonal antibodies in New Zealand White rabbits.

For example, such regions include, but are not limited to, amino acids 1-34, amino acids 118-135, amino acids 194-224, amino acids 280-290, and amino acids 690-710, of 24P4C12 variants 1. It is useful to conjugate the immunizing agent to a protein known to be immunogenic in the mammal being immunized. Examples of such immunogenic proteins include, but are not limited to, keyhole limpet hemocyanin (KLH), serum albumin, bovine thyroglobulin, and soybean trypsin inhibitor. In one embodiment, a peptide encoding amino acids 1-14 of 24P4C12 variant 1 was conjugated to KLH and used to immunize a rabbit. This antiserum exhibited a high titer to the peptide (>10,000) and recognized 24P4C12 in transfected 293T cells by Western blot and flow cytometry (Figure 24) and in stable recombinant PC3 cells by Western blot and immunohistochemistry (Figure 25). Alternatively the immunizing agent may include all or portions of the 24P4C12 variant proteins, analogs or fusion proteins thereof. For example, the 24P4C12 variant 1 amino acid sequence can be fused using recombinant DNA techniques to any one of a variety of fusion protein partners that are well known in the art, such as glutathione-S-transferase (GST) and HIS tagged fusion proteins. Such fusion proteins are purified from induced bacteria using the appropriate affinity matrix.

In one embodiment, a GST-fusion protein encoding amino acids 379-453, encompassing the third predicted extracellular loop of variant 1, is produced, purified, and used as immunogen. Other recombinant bacterial fusion proteins that may he employed include maltose binding protein, LacZ, thioredoxin, NusA, or an immunoglobulin constant region (see the section entitled "Production of 24P4C12 in Prokaryotic Systems' and Current Protocols In Molecular Biology, Volume 2, Unit 16, Frederick M. Ausubul et al. eds., 1995; Linsley, Brady, Urnes, Grosmaire, Damle, and Ledbetter, L.(1991) J.Exp. Med. 174, 561-566).

In addition to bacterial derived fusion proteins, mammalian expressed protein antigens are also used. These antigens are expressed from mammalian expression vectors such as the Tag5 and Fc-fusion vectors (see the Example entitled "Production of Recombinant 24P4C12 in Eukaryotic Systems"), and retains post-translational modifications such as glycosylations found in native protein. In two embodiments, the predicted 1st and third extracellular loops of variant 1, amino acids 59-227 and 379453 respectively, were each cloned into the Tag5 mammalian secretion vector and expressed in 293T cells (Figure 26). Each recombinant protein is then purified by metal chelate chromatography from tissue culture WO 2004/050828 PCT/US2002/038264 supernatants andlor lysates of 293T cells stably expressing the recombinant vector. The purified Tag5 24P4C12 protein is then used as immunogen.

During the immunization protocol, it is useful to mix or emulsify the antigen in adjuvants that enhance the immune response of the host animal. Examples of adjuvants include, but are not limited to, complete Freund's adjuvant (CFA) and MPL-TDM adjuvant (monophosphoryl Lipid A, synthetic trehalose dicorynomycolate).

In a typical protocol, rabbits are initially immunized subcutaneously with up to 200 pg, typically 100-200 pg, of fusion protein or peptide conjugated to KLH mixed in complete Freund's adjuvant (CFA). Rabbits are then injected subcutaneously every two weeks with up to 200 gg, typically 100-200 g, of the immunogen in incomplete Freund's adjuvant (IFA). Test bleeds are taken approximately 7-10 days following each immunization and used to monitor the titer of the antiserum by ELISA.

To test reactivity and specificity of immune serum, such as the rabbit serum derived from immunization with a KLHconjugated peptide encoding amino acids 1-14 of variant 1, the full-length 24P4C12 variant 1 cDNA is cloned into pCDNA 3.1 myc-his or retroviral expression vectors (Invitrogen, see the Example entitled "Production of Recombinant 24P4C12 in Eukaryotic Systems"). After transfection of the constructs into 293T cells or transduction of PC3 with 24P4C12 retrovirus, cell lysates are probed with the anti-24P4C12 serum and with anti-His antibody (Santa Cruz Biotechnologies, Santa Cruz, CA) to determine specific reactivity to denatured 24P4C12 protein using the Westem blot technique. As shown in Figures 24 and 25 the antiserum specifically recognizes 24P4C12 protein in 293T and PC3 cells. In addition, the immune serum is tested by fluorescence microscopy, flow cytometry, and immunohistochemistry (Figure 25) and immunoprecipitation against 293T and other recombinant 24P4C12-expressing cells to determine specific recognition of native protein. Western blot, immunoprecipitation, fluorescent microscopy, immunohistochemistry and flow cytometric techniques using cells that endogenously express 24P4C12 are also carried out to test reactivity and specificity.

Anti-serum from rabbits immunized with 24P4C12 variant fusion proteins, such as GST and MBP fusion proteins, are purified by depletion of antibodies reactive to the fusion partner sequence by passage over an affinity column containing the fusion partner either alone or in the context of an irrelevant fusion protein. For example, antiserum derived from a GST- 24P4C12 fusion protein encoding amino acids 379-453 of variant 1 is first purified by passage over a column of GST protein covalently coupled to AffiGel matrix (BioRad, Hercules, Calif.). The antiserum is then affinity purified by passage over a column composed of a MBP-fusion protein also encoding amino acids 379-453 covalently coupled to Affigel matrix. The serum is then further purified by protein G affinity chromatography to isolate the IgG fraction. Sera from other His-tagged antigens and peptide immunized rabbits as well as fusion partner depleted sera are affinity purified by passage over a column matrix composed of the original protein immunogen or free peptide.

Example 11: Generation of 24P4C12 Monoclonal Antibodies (mAbs) In one embodiment, therapeutic mAbs to 24P4C12 variants comprise those that react with epitopes specific for each variant protein or specific to sequences in common between the variants that would disrupt or modulate the biological function of the 24P4C12 variants, for example those that would disrupt the interaction with ligands and substrates or disrupt its biological activity. Immunogens for generation of such mAbs include those designed to encode or contain the entire 24P4C12 protein variant sequence, regions of the 24P4C12 protein variants predicted to be antigenic from computer analysis of the amino acid sequence (see, Figure 5, Figure 6, Figure 7, Figure 8, or Figure 9, and the Example entitled "Antigenicity Profiles"). Immunogens include peptides, recombinant bacterial proteins, and mammalian expressed Tag proteins and human and murine IgG FC fusion proteins. In addition, cells engineered to express high levels of a respective 24P4C12 variant, such as 293T-24P4C12 variant 1 or 300.19-24P4C12 variant Imurine Pre-B cells, are used to immunize mice.

WO 2004/050828 PCT/US2002/038264 To generate mAbs to a 24P4C12 variant, mice are first immunized intraperitoneally (IP) with, typically, 10-50 pg of protein immunogen or 107 24P4C12-expressing cells mixed in complete Freund's adjuvant. Mice are then subsequently immunized IP every 2-4 weeks with, typically, 10-50 ig of protein immunogen or 107 cells mixed in incomplete Freund's adjuvant. Alternatively, MPL-TDM adjuvant is used in immunizations. In one embodiment, mice were immunized as above with 300.19-24P4C12 cells in complete and then incomplete Freund's adjuvant, and subsequently sacrificed and the spleens harvested and used for fusion and hybridoma generation. As is can be seen in Figure 27, 2 hybridomas were generated whose antibodies specifically recognize 24P4C12 protein expressed in 293T cells by flow cytometry. In addition to the above protein and cell-based immunization strategies, a DNA-based immunization protocol is employed in which a mammalian expression vector encoding a 24P4C12 variant sequence is used to immunize mice by direct injection of the plasmid DNA.

In one embodiment, a Tag5 mammalian secretion vector encoding amino acids 59-227 of the variant 1 sequence (Figure 26) was used to immunize mice. Subsequent booster immunizations are then carried out with the purified protein. In another example, the same amino acids are cloned into an Fc-fusion secretion vector in which the 24P4C12 variant 1 sequence is fused at the amino-terminus to an IgK leader sequence and at the carboxyl-terminus to the coding sequence of the human or murine IgG Fc region. This recombinant vector is then used as immunogen. The plasmid immunization protocols are used in combination with purified proteins as above and with cells expressing the respective 24P4C12 variant.

During the immunization protocol, test bleeds are taken 7-10 days following an injection to monitor titer and specificity of the immune response. Once appropriate reactivity and specificity is obtained as determined by ELISA, Western blotting, immunoprecipitation, fluorescence microscopy, immunohistochemistry, and flow cytometric analyses, fusion and hybridoma generation is then carried out with established procedures well known in the art (see, Harlow and Lane, 1988).

In one embodiment for generating 24P4C12 variant 8 specific monoclonal antibodies, a peptide encoding amino acids 643-654 (RNPITPTGHVFQ) (SEQ ID NO: 46) of 24P4C12 variant 8 is synthesized, coupled to KLH and used as immunogen. Balb C mice are initially immunized intraperitoneally with 25 pg of the KLH-24P4C12 variant 8 peptide mixed in complete Freund's adjuvant. Mice are subsequently immunized every two weeks with 25 pg of the antigen mixed in incomplete Freund's adjuvant for a total of three immunizations. ELISA using the free peptide determines the reactivity of serum from immunized mice. Reaclivity and specificity of serum to full length 24P4C12 variant 8 protein is monitored by Western blotting, immunoprecipitation and flow cytometry using 293T cells transfected with an expression vector encoding the 24P4C12 variant 8 cDNA compared to cells transfected with the other 24P4C12 variants (see the Example entitled "Production of Recombinant 24P4C12 in Eukaryotic Systems"). Other recombinant 24P4C12 variant 8-expressing cells or cells endogenously expressing 24P4C12 variant 8 are also used. Mice showing the strongest specific reactivity to 24P4C12 variant 8 are rested and given a final injection of antigen in PBS and then sacrificed four days later. The spleens of the sacrificed mice are harvested and fused to SPO/2 myeloma cells using standard procedures (Harlow and Lane, 1988).

Supernatants from HAT selected growth wells are screened by ELISA, Western blot, immunoprecipitation, fluorescent microscopy, and flow cytometry to identify 24P4C12 variant B-specific antbody-producing clones. A similar strategy is also used to derive 24P4C12 variant 9-specific antibodies using a peptide encompassing amino acids 379-388 (PLPTQPATLG) (SEQ ID NO: 47).

The binding affinity of a 24P4C12 monoclonal antibody is determined using standard technologies. Affinity measurements quantify the strength of antibody to epitope binding and are used to help define which 24P4C12 monoclonal antibodies preferred for diagnostic or therapeutic use, as appreciated by one of skill in the art. The BIAcore system (Uppsala, Sweden) is a preferred method for determining binding affinity. The BIAcore system uses surface plasmon resonance (SPR, Welford K. 1991, Opt. Quant. Elect. 23:1; Morton and Myszka, 1998, Methods in Enzymology 295: 268) to WO 2004/050828 PCT/US2002/038264 monitor bimolecular interactions in real time. BIAcore analysis conveniently generates association rate constants, dissociation rate constants, equilibrium dissociation constants, and affinity constants.

Example 12: HLA Class I and Class II Binding Assays HLA class I and class II binding assays using purified HLA molecules are performed in accordance with disclosed protocols PCT publications WO 94/20127 and WO 94/03205; Sidney et Current Protocols in Immunology 18.3.1 .(1998); Sidney, et al., J. Immunol. 154:247 (1995); Sette, et at., Mol. Immunol. 31:813 (1994)). Briefly, purified MHC molecules (5 to 500 nM) are incubated with various unlabeled peptide inhibitors and 1-10 nM 125 -radiolabeled probe peptides as described. Following incubation, MHC-peptide complexes are separated from free peptide by gel filtration and the fraction of peptide bound is determined. Typically, in preliminary experiments, each MHC preparation is titered in the presence of fixed amounts of radiolabeled peptides to determine the concentration of HLA molecules necessary to bind of the total radioactivity. All subsequent inhibition and direct binding assays are performed using these HLA concentrations.

Since under these conditions [label]<[HLA] and ICso[HLA), the measured ICso values are reasonable approximations of the true Ko values. Peptide inhibitors are typically tested at concentrations ranging from 120 ugml to 1.2 ng/ml, and are tested in two to four completely independent experiments. To allow comparison of the data obtained in different experiments, a relative binding figure is calculated for each peptide by dividing the ICso of a positive control for inhibition by the ICso for each tested peptide (typically unlabeled versions of the radiolabeled probe peptide). For database purposes, and inter-experiment comparisons, relative binding values are compiled. These values can subsequently be converted back into ICso nM values by dividing the ICso nM of the positive controls for inhibition by the relative binding of the peptide of interest. This method of data compilation is accurate and consistent for comparing peptides that have been tested on different days, or with different lots of purified MHC.

Binding assays as outlined above may be used to analyze HLA supermotif and/or HLA motif-bearing peptides (see Table IV).

Example 13: Identification of HLA Supermotif- and Motif-Bearing CTL Candidate Epitopes HLA vaccine compositions of the invention can include multiple epitopes. The multiple epitopes can comprise .multiple HLA supermotifs or motifs to achieve broad population coverage. This example illustrates the identification and confirmation of supermotif- and motif-bearing epitopes for the inclusion in such a vaccine composition. Calculation of population coverage is performed using the strategy described below.

Computer searches and algorithms for identification of supermotif and/or motif-bearing epitopes The searches performed to identify the motif-bearing peptide sequences in the Example entitled "Antigenicity Profiles" and Tables VIII-XXI and XXII-XLIXemploy the protein sequence data from the gene product of 24P4C12 set forth in Figures 2 and 3, the specific search peptides used to generate the tables are listed in Table VII.

Computer searches for epitopes bearing HLA Class I or Class II supermotifs or motifs are performed as follows, 'All translated 24P4C12 protein sequences are analyzed using a text string search software program to identify potential peptide sequences containing appropriate HLA binding motifs; such programs are readily produced in accordance with information in the art in view of known motif/supermotif disclosures. Furthermore, such calculations can be made mentally.

Identified A2-, A3-, and DR-supermotif sequences are scored using polynomial algorithms to predict their capacity to bind to specific HLA-Class I or Class II molecules. These polynomial algorithms account for the impact of different amino acids at different positions, and are essentially based on the premise that the overall affinity (or AG) of peptide-HLA molecule interactions can be approximated as a linear polynomial function of the type: WO 2004/050828 PCT/US2002/038264 "AG" au 2 x ai x ai where a i is a coefficient which represents the effect of the presence of a given amino acid at a given position (i) along the sequence of a peptide of n amino acids. The crucial assumption of this method is that the effects at each position are essentially independent of each other independent binding of individual side-chains). When residue j occurs at position i in the peptide, it is assumed to contribute a constant amount j; to the free energy of binding of the peptide irrespective of the sequence of the rest of the peptide.

The method of derivation of specific algorithm coefficients has been described in Gulukota et al, J. Mol. Bid.

267:1258-126, 1997; (see also Sidney et al, Human Immunol. 45:79-93, 1996; and Southwood et al., J. Immuno. 160:3363- 3373, 1998). Briefly, for all i positions, anchor and non-anchor alike, the geometric mean of the average relative binding (ARB) of all peptides carrying j is calculated relative to the remainder of the group, and used as the estimate of j. For Class II peptides, if multiple alignments are possible, only the highest scoring alignment is utilized, following an iterative procedure.

To calculate an algorithm score of a given peptide in a test set, the ARB values corresponding to the sequence of the peptide are multiplied. If this product exceeds a chosen threshold, the peptide is predicted to bind. Appropriate thresholds are chosen as a function of the degree of stringency of prediction desired.

Selection of HLA-A2 supertype cross-reactive peptides Protein sequences from 24P4C12 are scanned utilizing motif identification software, to identify 9- 10- and 11mer sequences containing the HLA-A2-supermotif main anchor specificity. Typically, these sequences are then scored using the protocol described above and the peptides corresponding to the positive-scoring sequences are synthesized and tested for their capacity to bind purified HLA-A*0201 molecules in vitro (HLA-A0201 is considered a prototype A2 supertype molecule).

These peptides are then tested for the capacity to bind to additional A2-supertype molecules (A*0202, A*0203, A*0206, and A*6802). Peptides that bind to at least three of the five A2-supertype alleles tested are typically deemed A2supertype cross-reactive binders. Preferred peptides bind at an affinity equal to or less than 500 nM to three or more HLA- A2 supertype molecules.

Selection of HLA-A3 supermotif-bearing epitopes The 24P4C12 protein sequence(s) scanned above is also examined for the presence of peptides with the HLA-A3supermotif primary anchors. Peptides corresponding to the HLA A3 supermotif-bearing sequences are then synthesized and tested for binding to HLA-A*0301 and HLA-A*1101 molecules, the molecules encoded by the two most prevalent A3supertype alleles. The peptides that bind at least one of the two alleles with binding affinities of 5500 nM, often 200 nM, are then tested for binding cross-reactivity to the other common A3-supertype alleles A*3101, A*3301, and A*6801) to identify those that can bind at least three of the five HLA-A3-supertype molecules tested.

Selection of HLA-B7 supermotif bearing epitopes The 24P4C12 protein(s) scanned above is also analyzed for the presence of 9-10-, or 11-mer peptides with the HLA-B7-supermotif. Corresponding peptides are synthesized and tested for binding to HLA-B*0702, the molecule encoded by the most common B7-supertype allele the prototype B7 supertype allele). Peptides binding B*0702 with ICso of _500 nM are identified using standard methods. These peptides are then tested for binding to other common B7-supertype molecules B*3501, B*5101, B*5301, and 8*5401). Peptides capable of binding to three or more of the five 87supertype alleles tested are thereby identified.

WO 2004/050828 PCT/US2002/038264 Selection of Al and A24 motif-bearing epitopes To further increase population coverage, HLA-A1 and -A24 epitopes can also be incorporated into vaccine compositions. An analysis of the 24P4C12 protein can also be performed to identify HLA-A1- and A24-motif-containing sequences.

High affinity and/or cross-reactive binding epitopes that bear other motif and/or supermotifs are identified using analogous methodology.

Example 14: Confirmation of Immunogenicity Cross-reactive candidate CTL A2-supermotif-bearing peptides that are identified as described herein are selected to confirm in vitro immunogenicity. Confirmation is performed using the following methodology: Target Cell Lines for Cellular Screening: The .221A2.1 cell line, produced by transferring the HLA-A2.1 gene into the HLA-A, -C null mutant human Blymphoblastoid cell line 721.221, is used as the peptide-loaded target to measure activity of HLA-A2.1-restricted CTL. This cell line is grown in RPMI-1640 medium supplemented with antibiotics, sodium pyruvate, nonessential amino acids and heat inactivated FCS. Cells that express an antigen of interest, or transfectants comprising the gene encoding the antigen of interest, can be used as target cells to confirm the ability of peptide-specific CTLs to recognize endogenous antigen.

Primary CTL Induction Cultures: Generation of Dendritic Cells PBMCs are thawed in RPMI with 30 uigml DNAse, washed twice and resuspended in complete medium (RPMI-1640 plus 5% AB human serum, non-essential amino acids, sodium pyruvate, Lglutamine and penicillin/streptomycin). The monccytes are purified by plating 10 x 10 6 PBMC/well in a 6-well plate. After 2 hours at 37°C, the non-adherent cells are removed by gently shaking the plates and aspirating the supematants. The wells are washed a total of three times with 3 ml RPMI to remove most of the non-adherent and loosely adherent cells. Three ml of complete medium containing 50 ng/ml of GM-CSF and 1,000 U/ml of IL-4 are then added to each well. TNFa is added to the DCs on day 6 at 75 nglml and the cells are used for CTL induction cultures on day 7.

Induction of CTL with DC and Peptide: CD8+ T-cells are isolated by positive selection with Dynal immunomagnetic beads (Dynabeads® M-450) and the detacha-bead® reagent. Typically about 200-250x10 6 PBMC are processed to obtain 24x10 6 CD8* T-cells (enough for a 48-well plate culture). Briefly, the PBMCs are thawed in RPMI with 30pglml DNAse, washed once with PBS containing 1% human AB serum and resuspended in PBS/1% AB serum at a concentration of 20x10 6 cells/ml. The magnetic beads are washed 3 times with PBS/AB serum, added to the cells (140pl beads/20x10 6 cells) and incubated for 1 hour at 4 0 C with continuous mixing. The beads and cells are washed 4x with PBS/AB serum to remove the nonadherent cells and resuspended at 100x10 6 cells/ml (based on the original cell number) in PBS/AB serum containing 100pl/ml detacha-bead® reagent and 30 pg/ml DNAse. The mixture is incubated for 1 hour at room temperature with continuous mixing. The beads are washed again with PBS/AB/DNAse to collect the CD8+ T-cells. The DC are collected and centrifuged at 1300 rpm for 5-7 minutes, washed once with PBS with 1% BSA, counted and pulsed with 40pg/ml of peptde at a cell concentration of 1-2x106ml in the presence of 3pg/ml 12- microglobulin for 4 hours at 20 0 C. The DC are then irradiated (4,200 rads), washed 1 time with medium and counted again.

Setting up induction cultures: 0.25 ml cytokine-generated DC (at 1x10 5 cells/ml) are co-cultured with 0.25ml of CD8+ T-cells (at 2x106 cell/mi) in each well of a 48-well plate in the presence of 10 ng/ml of IL-7. Recombinant human is added the next day at a final concentration of 10 ng/ml and rhuman IL-2 is added 48 hours later at 10 IUfml.

Restimulation of the induction cultures with peptide-pulsed adherent cells: Seven and fourteen days after the primary induction, the cells are restimulated with peptide-pulsed adherent cells. The PBMCs are thawed and washed twice WO 2004/050828 PCT/US2002/038264 with RPMI and DNAse. The cells are resuspended at 5x10 6 cells/ml and irradiated at -4200 rads. The PBMCs are plated at 2x106 in 0.5 ml complete medium per well and incubated for 2 hours at 37°C. The plates are washed twice with RPMI by tapping the plate gently to remove the nonadherent cells and the adherent cells pulsed with 10pg/ml of peptide in the presence of 3 pg/ml P12 microglobulin in 0.25ml RPMI/5%AB per well for 2 hours at 37°C. Peptide solution from each well is aspirated and the wells are washed once with RPMI. Most of the media is aspirated from the induction cultures (CD8+ cells) and brought to 0.5 ml with fresh media. The cells are then transferred to the wells containing the peptide-pulsed adherent cells. Twenty four hours later recombinant human IL-10 is added at a final concentration of 10 ng/ml and recombinant human 112 is added the next day and again 2-3 days later at 501U/ml (Tsai et al, Critical Reviews in Immunology 18(1-2):65-75, 1998). Seven days later, the cultures are assayed for CTL activity in a slCr release assay. In some experiments the cultures are assayed for peptide-specific recognition in the in situ IFNy ELISA at the time of the second restimulation followed by assay of endogenous recognition 7 days later. After expansion, activity is measured in both assays for a side-by-side comparison.

Measurement of CTL Ivtic activity by 5 'Cr release.

Seven days after the second restimulation, cytotoxicity is determined In a standard (5 hr) 51 Cr release assay by assaying individual wells at a single E:T. Peptide-pulsed targets are prepared by incubating the cells with 10pg/ml peptide overnight at 370C.

Adherent target cells are removed from culture flasks with trypsin-EDTA. Target cells are labeled with 200pCi of 1 Cr sodium chromate (Dupont, Wilmington, DE) for 1 hour at 37 0 C. Labeled target cells are resuspended at 106 per ml and diluted 1:10 with K562 cells at a concentration of 3.3x10 6 /ml (an NK-sensitive erythroblastoma cell line used to reduce nonspecific lysis). Target cells (100 pl) and effectors (100pl) are plated in 96 well round-bottom plates and incubated for 5 hours at 37°C. At that time, 100 pi of supematant are collected from each well and percent lysis is determined according to the formula: [(cpm of the test sample- cpm of the spontaneous 51 Cr release sample)/(cpm of the maximal 6 Cr release samplecpm of the spontaneous siCr release sample)] x 100.

Maximum and spontaneous release are determined by incubating the labeled targets with 1% Triton X-100 and media alone, respectively. A positive culture is,defined as one in which the specific lysis (sample- background) is 10% or higher in the case of individual wells and is 15% or more at the two highest E:T ratios when expanded cultures are assayed.

In situ Measurement of Human IFNy Production as an Indicator of Peptide-specific and Endogenous Recognition Immulon 2 plates are coated with mouse anti-human IFNy monoclonal antibody (4 igiml 0.1M NaHCO3, pH8.2) ovemight at 4°C. The plates are washed with Ca 2 Mg 2 -free PBS/0.05% Tween 20 and blocked with PBS/10% FCS for two hours, after which the CTLs (100 p/well) and targets (100 pl/well) are added to each well, leaving empty wells for the standards and blanks (which received media only). The target cells, either peptide-pulsed or endogenous targets, are used at a concentration of 1x106 cells/ml. The plates are incubated for 48 hours at 37°C with 5% C02.

Recombinant human IFN-gamma is added to the standard wells starting at 400 pg or 1200pg/100 microliterlwell and the plate incubated for two hours at 37"C. The plates are washed and 100 pl of biotinylated mouse anti-human IFNgamma monoclonal antibody (2 microgramlml in PBS/3%FCS/0.05% Tween 20) are added and incubated for 2 hours at room temperature. After washing again, 100 microliter HRP-streptavidin (1:4000) are added and the plates incubated for one hour at room temperature. The plates are then washed 6x with wash buffer, 100 microliter/well developing solution (TMB 1:1) are added, and the plates allowed to develop for 5-15 minutes. The reaction is stopped with 50 microlitertwell 1M H3PO 4 and read at OD450. A culture is considered positive if it measured at least 50 pg of IFN-gamma/well above background and is twice the background level of expression.

WO 2004/050828 PCT/US2002/038264 CTL Expansion.

Those cultures that demonstrate specific lytic activity against peptide-pulsed targets andlor tumor targets are expanded over a two week period with anti-CD3. Briefly, 5x10 4 CD8+ cells are added to a T25 flask containing the following: 1x10 6 irradiated (4,200 rad) PBMC (autologous or allogeneic) per ml, 2x10 5 irradiated (8,000 rad) EBV- transformed cells per ml, and OKT3 (anti-CD3) at 30ng per ml in RPMI-1640 containing 10% human AB serum, non-essential amino acids, sodium pyruvate, 25pM 2-mercaptoethanol, L-glutamine and penicillinlstreptomycin. Recombinant human IL2 is added 24 hours later at a final concentration of 2001U/ml and every three days thereafter with fresh media at 501U/ml. The cells are split if the cell concentration exceeds 1x106/ml and the cultures are assayed between days 13 and 15 at E:T ratios of 30, 3 and 1:1 in the 51 Cr release assay or at 1x10 6 /ml in the in situ IFNy assay using the same targets as before the expansion.

Cultures are expanded in the absence of anti-CD3- as follows. Those cultures that demonstrate specific lytic activity against peptide and endogenous targets are selected and 5x10 4 CD8* cells are added to a T25 flask containing the ,following: 1x10 6 autologous PBMC per ml which have been peptide-pulsed with 10 pg/ml peptide for two hours at 37°C and irradiated (4,200 rad); 2x105 irradiated (8,000 rad) EBV-transformed cells per ml RPMI-1640 containing 10%(v/v) human AB serum, non-essential AA, sodium pyruvate, 25mM 2-ME, L-glutamine and gentamicin.

Immunogenicity ofA2 supermotif-bearing peptides A2-supermotif cross-reactive binding peptides are tested in the cellular assay for the ability to induce peptidespecific CTL in normal individuals. In this analysis, a peptide is typically considered to be an epitope if it induces peptidespecific CTLs in at least individuals, and preferably, also recognizes the endogenously expressed peptide.

Immunogenicity can also be confirmed using PBMCs isolated from patients bearing a tumor that expresses 24P4C12. Briefly, PBMCs are isolated from patients, re-stimulated with peptide-pulsed monocytes and assayed for the ability to recognize peptide-pulsed target cells as well as transfected cells endogenously expressing the antigen.

Evaluation of A*03/A11 immunoqenicity HLA-A3 supermotif-bearing cross-reactive binding peptides are also evaluated for immunogenicity using methodology analogous for that used to evaluate the immunogenidty of the HLA-A2 supermotif peptides.

Evaluation of B7 immunocenicity Immunogenicity screening of the B7-supertype cross-reactive binding peptides identified as set forth herein are confirmed in a manner analogous to the confirmation of A2-and A3-supermotif-bearing peptides.

Peptides bearing other supermotifsmotifs, HLA-A1, HLA-A24 etc. are also confirmed using similar methodology Example 15: Implementation of the Extended Suoermotif to Improve the Binding Capacity of Native Epitopes by Creating Analogs HLA motifs and supermotifs (comprising primary and/or secondary residues) are useful in the identification and preparation of highly cross-reactive native peptides, as demonstrated herein. Moreover, the definition of HLA motifs and supermotifs also allows one to engineer highly cross-reactive epitopes by identifying residues within a native peptide sequence which can be analoged to confer upon the peptide certain characteristics, e.g. greater cross-reactivity within the group of HLA molecules that comprise a supertype, and/or greater binding affinity for some or all of those HLA molecules.

Examples of analoging peptides to exhibit modulated binding affinity are set forth in this example.

Analoaina at Primary Anchor Residues Peptide engineering strategies are implemented to further increase the cross-reactivity of the epitopes. For example, the main anchors of A2-supermotif-bearing peptides are altered, for example, to introduce a preferred L, I, V, or M at position 2, and I or V at the C-terminus.

WO 2004/050828 PCT/US2002/038264 To analyze the cross-reactivity of the analog peptides, each engineered analog is initially tested for binding to the prototype A2 supertype allele A*0201, then, if A*0201 binding capacity is maintained, for A2-supertype cross-reactivity.

Alternatively, a peptide is confirmed as binding one or all supertype members and then analoged to modulate binding affinity to any one (or more) of the supertype members to add population coverage.

The selection of analogs for immunogenicity in a cellular screening analysis is typically further restricted by the capacity of the parent wild type (WT) peptide to bind at least weakly, bind at an ICso of 5000nM or less, to three of more A2 supertype alleles. The rationale for this requirement is that the WT peptides must be present endogenously in sufficient quantity to be biologically relevant. Analoged peptides have been shown to have increased immunogenicity and crossreactivity by T cells specific for the parent epitope (see, Parkhurst et al., J. Immunol. 157:2539, 1996; and Pogue et at., Proc. Natl. Acad. Sci. USA 92:8166, 1995).

In the cellular screening of these peptide analogs, it is important to confirm that analog-specific CTLs are also able to recognize the wild-type peptide and, when possible, target cells that endogenously express the epitope.

Analoging of HLA-A3 and B7-supermotif-bearino Deotides Analogs of HLA-A3 supermotif-bearing epitopes are generated using strategies similar to those employed in analoging HLA-A2 supermotif-bearing peptides. For example, peptides binding to 3/5 of the A3-supertype molecules are engineered at primary anchor residues to possess a preferred residue S, M, or A) at position 2.

The analog peptides are then tested for the ability to bind A*03 and A*11 (prototype A3 supertype alleles). Those peptides that demonstrate 500 nM binding capacity are then confirmed as having A3-supertype cross-reactivity.

Similarly to the A2- and A3- motif bearing peptides, peptides binding 3 or more B7-supertype alleles can be improved, where possible, to achieve increased cross-reactive binding or greater binding affinity or binding half life. 87 supermotif-bearing peptides are, for example, engineered to possess a preferred residue I, L, or F) at the C-terminal primary anchor position, as demonstrated by Sidney etal. Immunol. 157:3480-3490, 1996).

Analoging at primary anchor residues of other motif and/or supermotif-bearing epitopes is performed in a like manner.

The analog peptides are then be confirmed for immunogenicity, typically in a cellular screening assay. Again, it is generally important to demonstrate that analog-specific CTLs are also able to recognize the wild-type peptide and, when possible, targets that endogenously express the epitope.

Analoqing at Secondary Anchor Residues Moreover, HLA supermotifs are of value in engineering highly cross-reactive peptides and/or peptides that bind HLA molecules with increased affinity by identifying particular residues at secondary anchor positions that are associated with such properties. For example, the binding capacity of a 87 supermotif-bearing peptide with an F residue at position 1 is analyzed. The peptide is then analoged to, for example, substitute L for F at position 1. The analoged peptide is evaluated for increased binding affinity, binding half life and/or increased cross-reactivity. Such a procedure identifies analoged peptides with enhanced properties.

Engineered analogs with sufficiently improved binding capacity or cross-reactivity can also be tested for immunogenicity in HLA-B7-transgenic mice, following for example, IFA immunization or lipopeptide immunization. Analoged peptides are additionally tested for the ability to stimulate a recall response using PBMC from patients with 24P4C12expressing tumors.

Other analoging strategies WO 2004/050828 PCT/US2002/038264 Another form of peptide analoging, unrelated to anchor positions, involves the substitution of a cysteine with aamino butyric acid. Due to its chemical nature, cysteine has the propensity to form disulfide bridges and sufficiently alter the peptide structurally so as to reduce binding capacity. Substitution of a-amino butyric acid for cysteine not only alleviates this problem, but has been shown to improve binding and crossbinding capabilities in some instances (see, the review by Sette et al., In: Persistent Viral Infections, Eds. R. Ahmed and I. Chen, John Wiley Sons, England, 1999).

Thus, by the use of single amino acid substitutions, the binding properties and/or cross-reactivity of peptide ligands for HLA supertype molecules can be modulated.

Example 16: Identification and confirmation of 24P4C12-derived sequences with HLA-DR binding motifs Peptide epitopes bearing an HLA class II supermotif or motif are identified and confirmed as outlined below using methodology similar to that described for HLA Class I peptides.

Selection of HLA-DR-supermotif-bearinQ epitopes.

To identify 24P4C12-derived, HLA class II HTL epitopes, a 24P4C12 antigen is analyzed for the presence of sequences bearing an HLA-DR-motif or supermotif. Specifically, 15-mer sequences are selected comprising a DRsupermotif, comprising a 9-mer core, and three-residue N- and C-terminal flanking regions (15 amino adds total).

Protocols for predicting peptide binding to DR molecules have been developed (Southwood et J. Immunol.

160:3363-3373, 1998). These protocols, specific for individual DR molecules, allow the scoring, and ranking, of 9-mer core regions. Each protocol not only scores peptide sequences for the presence of DR-supermotif primary anchors at position 1 and position 6) within a 9-mer core, but additionally evaluates sequences for the presence of secondary anchors.

Using allele-specific selection tables (see, Southwood et aL, ibid.), it has been found that these protocols efficiently select peptide sequences with a high probability of binding a particular OR molecule. Additionally, it has been found that performing these protocols in tandem, specifically those for DR1, DR4w4, and DR7, can efficiently select DR cross-reactive peptides.

The 24P4C12-derived peptides identified above are tested for their binding capacity for various common HLA-DR molecules. All peptides are initially tested for binding to the DR molecules in the primary panel: DR1, DR4w4, and DR7.

Peptides binding at least two of these three DR molecules are then tested for binding to DR2w2 p1, DR2w2 p2, DR6w19, and DR9 molecules in secondary assays. Finally, peptides binding at least two of the four secondary panel DR molecules, and thus cumulatively at least four of seven different DR molecules, are screened for binding to DR4w15, DR5w1 1, and DR8w2 molecules in tertiary assays. Peptides binding at least seven of the ten DR molecules comprising the primary, secondary, and tertiary screening assays are considered cross-reactive DR binders. 24P4C12-derived peptides found to bind. common HLA-DR alleles are of particular interest.

Selection of DR3 motif petides Because HLA-DR3 is an allele that is prevalent in Caucasian, Black, and Hispanic populations, DR3 binding capacity is a relevant criterion in the selection of HTL epitopes. Thus, peptides shown to be candidates may also be assayed for their DR3 binding capacity. However, in view of the binding specificity of the DR3 motif, peptides binding only to SDR3 can also be considered as candidates for inclusion in a vaccine formulation.

To efficiently identify peptides that bind DR3, target 24P4C12 antigens are analyzed for sequences carrying one of the two DR3-specific binding motifs reported by Geluk etal. Immunol. 152:5742-5748,1994). The corresponding peptides are then synthesized and confirmed as having the ability to bind DR3 with an affinity of 1 pM or better, less than 1 pM. Peptides are found that meet this binding criterion and qualify as HLA class II high affinity binders.

DR3 binding epitopes identified in this manner are included in vaccine compositions with DR supermotif-bearing peptide epitopes.

WO 2004/050828 PCT/US20021038264 Similarly to the case of HLA class I motif-bearing peptides, the class II motif-bearing peptides are analoged to improve affinity or cross-reactivity. For example, aspartic acid at position 4 of the 9-mer core sequence is an optimal residue for DR3 binding, and substitution for that residue often improves DR 3 binding. Example 17: Immunoaenicitv of 24P4C12-derived HTL epitopes This example determines immunogenic DR supermotif- and DR3 motif-bearing epitopes among those identified using the methodology set forth herein.

Immunogenicity of HTL epitopes are confirmed in a manner analogous to the determination of immunogenicity of CTL epitopes, by assessing the ability to stimulate HTL responses and/or by using appropriate transgenic mouse models.

Immunogenicity is determined by screening for: in vitro primary induction using normal PBMC or recall responses from patients who have 24P4C12-expressing tumors.

Example 18: Calculation of phenotypic frequencies of HLA-supertypes in various ethnic backgrounds to determine breadth of population coverage This example illustrates the assessment of the breadth of population coverage of a vaccine composition comprised of multiple epitopes comprising multiple supermotifs and/or motifs.

In order to analyze population coverage, gene frequencies of HLA alleles are determined. Gene frequencies for each HLA allele are calculated from antigen or allele frequencies utilizing the binomial distribution formulae gf=1-(SQRT(1af)) (see, Sidney et Human Immuno. 45:79-93, 1996). To obtain overall phenotypic frequencies, cumulative gene frequencies are calculated, and the cumulative antigen frequencies derived by the use of the inverse formula [af=1-(1-Cgf)2].

Where frequency data is not available at the level of DNA typing, correspondence to the serologically defined antigen frequencies is assumed. To obtain total potential supertype population coverage no linkage disequilibrium is assumed, and only alleles confirmed to belong to each of the supertypes are included (minimal estimates). Estimates of total potential coverage achieved by inter-loci combinations are made by adding to the A coverage the proportion of the non-A covered population that could be expected to be covered by the B alleles considered total=A+B*(1-A)). Confirmed members of the A3-like supertype are A3, All11, A31, A'3301, and A*6801. Although the A3-like supertype may also include A34, A66, and A*7401, these alleles were not included in overall frequency calculations. Likewise, confirmed members of the A2-like supertype family are A*0201, A*0202, A*0203, A*0204, A*0205, A*0206, A*0207, A*6802, and A*6901. Finally, the B7-like supertype-confirmed alleles are: B7, B*3501-03, B51, B*5301, B*5401, B*5501-2, B*5601, 8*6701, and B*7801 (potentially also B*1401, B*3504-06, B*4201, and B'5602).

Population coverage achieved by combining the A2-, A3- and B7-supertypes is approximately 86% in five major ethnic groups. Coverage may be extended by including peptides bearing the Al and A24 motifs. On average, Al is present in 12% and A24 in 29% of the population across five different major ethnic groups (Caucasian, North American Black, Chinese, Japanese, and Hispanic). Together, these alleles are represented with an average frequency of 39% in these same ethnic populations. The total coverage across the major ethnicities when Al and A24 are combined with the coverage of the A2-, A3- and B7-supertype alleles is see, Table IV An analogous approach can be used to estimate population coverage achieved with combinations of class II motif-bearing epitopes.

Immunogenicity studies in humans Bertoni et J. Clin. Invest. 100:503, 1997; Doolan et al, Immunity 7:97, 1997; and Threlkeld et al., J. Immunol. 159:1648, 1997) have shown that highly cross-reactive binding peptides are almost always recognized as epitopes. The use of highly cross-reactive binding peptides is an important selection criterion in identifying candidate epitopes for inclusion in a vaccine that is immunogenic in a diverse population.

WO 2004/050828 PCT/US2002/038264 With a sufficient number of epitopes (as disclosed herein and from the art), an average population coverage is predicted to be greater than 95% in each of five major ethnic populations. The game theory Monte Carlo simulation analysis, which is known in the art (see eg., Osborne, M.J. and Rubinstein, A. "Acourse in game theory" MIT Press, 1994), can be used to estimate what percentage of the individuals in a population comprised of the Caucasian, North American Black, Japanese, Chinese, and Hispanic ethnic groups would recognize the vaccine epitopes described herein. A preferred percentage is 90%. A more preferred percentage is Example 19: CTL Recognition Of Endogenously Processed Antigens After Priming This example confirms that CTL induced by native or analoged peptide epitopes identified and selected as described herein recognize endogenously synthesized, native antigens.

Effector cells isolated from transgenic mice that are immunized with peptide epitopes, for example HLA-A2 supermotif-bearing epitopes, are re-stimulated in vitro using peptide-coated stimulator cells. Six days later, effector cells are assayed for cytotoxicity and the cell lines that contain peptide-specific cytotoxic activity are further re-stimulated. An additional six days later, these cell lines are tested for cytotoxic activity on 51 Cr labeled Jurkat-A2.1/Kb target cells in the absence or presence of peptide, and also tested on 51Cr labeled target cells bearing the endogenously synthesized antigen, i.e. cells that are stably transfected with 24P4C12 expression vectors.

The results demonstrate that CTL lines obtained from animals primed with peptide epitope recognize endogenously synthesized 24P4C12 antigen. The choice of transgenic mouse model to be used for such an analysis depends upon the epitope(s) that are being evaluated. In addition to HLA-A*0201/Kb transgenic mice, several other transgenic mouse models including mice with human A 1, which may also be used to evaluate A3 epitopes, and B7 alleles have been characterized and others transgenic mice for HLA-A1 and A24) are being developed. HLA-DR1 and HLA- DR3 mouse models have also been developed, which may be used to evaluate HTL epitopes.

Example 20: Activity Of CTL-HTL Conjugated Epitopes In Transgenic Mice This example illustrates the induction of CTLs and HTLs in transgenic mice, by use of a 24P4C12-derived CTL and HTL peptide vaccine compositions. The vaccine composition used herein comprise peptides to be administered to a patient with a 24P4C12-expressing tumor. The peptide composition can comprise multiple CTL andlor HTL epitopes. The epitopes are identified using methodology as described herein. This example also illustrates that enhanced immunogenicity can be achieved by inclusion of one or more HTL epitopes in a CTL vaccine composition; such a peptide composition can comprise an HTL epitope conjugated to a CTL epitope. The CTL epitope can be one that binds to multiple HLA family members at an affinity of 500 nM or less, or analogs of that epitope. The peptides may be lipidated, if desired.

Immunization procedures: Immunization of transgenic mice is performed as described (Alexander et al., J.

Immunol. 159:4753-4761, 1997). For example, A2/Kb mice, which are transgenic for the human HLA A2.1 allele and are used to confirm the immunogenicity of HLA-A*0201 motif- or HLA-A2 supermotif-bearing epitopes, and are primed subcutaneously (base of the tail) with a 0,1 ml of peptide in Incomplete Freund's Adjuvant, or if the peptide composition is a lipidated CTUHTL conjugate, in DMSO/saline, or if the peptide composition is a polypeptide, in PBS or Incomplete Freund's Adjuvant. Seven days after priming, splenocytes obtained from these animals are restimulated with syngenic irradiated LPSactivated lymphoblasts coated with peptide.

Cell lines: Target cells for peptide-specific cytotoxicity assays are Jurkat cells transfected with the HLA-A2.1/Kb chimeric gene Vitiello et al, J. Exp. Med. 173:1007, 1991) WO 2004/050828 PCT/US2002/038264 In vitro CTL activation: One week after priming, spleen cells (30x10 6 cells/flask) are co-cuitured at 37'C with syngeneic, irradiated (3000 rads), peptide coated lymphoblasts (10x10 6 cells/flask) in 10 ml of culture mediumlT25 flask.

After six days, effector cells are harvested and assayed for cytotoxic activity.

Assay for cytotoxic activity: Target cells (1.0 to 1.5x106) are incubated at 37"C in the presence of 200 pl of 51 Cr.

After 60 minutes, cells are washed three times and resuspended in R10 medium. Peptide is added where required at a concentration of 1 Jg/ml. For the assay, 104 s 5 Cr-labeled target cells are added to different concentrations of effector cells (final volume of 200 pl) in U-bottom 96-well plates. After a six hour incubation period at 37°C, a 0.1 ml aliquot of supernatant is removed from each well and radioactivity is determined in a Micromedic automatic gamma counter. The percent specific lysis is determined by the formula: percent specific release 100 x (experimental release spontaneous release)/(maximum release spontaneous release). To facilitate comparison between separate CTL assays run under the same conditions, S'Cr release data is expressed as lytic units/106 cells. One lytic unit is arbitrarily defined as the number of effector cells required to achieve 30% lysis of 10,000 target cells in a six hour 5 1 Cr release assay. To obtain specific lytic units/106, the lytic units/10 6 obtained in the absence of peptide is subtracted from the lytic units/10 6 obtained in the presence of peptide. For example, if 30% 51 Cr release is obtained at the effector target ratio of 50:1 5x105 effector cells for 10,000 targets) in the absence of peptide and 5:1 5x10 4 effector cells for 10,000 targets) in the presence of peptide, the specific lytic units would be: [(1/50,000)-(1/500,000)] x 106 18 LU.

The results are analyzed to assess the magnitude of the CTL responses of animals injected with the immunogenic CTL/HTL conjugate vaccine preparation and are compared to the magnitude of the CTL response achieved using, for example, CTL epitopes as outlined above in the Example entitled "Confirmation of Immunogenicity." Analyses similar to this may be performed to confirm the immunogenicity of peptide conjugates containing multiple CTL epitopes and/or multiple HTL epitopes. In accordance with these procedures, it is found that a CTL response is induced, and concomitantly that an HTL response is induced upon administration of such compositions.

Example 21: Selection of CTL and HTL epitopes for inclusion in a 24P4C12-specific vaccine.

This example illustrates a procedure for selecting peptide epitopes for vaccine compositions of the invention. The peptides in the composition can be in the form of a nucleic acid sequence, either single or one or more sequences minigene) that encodes peptide(s), or can be single and/or polyepitopic peptides.

The following principles are utilized when selecting a plurality of epitopes for inclusion in a vaccine composition.

Each of the following principles is balanced in order to make the selection.

Epitopes are selected which, upon administration, mimic immune responses that are correlated with 24P4C12 clearance. The number of epitopes used depends on observations of patients who spontaneously clear 24P4C12. For example, if it has boon observed that patients who spontaneously dear 24P4C12-expressing cells generate an immune response to at least three epitopes from 24P4C12 antigen, then at least three epitopes should be included for HLA class I. A similar rationale is used to determine HLA class II epitopes.

Epitopes are often selected that have a binding affinity of an ICso of 500 nM or less for an HLA class I molecule, or for class II, an IC5o of 1000 nM or less; or HLA Class I peptides with high binding scores from the BIMAS web site, at URL bimas.dcrt.nih.govl.

In order to achieve broad coverage of the vaccine through out a diverse population, sufficient supermotif bearing peptides, or a sufficient array of allele-specific motif bearing peptides, are selected to give broad population coverage. In one embodiment, epitopes are selected to provide at least 80% population coverage. A Monte Carlo analysis, a statistical evaluation known in the art, can be employed to assess breadth, or redundancy, of population coverage.

WO 2004/050828 PCT/US2002/038264 When creating polyepitopic compositions, or a minigene that encodes same, it is typically desirable to generate the smallest peptide possible that encompasses the epitopes of interest. The principles employed are similar, if not the same, as those employed when selecting a peptide comprising nested epitopes. For example, a protein sequence for the vaccine composition is selected because it has maximal number of epitopes contained within the sequence, it has a high concentration of epitopes. Epitopes may be nested or overlapping frame shifted relative to one another). For example, with overlapping epitopes, two 9-mer epitopes and one 10-mer epitope can be present in a 10 amino acid peptide. Each epitope can be exposed and bound by an HLA molecule upon administration of such a peptide. A multi-epitopic, peptide can be generated synthetically, recombinantly, or via cleavage from the native source. Alternatively, an analog can be made of this native sequence, whereby one or more of the epitopes comprise substitutions that alter the cross-reactivity and/or binding affinity properties of the polyepitopic peptide. Such a vaccine composition is administered for therapeutc or prophylactic purposes. This embodiment provides for the possibility that an as yet undiscovered aspect of immune system processing will apply to the native nested sequence and thereby facilitate the production of therapeutic or prophylactic immune response-inducing vaccine compositions. Additionally such an embodiment provides for the possibility of motifbearing epitopes for an HLA makeup that is presently unknown. Furthermore, this embodiment (absent the creating of any analogs) directs the immune response to multiple peptide sequences that are actually present in 24P4C12, thus avoiding the need to evaluate any junctional epitopes. Lastly, the embodiment provides an economy of scale when producing nucleic acid vaccine compositions. Related to this embodiment, computer programs can be derived in accordance with principles in the art, which identify in a target sequence, the greatest number of epitopes per sequence length.

A vaccine composition comprised of selected peptides, when administered, is safe, efficacious, and elicits an immune response similar in magnitude to an immune response that controls or clears cells that bear or overexpress 24P4C12.

Example 22: Construction of "Miniaene" Multi-Epitope DNA Plasmids This example discusses the construction of a minigene expression plasmid. Minigene plasmids may, of course, contain various configurations of B cell, CTL and/or HTL epitopes or epitope analogs as described herein.

A minigene expression plasmid typically includes multiple CTL and HTL peptide epitopes. In the present example, HLA-A2, -A3, -B7 supermotif-bearing peptide epitopes and HLA-A1 and -A24 motif-bearing peptide epitopes are used in conjunction with DR supermotif-bearing epitopes and/or DR3 epitopes. HLA class I supermotif or motif-bearing peptide epitopes derived 24P4C12, are selected such that multiple supermotifs/motifs are represented to ensure broad population coverage. Similarly, HLA class II epitopes are selected from 24P4C12 to provide broad population coverage, i.e. both HLA DR-1-4-7 supermotif-bearing epitopes and HLA DR-3 motif-bearing epitopes are selected for inclusion in the minigene construct. The selected CTL and HTL epitopes are then incorporated into a minigene for expression in an expression vector.

Such a construct may additionally include sequences that direct the HTL epitopes to the endoplasmic reticulum.

For example, the li protein may be fused to one or more HTL epitopes as described in the art, wherein the CLIP sequence of the Ii protein is removed and replaced with an HLA class II epitope sequence so that HLA class II epitope is directed to the endoplasmic reticulum, where the epitope binds to an HLA class II molecules.

This example illustrates the methods to be used for construction of a minigene-bearing expression plasmid. Other expression vectors that may be used for minigene compositions are available and known to those of skill in the art.

The minigene DNA plasmid of this example contains a consensus Kozak sequence and a consensus murine kappa Ig-light chain signal sequence followed by CTL and/or HTL epitopes selected in accordance with principles disclosed herein. The sequence encodes an open reading frame fused to the Myc and His antibody epitope tag coded for by the pcDNA 3.1 Myc-His vector.

WO 2004/050828 PCT/US2002/038264 Overlapping oligonucleotides that can, for example, average about 70 nucleotides in length with 15 nucleotide overlaps, are synthesized and HPLC-purified. The oligonucleotides encode the selected peptide epitopes as well as appropriate linker nudeotides, Kozak sequence, and signal sequence. The final multiepitope minigeneis assembled by extending the overlapping oligonucleotidesin three sets of reactions using PCR. A PerkinfElmer 9600 PCR machine is used and a total of 30 cycles are performed using the following conditions: 95°C for 15 sec, annealing temperature below the lowest calculated Tm of each primer pair) for 30 sec, and 72"C for 1 min.

For example, a minigene is prepared as follows. For a first PCR reaction, 5 pg of each of two oligonucleotides are annealed and extended: In an example using eight oligonuceotides, four pairs of primers, oligonucleotides 1+2, 3+4, 5+6, and 7+8 are combined in 100 pi reactions containing Pfu polymerase buffer (lx= 10 mM KCL, 10 mM (NH4)2S04, mM Tris-chloride, pH 8.75, 2 mM MgSO4, 0.1% Triton X-100, 100 pgfml BSA), 0.25 mM each dNTP, and 2.5 U of Pfu polymerase. The full-length dimer products are gel-purified, and two reactions containing the product of 1+2 and 3+4, and the product of 5+6 and 7+8 are mixed, annealed, and extended for 10 cycles. Half of the two reactions are then mixed, and cycles of annealing and extension carried out before flanking primers are added to amplify the full length product. The fulllength product is gel-purified and cloned into pCR-blunt (Invitrogen) and individual clones are screened by sequencing.

Example 23: The Plasmid Construct and the Degree to Which It Induces Immunogenicity.

The degree to which a plasmid construct, for example a plasmid constructed in accordance with the previous Example, is able to induce immunogenicily is confirmed in vitro by determining epitope presentation by APC following transduction or transfection of the APC with an epitope-expressing nucleic acid construct. Such a study determines "antigenicity" and allows the use of human APC. The assay determines the ability of the epitope to be presented by the APC in a context that is recognized by a T cell by quantifying the density of epitope-HLA dass I complexes on the cell surface.

Quantitation can be performed by directly measuring the amount of peptide eluted from the APC (see, Sijts et J.

Immunol. 156:683-692, 1996; Demotz et al, Nature 342:682-684, 1989); or the number of peptide-HLA class I complexes can be estimated by measuring the amount of lysis or lymphokine release induced by diseased or transfected target cells, and then determining the concentration of peptide necessary to obtain equivalent levels of lysis or lymphokine release (see, Kageyama et al, J. Immunol. 154:567-576, 1995).

Altematively, immunogenicity is confirmed through in vivo injections into mice and subsequent in vitro assessment of CTL and HTL activity, which are analyzed using cytotoxicty and proliferation assays, respectively, as detailed in Alexander ef al., mmunity 1:751-761, 1994.

For example, to confirm the capacity of a DNA minigene construct containing at least one HLA-A2 supermotif peptide to induce CTLs in viva, HLA-A2.1/Kb transgenic mice, for example, are immunized intramuscularly with 100 pg of naked cDNA. As a means of comparing the level of CTLs induced by cDNA immunization, a control group of animals is also immunized with an actual peptide composition that comprises multiple epitopes synthesized as a single polypeptide as they would be encoded by the minigene.

Splenocytes from immunized animals are stimulated twice with each of the respective compositions (peptide epitopes encoded in the minigene or the polyepitopic peptide), then assayed for peptide-specific cytotoxic activity in a 51 Cr release assay. The results indicate the magnitude of the CTL response directed against the A2-restricted epitope, thus indicating the in vivo immunogenicity of the minigene vaccine and polyepitopic vaccine.

It is, therefore, found that the minigene elicits immune responses directed toward the HLA-A2 supermotif peptide epitopes as does the polyepitopic peptide vaccine. A similar analysis is also performed using other HLA-A3 and HLA-B7 transgenic mouse models to assess CTL induction by HLA-A3 and HLA-B7 motif or supermotif epitopes, whereby it is also found that the minigene elicits appropriate immune responses directed toward the provided epitopes.

WO 2004/050828 PCT/US2002/038264 To confirm the capacity of a class II epitope-encoding minigene to induce HTLs in vivo, DR transgenic mice, or for those epitopes that cross react with the appropriate mouse MHC molecule, I-Ab-restricted mice, for example, are immunized intramuscularly with 100 pg of plasmid DNA. As a means of comparing the level of HTLs induced by DNA immunization, a group of control animals is also immunized with an actual peptide composition emulsified in complete Freund's adjuvant.

CD4+ T cells, i.e. HTLs, are purified from splenocytes of immunized animals and stimulated with each of the respective compositions (peptides encoded in the minigene). The HTL response is measured using a 3 H-thymidine incorporation proliferation assay, (see, Alexander et a. Immunity 1:751-761, 1994). The results indicate the magnitude of the HTL response, thus demonstrating the in vivo immunogenicity of the minigene.

DNA minigenes, constructed as described in the previous Example, can also be confirmed as a vaccine in combination with a boosting agent using a prime boost protocol. The boosting agent can consist of recombinant protein Barnett et Aids Res. and Human Retrovirses 14, Supplement 3:S299-S309, 1998) or recombinant vaccinia, for example, expressing a minigene or DNA encoding the complete protein of interest (see, Hanke et Vaccine 16:439- 445, 1998; Sedegah etal., Proc. Natl. Acad. Sci USA 95:7648-53, 1998; Hanke and McMichael, Immunol. Letters 66:177- 181, 1999; and Robinson etal., Nature Med. 5:526-34, 1999).

For example, the efficacy of the DNA minigene used in a prime boost protocol is initially evaluated in transgenic mice. In this example, A2.1/Kb transgenic mice are immunized IM with 100 Ig of a DNA minigene encoding the immunogenic peptides including at least one HLA-A2 supermolif-bearing peptide. After an incubation period (ranging from 3- 9 weeks), the mice are boosted IP with 10 7 pfulmouse of a recombinant vaccinia virus expressing the same sequence encoded by the DNA minigene. Control mice are immunized with 100 ig of DNA or recombinant vaccinia without the minigene sequence, or with DNA encoding the minigene, but without the vaccinia boost. After an additional incubation period of two weeks, splenocytes from the mice are immediately assayed for peptide-specific activity in an ELISPOT assay.

Additionally, splenocytes are stimulated in vitro with the A2-restricted peptide epitopes encoded in the minigene and recombinant vaccinia, then assayed for peptide-specific activity in an alpha, beta andlor gamma IFN ELISA.

It is found that the minigene utilized in a prime-boost protocol elicits greater immune responses toward the HLA-A2 supermotif peptides than with DNA alone. Such an analysis can also be performed using HLA-A11 or HLA-B7 transgenic mouse models to assess CTL induction by HLA-A3 or HLA-B7 motif or supermotif epitopes. The use of prime boost protocols in humans is described below in the Example entitled "Induction of CTL Responses Using a Prime Boost Protocol." Example 24: Peptide Compositions for Prophylactic Uses Vaccine compositions of the present invention can be used to prevent 24P4C12 expression in persons who are at risk for tumors that bear this antigen. For example, a polyepitopic peptide epitope composition (or a nucleic acid comprising the same) containing multiple CTL and HTL epitopes such as those selected in the above Examples, which are also selected to target greater than 80% of the population, is administered to individuals at risk for a 24P4C12-associated tumor.

For example, a peptide-based composition is provided as a single polypeptide that encompasses multiple epitopes. The vaccine is typically administered in a physiological solution that comprises an adjuvant, such as Incomplete Freunds Adjuvant. The dose of peptide for the initial immunization is from about 1 to about 50,000 jg, generally 100-5,000 ag, for a 70 kg patient. The initial administration of vaccine is followed by booster dosages at 4 weeks followed by evaluation of the magnitude of the immune response in the patient, by techniques that determine the presence of epitopespecific CTL populations in a PBMC sample. Additional booster doses are administered as required. The composition is found to be both safe and efficacious as a prophylaxis against 24P4C12-associated disease.

Alternatively, a composition typically comprising transfecting agents is used for the administration of a nucleic acidbased vaccine in accordance with methodologies known in the art and disclosed herein.

WO 2004/050828 PCT/US2002/038264 Example 25: Polyepitopic Vaccine Compositions Derived from Native 24P4C12 Sequences A native 24P4C12 polyprotein sequence is analyzed, preferably using computer algorithms defined for each class I and/or class II supermotif or motif, to identify "relatively short" regions of the polyprotein that comprise multiple epitopes, The "relatively short" regions are preferably less in length than an entire native antigen. This relatively short sequence that contains multiple distinct or overlapping, 'nested" epitopes can be used to generate a minigene construct. The construct is engineered to express the peptide, which corresponds to the native protein sequence. The "relatively short" peptide is generally less than 250 amino acids in length, often less than 100 amino acids in length, preferably less than 75 amino acds in length, and more preferably less than 50 amino acids in length. The protein sequence of the vaccine composition is selected because it has maximal number of epitopes contained within the sequence, it has a high concentration of epitopes. As noted herein, epitope motifs may be nested or overlapping frame shifted relative to one another). For example, with overlapping epitopes, two 9-mer epitopes and one 10-mer epitope can be present in a 10 amino acid peptide.

Such a vaccine composition is administered for therapeutic or prophylactic purposes.

The vaccine composition will include, for example, multiple CTL epitopes from 24P4C12 antigen and at least one HTL epitope. This polyepitopic native sequence is administered either as a peptide or as a nucleic acid sequence which encodes the peptide. Alternatively, an analog can be made of this native sequence, whereby one or more of the epitopes comprise substitutions that alter the cross-reactivity and/or binding affinity properties of the polyepitopic peptide.

The embodiment of this example provides for the possibility that an as yet undiscovered aspect of immune system processing will apply to the native nested sequence and thereby facilitate the production of therapeutic or prophylactic immune response-inducing vaccine compositions. Additionally, such an embodiment provides for the possibility of motifbearing epitopes for an HLA makeup(s) that is presently unknown. Furthermore, this embodiment (excluding an analoged embodiment) directs the immune response to multiple peptide sequences that are actually present in native 24P4C12, thus avoiding the need to evaluate any junctional epitopes. Lastly, the embodiment provides an economy of scale when producing peptide or nucleic acid vaccine compositions.

Related to this embodiment, computer programs are available in the art which can be used to identify in a target sequence, the greatest number of epitopes per sequence length.

Example 26: Polvepitopic Vaccine Compositions from Multiple Antigens The 24P4C12 peptide epitopes of the present invention are used in conjunction with epitopes from other target tumor-associated antigens, to create a vaccine composition that is useful for the prevention or treatment of cancer that expresses 24P4C12 and such other antigens. For example, a vaccine composition can be provided as a single polypeptide that incorporates multiple epitopes from 24P4C12 as well as tumor-associated antigens that are often expressed with a target cancer associated with 24P4C12 expression, or can be administered as a composition comprising a cocktail of one or more discrete epitopes. Alternatively, the vaccine can be administered as a minigene constructor as dendritic cells which have been loaded with the peptide epitopes in vitro.

Example 27: Use of peptides to evaluate an immune response Peptides of the invention may be used to analyze an immune response for the presence of specific antibodies, CTL or HTL directed to 24P4C12. Such an analysis can be performed in a manner described by Ogg et al, Science 279:2103-2106, 1998. In this Example, peptides in accordance with the invention are used as a reagent for diagnostic or prognostic purposes, not as an immunogen.

WO 2004/050828 PCT/US2002/038264 In this example highly sensitive human leukocyte antigen tetrameric complexes ("tetramers") are used for a crosssectional analysis of, for example, 24P4C12 HLA-A*0201-specific CTL frequencies from HLA A*0201-positive individuals at different stages of disease or following immunization comprising a 24P4C12 peptide containing an A*0201 motif. Tetrameric complexes are synthesized as described (Musey et al., N. Engl. J. Mad. 337:1267,1997). Briefly, purified HLA heavy chain (A*0201 in this example) and P2-microglobulin are synthesized by means of a prokaryotic expression system. The heavy chain is modified by deletion of the transmembrane-cytosolic tail and COOH-terminal addition of a sequence containing a BirA enzymatic biotinylation site. The heavy chain, p2-microglobulin, and peptide are refolded by dilution. The refolded product is isolated by fast protein liquid chromatography and then biotinylated by BirA in the presence of biotin (Sigma, St. Louis, Missouri), adenosine 5' triphosphate and magnesium. Streptavidin-phycoerythrin conjugate is added in a 1:4 molar ratio, and the tetrameric product is concentrated to 1 mg/ml. The resulting product is referred to as tetramerphycoerythrin.

For the analysis of patient blood samples, approximately one million PBMCs are centrifuged at 300g for 5 minutes and resuspended in 50 pl of cold phosphate-buffered saline. Tri-color analysis is performed with the tetramer-phycoerythrin, along with anti-CD8-Tricolor, and anti-CD38. The PBMCs are incubated with tetramer and antibodies on ice for 30 to 60 min and then washed twice before formaldehyde fixation. Gates are applied to contain >99.98% of control samples. Controls for the tetramers include both A*0201-negative individuals and A*0201-positive non-diseased donors. The percentage of cells stained with the tetramer is then determined by flow cytometry. The results indicate the number of cells in the PBMC sample that contain epitope-restricted CTLs, thereby readily indicating the extent of immune response to the 24P4C12 epitope, and thus the status of exposure to 24P4C12, or exposure to a vaccine that elicits a protective or therapeutic response.

Example 28: Use of Peptide Epitopes to Evaluate Recall Responses The peptide epitopes of the invention are used as reagents to evaluate T cell responses, such as acute or recall responses, in patients. Such an analysis may be performed on patients who have recovered from 24P4C12-associated disease or who have been vaccinated with a 24P4C12 vaccine.

For example, the class I restricted CTL response of persons who have been vaccinated may be analyzed. The vaccine may be any 24P4C12 vaccine. PBMC are collected from vaccinated individuals and HLA typed. Appropriate peptide epitopes of the invention that, optimally, bear supermotifs to provide cross-reactivity with multiple HLA supertype family members, are then used for analysis of samples derived from individuals who bear that HLA type.

PBMC from vaccinated individuals are separated on Ficoll-Histopaque density gradients (Sigma Chemical Co., St.

Louis, MO), washed three times in HBSS (GIBCO Laboratories), resuspended in RPMI-1640 (GIBCO Laboratories) supplemented with L-glutamine (2mM), penicillin (50U/ml), streptomycin (50 gg/ml), and Hepes (10mM) containing heat-inactivated human AB serum (complete RPMI) and plated using microculture formats. A synthetic peptide comprising an epitope of the invention is added at 10 pg/lm to each well and HBV core 128-140 epitope is added at 1 pg/ml to each well as a source of T cell help during the first week of stimulation.

In the microculture format, 4 x 105 PBMC are stimulated with peptide in 8 replicate cultures in 96-well round bottom plate in 100 pi/well of complete RPMI. On days 3 and 10, 100 pl of complete RPMI and 20 U/ml final concentration of rlL-2 are added to each well. On day 7 the cultures are transferred into a 96-well flat-bottom plate and restimulated with peptide, rlL-2 and 105 irradiated (3,000 rad) autologous feeder cells. The cultures are tested for cytatoxic activity on day 14. A positive CTL response requires two or more of the eight replicate cultures to display greater than 10% specific 5'Cr release, based on comparison with non-diseased control subjects as previously described (Rehermann, et Nafure Med.

2:1104,1108, 1996; Rehermann et al, J. Clin. Invest. 97:1655-1665, 1996; and Rehermann etal. J. Clin. Invest. 98:1432- 1440, 1996).

WO 2004/050828 PCT/US2002/038264 Target cell lines are autologous and allogeneic EBV-transformed B-LCL that are either purchased from the American Society for Histocompatibility and Immunogenetcs (ASHI, Boston, MA) or established from the pool of patients as described (Guilhot, et a J. Virol. 66:2670-2678, 1992).

Cytotoxicity assays are performed in the following manner. Target cells consist of either allogeneic HLA-matched or autologous EBV-transformed B lymphoblastoid cell line that are incubated overnight with the synthetic peptide epitope of the invention at 10 pM, and labeled with 100 pCi of 51 Cr (Amersham Corp., Arlington Heights, IL) for 1 hour after which they are washed four times with HBSS.

Cytolytic activity is determined in a standard 4-h, split well 5 sCr release assay using U-bottomed 96 well plates containing 3,000 targets/well. Stimulated PBMC are tested at effector/target ratios of 20-50:1 on day 14. Percent cytotoxicity is determined from the formula: 100 x [(experimental release-spontaneous release)/maximum releasespontaneous release)]. Maximum release is determined by lysis of targets by detergent Triton X-100; Sigma Chemical Co., St. Louis, MO). Spontaneous release is <25% of maximum release for all experiments.

The results of such an analysis indicate the extent to which HLA-restricted CTL populations have been stimulated by previous exposure to 24P4C12 or a 24P4C12 vaccine.

Similarly, Class II restricted HTL responses may also be analyzed. Purified PBMC are cultured in a 96-well flat bottom plate at a density of 1.5x105 cells/well and are stimulated with 10 pg/ml synthetic peptide of the invention, whole 24P4C12 antigen, or PHA. Cells are routinely plated in replicates of 4-6 wells for each condition. After seven days of culture, the medium is removed and replaced with fresh medium containing 10U/ml IL-2. Two days later, 1 pCi 3 H-thymidine is added to each well and incubation is continued for an additional 18 hours. Cellular DNA is then harvested on glass fiber mats and analyzed for 3 H-thymidine incorporation. Antigen-specific T cell proliferation is calculated as the ratio of 3

H-

thymidine incorporation in the presence of antigen divided by the 3H-thymidine incorporation in the absence of antigen.

Example 29: Induction Of Specific CTL Response In Humans A human clinical trial for an immunogenic composition comprising CTL and HTL epitopes of the invention is set up as an IND Phase I, dose escalation study and carried out as a randomized, double-blind, placebo-controlled trial. Such a trial is designed, for example, as follows: A total of about 27 individuals are enrolled and divided into 3 groups: Group I: 3 subjects are injected with placebo and 6 subjects are injected with 5 p.g of peptide composition; Group II: 3 subjects are injected with placebo and 6 subjects are injected with 50 pg peptide composition; Group II: 3 subjects are injected with placebo and 6 subjects are injected with 500 pg of peptide composition.

After 4 weeks following the first injection, all subjects receive a booster inoculation at the same dosage.

The endpoints measured in this study relate to the safety and tolerability of the peptide composition as well as its immunogenicity. Cellular immune responses to the peptide composition are an index of the intrinsic activity of this the peptide composition, and can therefore be viewed as a measure of biolog cal efficacy. The following summarize the clinical and laboratory data that relate to safety and eficacy endpoints.

Safety: The incidence of adverse events is monitored in the placebo and drug treatment group and assessed in terms of degree and reversibility.

Evaluation of Vaccine Efficacy: For evaluation of vaccine efficacy, subjects are bled before and after injection.

Peripheral blood mononuclear cells are isolated from fresh heparinized blood by Ficoll-Hypaque density gradient centrifugation, aliquoted in freezing media and stored frozen. Samples are assayed for CTL and HTL activity.

The vaccine is found to be both safe and efficacious.

WO 2004/050828 PCT/US2002/038264 Example 30: Phase II Trials In Patients Expressing 24P4C12 Phase II trials are performed to study the effect of administering the CTL-HTL peptide compositions to patients having cancer that expresses 24P4C12. The main objectives of the trial are to determine an effective dose and regimen for inducing CTLs in cancer patients that express 24P4C12, to establish the safety of inducing a CTL and HTL response in these patients, and to see to what extent activation of CTLs improves the clinical picture of these patients, as manifested, by the reduction andlor shrinking of lesions. Such a study is designed, for example, as follows: The studies are performed in multiple centers. The trial design is an open-label, uncontrolled, dose escalation protocol wherein the peptide composition is administered as a single dose followed six weeks later by a single booster shot of the same dose. The dosages are 50, 500 and 5,000 micrograms per injection. Drug-assocated adverse effects (severity and reversibility) are recorded.

There are three patient groupings. The first group is injected with 50 micrograms of the peptide composition and the second and third groups with 500 and 5,000 micrograms of peptide composition, respectively. The patients within each group range in age from 21-65 and represent diverse ethnic backgrounds. All of them have a tumor that expresses 24P4C12.

Clinical manifestations or antigen-specific T-cell responses are monitored to assess the effects of administering the peptide compositions. The vaccine composition is found to be both safe and efficacious in the treatment of 24P4C12associated disease.

Example 31: Induction of CTL Responses Using a Prime Boost Protocol A prime boost protocol similar in its underlying principle to that used to confirm the efficacy of a DNA vaccine in transgenic mice, such as described above in the Example entitled "The Plasmid Construct and the Degree to Which It Induces Immunogenicity," can also be used for the administration of the vaccine to humans. Such a vaccine regimen can include an initial administration of, for example, naked DNA followed by a boost using recombinant virus encoding the vaccine, or recombinant protein/polypeptide or a peptide mixture administered in an adjuvant.

For example, the initial immunization may be performed using an expression vector, such as that constructed in the Example entitled "Construction of "Minigene" Multi-Epitope DNA Plasmids" in the form of naked nucleic acid administered IM (or SC or ID) in the amounts of 0.5-5 mg at multiple sites. The nucleic acid (0.1 to 1000 pg) can also be administered using a gene gun. Following an incubation period of 34 weeks, a booster dose is then administered. The booster can be recombinant fowlpox virus administered at a dose of 5-10 7 to 5x109 pfu. An alternative recombinant virus, such as an MVA, canarypox, adenovirus, or adeno-associaled virus, can also be used for the booster, or the polyepitopic protein or a mixture of the peptides can be administered. For evaluation of vaccine efficacy, patient blood samples are obtained before immunization as well as at intervals following administration of the initial vaccine and booster doses of the vaccine.

Analysis of the results indicates that a magnitude of response sufficient to achieve a therapeutic or protective immunity against 24P4C12 is generated.

Example 32: Administration of Vaccine Compositions Using Dendritic Cells (DC) Vaccines comprising peptide epitopes of the invention can be administered using APCs, or "professional" APCs such as DC. In this example, peptide-pulsed DC are administered to a patient to stimulate a CTL response in vive. In this method, dendritic cells are isolated, expanded, and pulsed with a vaccine comprising peptide CTL and HTL epitopes of the invention. The dendritic cells are infused back into the patient to elicit CTL and HTL responses in vive. The induced CTL WO 2004/050828 PCT/US2002/038264 and HTL then destroy or facilitate destruction, respectively, of the target cells that bear the 24P4C12 protein from which the epitopes in the vaccine are derived.

For example, a cocktail of epitope-comprising peptides is administered ex vivo to PBMC, or isolated DC therefrom.

A pharmaceutical to facilitate harvesting of DC can be used, such as Progenipoietin T M (Monsanto, St. Louis, MO) or GM- CSFIIL-4. After pulsing the DC with peptides, and prior to reinfusion into patients, the DC are washed to remove unbound peptides.

As appreciated clinically, and readily determined by one of skill based on clinical outcomes, the number of DC reinfused into the patient can vary (see, Nature Med. 4:328, 1998; Nature Med. 2:52, 1996 and Prostate 32:272, 1997).

Although 2-50 x 10 6 DC per patient are typically administered, larger number of DC, such as 10 7 or 108 can also be provided.

Such cell populations typically contain between 50-90% DC.

In some embodiments, peptide-loaded PBMC are injected into patients without purification of the DC. For example, PBMC generated after treatment with an agent such as Progenipoietin T M are injected into patients without purification of the DC. The total number of PBMC that are administered often ranges from 108 to 1010. Generally, the cell doses injected into patients is based on the percentage of DC in the blood of each patient, as determined, for example, by immunofluorescence analysis with specific anti-DC antibodies. Thus, for example, if Progenipoietin T M mobilizes 2% DC in the peripheral blood of a given patient, and that patient is to receive 5 x 106 DC, then the patient will be injected with a total of 2.5 x 108 peptide-loaded PBMC. The percent DC mobilized by an agent such as Progenipoietin T M is typically estimated to be between 2-10%, but can vary as appreciated by one of skill in the art.

Ex vio activation of CTL/HTL responses Alternatively, ex vivo CTL or HTL responses to 24P4C12 antigens can be induced by incubating, in tissue culture, the patient's, or genetically compatible, CTL or HTL precursor cells together with a source of APC, such as DC, and immunogenic peptides. After an appropriate incubation time (typically about 7-28 days), in which the precursor cells are activated and expanded into effector cells, the cells are infused Into the patient, where they will destroy (CTL) or facilitate destruction (HTL) of their specific target cells, tumor cells.

Example 33: An Alternative Method of Identifying and Confirming Motif-Bearing Peptides Another method of identifying and confirming motif-bearing peptides is to elute them from cells bearing defined MHC molecules. For example, EBV transformed B cell lines used for tissue typing have been extensively characterized to determine which HLA molecules they express. In certain cases these cells express only a single type of HLA molecule.

These cells can be transfected with nucleic acids that express the antigen of interest, e.g. 24P4C12. Peptides produced by endogenous antigen processing of peptides produced as a result of transfection will then bind to HLA molecules within the cell and be transported and displayed on the cell's surface. Peptides are then eluted from the HLA molecules by exposure to mild acid conditions and their amino acid sequence determined, by mass spectral analysis Kubo et al, J.

Immunol. 152:3913, 1994). Because the majority of peptides that bind a particular HLA molecule are motif-bearing, this is an atemative modality for obtaining the motif-bearing peptides correlated with the particular HLA molecule expressed on the cell.

Alternatively, cell lines that do not express endogenous HLA molecules can be transfected with an expression construct encoding a single HLA allele. These cells can then be used as described, they can then be transfected with nucleic acids that encode 24P4C12 to isolate peptides corresponding to 24P4C12 that have been presented on the cell surface. Peptides obtained from such an analysis will bear motif(s) that correspond to binding to the single HLA allele that is expressed in the cell.

WO 2004/050828 PCT/US2002/038264 As appreciated by one in the art, one can perform a similar analysis on a cell bearing more than one HLA allele and subsequently determine peptides specific for each HLA allele expressed. Moreover, one of skill would also recognize that means other than transfection, such as loading with a protein antigen, can be used to provide a source of antigen to the cell.

Example 34: Complementary Polynucleotides Sequences complementary to the 24P4C12-encoding sequences, or any parts thereof, are used to detect, decrease, or inhibit expression of naturally occurring 24P4C12. Although use ofoligonucleotides comprising from about to 30 base pairs is described, essentially the same procedure is used with smaller or with larger sequence fragments.

Appropriate oligonucleotides are designed using, OLIGO 4.06 software (National Biosciences) and the coding sequence of 24P4C12. To inhibit transcription, a complementary oligonucleotide is designed from the most unique 5' sequence and used to prevent promoter binding to the coding sequence. To inhibit translation, a complementary oligonucleotide is designed to prevent ribosomal binding to a 24P4C12-encoding transcript.

Example 35: Purification of Naturally-occurring or Recombinant 24P4C12 Using 24P4C12-Specific Antibodies Naturally occurring or recombinant 24P4C12 is substantially purified by immunoaffinity chromatography using antibodies specific for 24P4C12. An immunoaffinity column is constructed by covalently coupling anti-24P4C12 antibody to an activated chromatographic resin, such as CNBr-activated SEPHAROSE (Amersham Pharmacia Biotech). After the coupling, the resin is blocked and washed according to the manufacturer's instructions.

Media containing 24P4C12 are passed over the immunoaffinity column, and the column is washed under conditions that allow the preferential absorbance of 24P4C12 high ionic strength buffers in the presence of detergent).

The column is eluted under conditions that disrupt antibody/24P4C12 binding a buffer of pH 2 to pH 3, or a high concentration of a chaotrope, such as urea or thiocyanate ion), and GCR.P is collected.

Example 36: Identification of Molecules Which Interact with 24P4C12 24P4C12, or biologically active fragments thereof, are labeled with 121 1 Bolton-Hunter reagent. (See, Bolton etal. (1973) Biochem. J. 133:529.) Candidate molecules previously arrayed in the wells of a multi-well plate are incubated with the labeled 24P4C12, washed, and any wells with labeled 24P4C12 complex are assayed. Data obtained using different concentrations of 24P4C12 are used to calculate values for the number, affinity, and association of 24P4C12 with the candidate molecules.

Example 37: In Vivo Assay for 24P4C12 Tumor Growth Promotion The effect of the 24P4C12 protein on tumor cell growth is evaluated in vivo by evaluating tumor development and growth of cells expressing or lacking 24P4C12. For example, SCID mice are injected subcutaneously on each flank with 1 x 106 of either 3T3, prostate, colon, ovary, lung, or bladder cancer cell lines PC3, Caco, PA-1, CaLu or J82 cells) containing tkNeo empty vector or 24P4C12. At least two strategies may be used: Constitutive 24P4C12 expression under regulation of a promoter, such as a constitutive promoter obtained from the genomes of viruses such as polyoma virus, fowlpox virus (UK 2,211,504 published 5 July 1989), adenovirus (such as Adenovirus bovine papilloma virus, avian sarcoma virus, cytomegalovirus, a retrovirus, hepatitis-B virus and Simian Virus 40 (SV40), or from heterologous mammalian promoters, the actin promoter or an immunoglobulin promoter, provided such promoters are compatible with the host cell systems, and Regulated expression under control of an inducible vector system, such as ecdysone, tetracycline, etc., provided such promoters are compatible with the host cell systems. Tumor volume is then monitored by caliper WO 2004/050828 PCT/US2002/038264 measurement at the appearance of palpable tumors and followed over time to determine if 24P4C12-expressing cells grow at a faster rate and whether tumors produced by 24P4C12-expressing cells demonstrate characteristics of altered aggressiveness enhanced metastasis, vascularization, reduced responsiveness to chemotherapeutic drugs). As shown in figure 31 and Figure 32, 24P4C12 has a profound effect on tumor growth in SCID mice. The prostate cancer cells PC3 and PC3-24P4C12 were injected subcutaneously in the right flank of SCID mice. Tumor growth was evaluated by caliper measurements An increase in tumor growth was observed in PC3-24P4C12 tumors within 47 days of injection (fig 31). In addition, subcutaneous injection of 3T3-24P4C12 induced tumor formation in SCID mice (Figure 32). This finding is significant as control 3T3 cells fail to form tumors, indicating that 24P4C12 has several tumor enhancing capabilities, including transformation, as well as tumor initiation and promotion.

Example 38: 24P4C12 Monoclonal Antibody-mediated Inhibition of Prostate Tumors In Vivo.

The significant expression of 24P4C12 in cancer tissues, together with its restrictive expression in normal tissues and cell surface localization, make 24P4C12 a good target for antibody therapy. Similarly, 24P4C12 is a target for T cellbased immunotherapy. Thus, the therapeutic efficacy of anti-24P4C12 mAbs in human prostate cancer xenograft mouse models is evaluated by using recombinant cell lines such as PC3-24P4C12, and 3T3-24P4C12 (see, Kaighn, ef aL, Invest Urol, 1979. 17(1): p. 16-23), as well as human prostate xenograft models such as LAPC9 (Saffran et al, Proc Nati Acad Sci U S A. 2001,98:2658). Similarly, anti-24P4C12 mAbs are evaluated in xenograft models of human bladder cancer colon cancer, ovarian cancer or lung cancer using recombinant cell lines such as J82-24P4C12, Caco-24P4C12, PA- 24P4C1 or CaLu-24P4C12, respectively.

Antibody efficacy on tumor growth.and metastasis formation is studied, in a mouse orthotopic bladder cancer xenograft model, and a mouse prostate cancer xenograft model, The antibodies can be unconjugated, as discussed in this Example, or can be conjugated to a therapeutic modality, as appreciated in the art. Ant-24P4C12 mAbs inhibit formation of prostate and bladder xenografts. Anti-24P4C12 mAbs also retard the growth of established orthotopic tumors and prolonged survival of tumor-bearing mice. These results indicate the utility of anti-24P4C12 mAbs in the treatment of local and advanced stages of prostate, colon, ovarian, lung and bladder cancer. (See, Saffran, et al., PNAS 10:1073-1078 or www.pnas.org/cgildoi/1 0.1073/pnas.051624698).

Administration of the anti-24P4C12 mAbs led to retardation of established orthotopic tumor growth and inhibition of metastasis to distant sites, resulting in a significant prolongation in the survival of tumor-bearing mice. These studies indicate that 24P4C12 as an attractive target for immunotherapy and demonstrate the therapeutic potential of anti-24P4C12 mAbs for the treatment of local and metastatic cancer. This example demonstrates that unconjugated 24P4C12 monoclonal antibodies are effective to inhibit the growth of human prostate, colon, ovarian, lung and bladder cancer tumor xenografts grown in SCID mice; accordingly a combination of such efficacious monoclonal antibodies is also effective.

Tumor inhibition using multiple unconjugated 24P4C12 mAbs Materials and Methods 24P4C12 Monoclonal Antibodies: Monoclonal antibodies are raised against 24P4C12 as described in the Example entitled "Generation of 24P4C12 Monoclonal Antibodies (mAbs)." The antibodies are characterized by ELISA, Western blot, FACS, and immunoprecipitation for their capacity to bind 24P4C12. Epitope mapping data for the anti-24P4C12 mAbs, as determined by ELISA and Western analysis, recognize epitopes on the 24P4C12 protein. Immunohistochemical analysis of prostate cancer tissues and cells with these antibodies is performed.

WO 2004/050828 PCT/US2002/038264 The monoclonal antibodies are purified from ascites or hybridoma tissue culture supernatants by Protein-G Sepharose chromatography, dialyzed against PBS, filter sterilized, and stored at -20°C. Protein determinations are performed by a Bradford assay (Bio-Rad, Hercules, CA). A therapeutic monoclonal antibody or a cocktail comprising a mixture of individual monoclonal antibodies is prepared and used for the treatment of mice receiving subcutaneous or orthotopic injections of SCABER, J82, A498, 769P, CaOvl or PA1 tumor xenografts.

Cell Lines The prostate, colon, ovarian, lung and bladder cancer carcinoma cell lines,, Caco, PA-1, CaLu or J82 cells as well as the fibroblast line NIH 3T3 (American Type Culture Collection) are maintained in media supplemented with L-glutamine and 10% FBS.

PC3-24P4C12, Caco-24P4C12, PA-24P4C12, CaLu-24P4C12 or J82-24P4C12 cells and 3T3-24P4C12 cell populations are generated by retroviral gene transfer as described in Hubert, et al., Proc Nati Acad Sci U S A, 1999.

96(25): 14523.

Xenoqraft Mouse Models.

Subcutaneous tumors are generated by injection of 1 x 10 6 cancer cells mixed at a 1:1 dilution with Matrigel (Collaborative Research) in the right flank of male SCID mice. To test antibody efficacy on tumor formation, i.p. antibody injections are started on the same day as tumor-cell injections. As a control, mice are injected with either purified mouse IgG (ICN) or PBS; or a purified monoclonal antibody that recognizes an irrelevant antigen not expressed in human cells. Tumor sizes are determined by caliper measurements, and the tumor volume is calculated as: Length x Width x Height Mice with s.c. tumors greater than 1.5 cm in diameter are sacrificed.

Orthotopic injections are performed under anesthesia by using ketamine/xylazine. For bladder orthotopic studies, an incision is made through the abdomen to expose the bladder, and tumor cells (5 x 105) mixed with Matrigel are injected into the bladder wall in a 10-pl volume. To monitor tumor growth, mice are palpated and blood is collected on a weekly basis to measure BTA levels. For prostate orthopotic models, an incision is made through the abdominal muscles to expose the bladder and seminal vesicles, which then are delivered through the incision to expose the dorsal prostate. Tumor cells e.g.

LAPC-9 cells (5 x 10) mixed with Matrigel are injected into the prostate in a 10-JI volume (Yoshida Yet al, Anticancer Res.

1998, 18:327; Ahn et al, Tumour Biol. 2001, 22:146). To monitor tumor growth, blood is collected on a weekly basis measuring PSA levels. Similar procedures are followed for lung and ovarian xenograft models. The mice are segregated into groups for the appropriate treatments, with anti-24P4C12 or control mAbs being injected I.p.

Anti-24P4C12 mAbs Inhibit Growth of 24P4C12-Exoressing Xenograft-Cancer Tumors The effect of anti-24P4C12 mAbs on tumor formation is tested on the growth and progression of bladder, and prostate cancer xenografts using PC3-24P4C12, Caco-24P4C12, PA-24P4C12, CaLu-24P4C12 or J82-24P4C12 orthotopic models. As compared with the s.c. tumor model, the orthotopic model, which requires injection of tumor cells directly in the mouse prostate, colon, ovary, lung and bladder, respectively, results in a local tumor growth, development of metastasis in distal sites, deterioration of mouse health, and subsequent death (Saffran, et al., PNAS supra; Fu, et al., Int J Cancer, 1992. 52(6): p. 987-90; Kubota, J Cell Biochem, 1994.56(1): p. The features make the orthotopic model more representative of human disease progression and allowed us to follow the therapeutic effect of mAbs on clinically relevant end points.

Accordingly, tumor cells are injected into the mouse organs, and 2 days later, the mice are segregated into two groups and treated with either: a) 200-500pg, of anti-24P4C12 Ab, or b) PBS three times per week for two to five weeks.

A major advantage of the orthotopic cancer models is the ability to study the development of metastases.

Formation of metastasis in mice bearing established orthotopic tumors is studies by IHC analysis on lung sections using an WO 2004/050828 PCT/US2002/038264 antibody against a tumor-specific cell-surface protein such as anti-CK20 for bladder cancer, anti-STEAP-1 for prostate cancer models (Lin S et al, Cancer Detect Prev. 2001;25:202; Saffran, et al., PNAS supra).

Mice bearing established orthotopic tumors are administered 1000pg injections of either anti-24P4C12 mAb or PBS over a 4-week period. Mice in both groups are allowed to establish a high tumor burden, to ensure a high frequency of metastasis formation in mouse lungs. Mice then are killed and their bladders, livers, bone and lungs are analyzed for the presence of tumor cells by IHC analysis.

These studies demonstrate a broad anti-tumor efficacy of anti-24P4C12 antibodies on initiation and progression of prostate and kidney cancer in xenograft mouse models. Anti-24P4C12 antibodies inhibit tumor formation of tumors as well as retarding the growth of already established tumors and prolong the survival of treated mice. Moreover, anti-24P4C12 mAbs demonstrate a dramatic inhibitory effect on the spread of local bladder and prostate tumor to distal sites, even in the presence of a large tumor burden. Thus, anti-24P4C12 mAbs are efficacious on major clinically relevant end points (tumor growth), prolongation of survival, and health.

Example 39: Therapeutic and Diagnostic use of Anti-24P4C12 Antibodies in Humans.

Anti-24P4C12 monoclonal antibodies are safely and effectively used for diagnostic, prophylactic, prognostic and/or therapeutic purposes in humans. Western blot and immunohistochemical analysis of cancer tissues and cancer xenografts with anti-24P4C12 mAb show strong extensive staining in carcinoma but significantly lower or undetectable levels in normal tissues. Detection of 24P4C12 in carcinoma and in metastatic disease demonstrates the usefulness of the mAb as a diagnostic and/or prognostic indicator. Anti-24P4C12 antibodies are therefore used in diagnostic applications such as immunohistochemistry of kidney biopsy specimens to detect cancer from suspect patients.

As determined by flow cytometry, anti-24P4C12 mAb specifically binds to carcinoma cells. Thus, anti-24P4C12 antibodies are used in diagnostic whole body imaging applications, such as radioimmunoscintigraphy and radioimmunotherapy, (see, Potamianos et al. Anticancer Res 20(2A):925-948 (2000)) for the detection of localized and metastatic cancers that exhibit expression of 24P4C12. Shedding or release of an extracellular domain of 24P4C12 into the extracellular milieu, such as that seen for alkaline phosphodiesterase B10 (Meerson, N. Hepatology 27:563-568 (1998)), allows diagnostic detection of 24P4C12 by anti-24P4C12 antibodies in serum and/or urine samples from suspect patients.

Anti-24P4C12 antibodies that specifically bind 24P4C12 are used in therapeutic applications for the treatment of cancers that express 24P4C12. Anti-24P4C12 antibodies are used as an unconjugated modality and as conjugated form in which the antibodies are attached to one of various therapeutic or imaging modalities well known in the art, such as a prodrugs, enzymes or radiolsotopes. In preclinical studies, unconjugated and conjugated anti-24P4C12 antibodies are tested for efficacy of tumor prevention and growth inhibition in the SCID mouse cancer xenograft models, kidney cancer .models AGS-K3 and AGS-K6, (see, the Example entitled '24P4C12 Monoclonal Antibody-mediated Inhibition of Bladder and Lung Tumors In Vivo). Either conjugated and unconjugated anti-24P4C12 antibodies are used as a therapeutic modality in human clinical trials either alone or in combination with other treatments as described in following Examples.

Example 40: Human Clinical Trials for the Treatment and Diagnosis of Human Carcinomas through use of Human Anti-24P4C12 Antibodies In vivo Antibodies are used in accordance with the present invention which recognize an epitope on 24P4C12, and are used in the treatment of certain tumors such as those listed in Table I. Based upon a number of factors, including 24P4C12 expression levels, tumors such as those listed in Table I are presently preferred indications. In connection with each of these indications, three clinical approaches are successfully pursued.

WO 2004/050828 PCT/US2002/038264 Adjunctive therapy: In adjunctive therapy, patients are treated with anti-24P4C12 antibodies in combination with a chemotherapeutic or antineoplastic agent and/or radiation therapy. Primary cancer targets, such as those listed in Table I, are treated under standard protocols by the addition anti-24P4C12 antibodies to standard first and second line therapy. Protocol designs address effectiveness as assessed by reduction in tumor mass as well as the ability to reduce usual doses of standard chemotherapy. These dosage reductions allow additional and/or prolonged therapy by reducing dose-related toxicity of the chemotherapeutic agent. Anti-24P4C12 antibodies are utilized in several adjunctive clinical trials in combination with the chemotherapeutic or antineoplastic agents adriamycin (advanced prostrate carcinoma), cisplatin (advanced head and neck and lung carcinomas), taxol (breast cancer), and doxorubicin (preclinical).

II.) Monotherapy: In connection with the use of the anti-24P4C12 antibodies in monotherapy of tumors, the antibodies are administered to patients without a chemotherapeutic or antineoplastic agent. In one embodiment, monotherapy is conducted clinically in end stage cancer patients with extensive metastatic disease. Patients show some disease stabilization. Trials demonstrate an effect in refractory patients with cancerous tumors.

III.) Imaging Agent: Through binding a radionuclide iodine or yttrium (I 13 1, Y 90 to anti-24P4C12 antibodies, the radiolabeled antibodies are utilized as a diagnostic and/or imaging agent In such a role, the labeled antibodies localize to both solid tumors, as well as, metastatic lesions of cells expressing 24P4C12. In connection with the use of the anti-24P4C12 antibodies as imaging agents, the antibodies are used as an adjunct to surgical treatment of solid tumors, as both a pre-surgical screen as well as a post-operative follow-up to determine what tumor remains and/or returns.

In one embodiment, a (11 1 ln)-24P4C12 antibody is used as an imaging agent in a Phase I human clinical trial in patients having a carcinoma that expresses 24P4C12 (by analogy see, Divgi et al. Natl. Cancer Inst 83:97-104 (1991)).

Patients are followed with standard anterior and posterior gamma camera. The results indicate that primary lesions and metastatic lesions are identified Dose and Route of Administration As appreciated by those of ordinary skill in the art, dosing considerations can be determined through comparison with the analogous products that are in the clinic. Thus, anti-24P4C12 antibodies can be administered with doses in the range of 5 to 400 mg/m 2, with the lower doses used, in connection with safety studies. The affinity of anti-24P4C12 antibodies relative to the affinity of a known antibody for its target is one parameter used by those of skill in the art for determining analogous dose regimens. Further, anti-24P4C12 antibodies that are fully human antibodies, as compared to the chimeric antibody, have slower clearance; accordingly, dosing in patients with such fully human anti-24P4C12 antibodies can be lower, perhaps in the range of 50 to 300 mg/m 2 and still remain efficacious. Dosing in mg/m 2 as opposed to the conventional measurement of dose in mg/kg, is a measurement based on surface area and is a convenient dosing measurement that is designed to include patients of all sizes from infants to adults.

Three distinct delivery approaches are useful for delivery of anti-24P4C12 antibodies. Conventional intravenous delivery is one standard delivery technique for many tumors. However, in connection with tumors in the peritoneal cavity, such as tumors of the ovaries, biliary duct, other ducts, and the like, intraperitoneal administration may prove favorable for obtaining high dose of antibody at the tumor and to also minimize antibody clearance. In a similar manner, certain solid tumors possess vasculature that is appropriate for regional perfusion. Regional perfusion allows for a high dose of antibody at the site of a tumor and minimizes short term clearance of the antibody.

Clinical Development Plan (CDP) Overview: The CDP follows and develops treatments of anti-24P4C12 antibodies in connection with adjunctive therapy, monotherapy, and as an imaging agent. Trials initially demonstrate safety and thereafter confirm efficacy in repeat doses. Trails are open label comparing standard chemotherapy with standard therapy plus anti-24P4C12 antibodies. As will WO 2004/050828 PCT/US2002/038264 be appreciated, one criteria that can be utilized in connection with enrollment of patients is 24P4C12 expression levels in their tumors as determined by biopsy As with any protein or antibody infusion-based therapeutic, safety concems are related primarily to cytokine release syndrome, hypotension, fever, shak ng, chills; (ii) the development of an immunogenic response to the material development of human antibodies by the patient to the antibody therapeutic, or HAHA response); and, (iii) toxicity to normal cells that express 24P4C12. Standard tests and follow-up are utilized to monitor each of these safety concerns.

Anti-24P4C12 antibodies are found to be safe upon human administration.

Example 41: Human Clinical Trial Adiunctive Therapy with Human Anti.24P4C12 Antibody and Chemotherapeutic Agent A phase I human clinical trial is initiated to assess the safety of six intravenous doses of a human anti-24P4C12 antibody in connection with the treatment of a solid tumor, a cancer of a tissue listed in Table I. In the study, the safety of single doses of anti-24P4C12 antibodies when utilized as an adjunctive therapy to an antineoplastic or chemotherapeutic agent as defined herein, such as, without limitation: cisplatin, topotecan, doxorubicin, adriamycin, taxol, or the like, is assessed. The trial design includes delivery of six single doses of an anti-24P4C12 antibody with dosage of antibody escalating from approximately about 25 mg/m 2 to about 275 mg/m2 over the course of the treatment in accordance with the following schedule: Day0 Day 7 Day14 Day 21 Day28 mAb Dose 25 75 125 175 225 275 mg/m 2 mg/m 2 mg/m 2 mg/m 2 mg/m 2 mg/m 2 Chemotherapy (standard dose) Patients are closely followed for one-week following each administration of antibody and chemotherapy. In particular, patients are assessed for the safety concerns mentioned above: cytokine release syndrome, hypotension, .fever, shaking, chills; (ii) the development of an immunogenic response to the material development of human antibodies by the patient to the human antibody therapeutic,.or HAHA response); and, (iii) toxicity to normal cells that express 24P4C12. Standard tests and follow-up are utilized to monitor each of these safety concerns. Patients are also assessed for clinical outcome, and particularly reduction in tumor mass as evidenced by MRI or other imaging.

The anti-24P4C12 antibodies are demonstrated to be safe and efficacious, Phase II trials confirm the efficacy and refine optimum dosing.

Example 42: Human Clinical Trial: Monotherapy with Human Anti-24P4C12 Antibody Anti-24P4C12 antibodies are safe in connection with the above-discussed adjunctive trial, a Phase II human clinical trial confirms the efficacy and optimum dosing for monotherapy. Such trial is accomplished, and entails the same safety and outcome analyses, to the above-described adjunctive trial with the exception being that patients do not receive chemotherapy concurrently with the receipt of doses of anti-24P4C12 antibodies.

Example 43: Human Clinical Trial: Diagnostic Imaging with Anti-24P4C12 Antibody Once again, as the adjunctive therapy discussed above is safe within the safety criteria discussed above, a human clinical trial is conducted concerning the use of anti-24P4C12 antibodies as a diagnostic imaging agent. The protocol is WO 2004/050828 PCT/US2002/038264 designed in a substantially similar manner to those described in the art, such as in Divgi etal. J. Natl. Cancer Inst. 83:97-104 (1991). The antibodies are found to be both safe and efficacious when used as a diagnostic modality.

Example 44: Homology Comparison of 24P4C12 to Known Sequences The 24P4C12 protein of Figure 3 has 710 amino acids with calculated molecular weight of 79.3 kDa, and pl of 8.9.

Several variants of 24P4C12 have been identified, including 4 SNPs (namely v.1, v.3, v.5, v.6) and 3 splice variants (namely v.7, v.8 and v.9) (figures 10 and 11). 24P4C12 variants v.3, v.5, and v.6 differ from 24P4C12 v.1 by 1 amino acid each, at aa positions 187, 326 and 436, respectively. Variant v.7 carries a deletion of 111 aa long starting at aa 237, while variant v.8 and v.9 contain insertions at aa 642 and 378, respectively. The 24P4C12 protein exhibits homology to a previously cloned human gene, namely NG22 also known as chorine transporter-like protein 4 (gi 14249468). It shows 99% identity and 99% homology to the CTL4 protein over the length of that protein (Figure 24P4C12 is a multi-transmembrane protein, predicted to carry 10, 11 or 13 transmembrane domains. Bioinformatic analysis indicates that the 24P4C12 protein localizes to the plasma membrane with some endoplasmic reticulum localization (see Table Recent evidence indicates that the 24P4C12 protein is a 10 transmembrane protein that localizes to the cell surface (O'Regan S et al PNAS 2000, 97:1835).

Choline as an essential component of cell membranes that plays an important role in cell integrity, growth and survival of normal and tumor cells. Choline accumulates at increased concentration in tumor cells relative to their normal counterparts and as such constitutes a tool for the detection of cancer cells by magnetic resonance imaging (Kurhanewicz J et al, J Magn Reson Imaging. 2002.). In addition to its role in maintaining membrane integrity, choline mediates signal transduction event from the membrane to the nucleus (Spiegel S, Milstien S. J Membr Biol. 1995, 146:225). Choline metabolites include sphingosylphosphorylcholine and lysophosphatidylcholine, both of which activate G-protein coupled receptors (Xu F et al Biochim Biophys Acta 2002, 1582:81). In addition, choline results in the activation of kinase pathways including Raf-1 (Lee M, Han SS, Cell Signal 2002, 14:373.). Choline also plays a role in regulating DNA methylation and regulation of gene expression. For example, choline methabolites regulate the expression of cytokines and chemokines essential for tumor growth (Schwartz BM et al, Gynecol Oncol. 2001,81:291; Denda A et al, Carcinogenesis. 2002, 23:245).

Due to its effect on cell signaling and gene expression, choline controls cell growth and survival (Holmes-McNary MQet al, J Biol Chem. 2001, 276:41197; Albright et al, FASEB 1996,10:510). Choline deficiency results in cell death, apoptosis and transformation, while accumulation of choline is associated with tumor growth (Zeisel S et al, Carcinogenesis 1997, 18:731).

,Accordingly, when 24P4C12 functions as a regulator of tumor formaton, cell proliferation, invasion or cell signaling, 24P4C12 is used for therapeutic, diagnostic, prognostic and/or preventative purposes.

Example 45: Identification and Confirmation of Potential Signal Transduction Pathways Many mammalian proteins have been reported to interact with signaling molecules and to participate in regulating signaling pathways. (J Neurochem. 2001; 76:217-223). In particular, choline have been reported to activate MAK cascades as well as G proteins, and been associated with the DAG and ceramide and sphingophosphorylcholine signaling pathway (Cummings et al, above). In addition, choline transmit its signals by regulating choline-kinase and phospholipase activity, 'resulting in enhance tumorigenic effect (Ramirez et al, Oncogene. 2002, 21:4317; Lucas et al, Oncogene. 2001, 20:1110; Chung T et al, Cell Signal. 2000, 12:279).

Using immunoprecipitation and Western blotting techniques, proteins are identified that associate with 24P4C12 and mediate signaling events. Several pathways known to play a role in cancer biology can be regulated by 24P4C12, including phospholipid pathways such as P13K, AKT, etc, adhesion and migration pathways, including FAK, Rho, Rac-1, etc, as well as mitogenicfsurvival cascades such as ERK, p38, etc (Cell Growth Differ. 2000,11:279; J Biol Chem. 1999.

274:801; Oncogene. 2000, 19:3003; J. Cell Biol. 1997, 138:913). Using Western blotting and other techniques, the ability of WO 2004/050828 PCT/US2002/038264 24P4C12 to regulate these pathways is confirmed. Cells expressing or lacking 24P4C12 are either left untreated or stimulated with cytokines, androgen and anti-integrin antibodies. Cell lysates are analyzed using anti-phospho-specific antibodies (Cell Signaling, Santa Cruz Biotechnology) in order to detect phosphorylation and regulation of ERK, p38, AKT, P13K, PLC and other signaling molecules.

To confirm that 24P4C12 directly or indirectly activates known signal transduction pathways in cells, luciferase (luc) based transcriptional reporter assays are carried out in cells expressing individual genes. These transcriptional reporters contain consensus-binding sites for known transcription factors that lie downstream of well-characterized signal transduction pathways. The reporters and examples of these associated transcription factors, signal transduction pathways, and activation stimuli are listed below.

1. NFkB-luc, NFkB/Rel; Ik-kinase/SAPK; growthlapoptosislstress 2. SRE-luc, SRF/TCFiELK1; MAPKISAPK; growthldifferentiation 3. AP-1-luc, FOS/JUN; MAPK/SAPK/PKC; growth/apoptosis/stress 4. ARE-luc, androgen receptor; steroids/MAPK; growth/differenliation/apoptosis p53-luc, p53; SAPK; growth/differentiation/apoptosis 6. CRE-luc, CREB/ATF2; PKA/p38; growth/apoptosis/stress 7. TCF-luc, TCF/Lef; I-catenin, Adhesion/invasion Gene-mediated effects can be assayed in cells showing mRNA expression. Luciferase reporter plasmids can be introduced by lipid-mediated transfection (TFX-50, Promega). Luciferase activity, an indicator of relative transcriptional activity, is measured by incubation of cell extracts with ludferin substrate and luminescence of the reaction is monitored in a luminometer.

Signaling pathways activated by 24P4C12 are mapped and used for the identification and validation of therapeutic targets. When 24P4C12 is involved in cell signaling, it is used as target for diagnostic, prognostic, preventative and/or therapeutic purposes.

Example 46: 24P4C12 Functions as a Choline transporter Sequence and homology analysis of 24P4C12 indicate that 24P4C12 carries a transport domain and that 24P4C12 functions as a choline transporter. In order to confirm that 24P4C12 transports choline, primary and tumor cells, includeing prostate, colon, bladder and lung lines, are grown in the presence and absence of 3 H-choline. Radioactive choline uptake is measured by counting incorporated counts per minutes (cpm). Parental 24P4C12 negative cells are compared to 24P4C12expressing cells using this and similar assays. Similarly, parental and 24P4C12-expressing cells can be compared for choline content using NMR spectroscopy. These assay systems can be used to identify small molecules and antibodies that interfere with choline uptake and/or with the function of 24P4C12.

Thus, compounds and small molecules designed to inhibit 24P4C12 function and downstream signaling events are used for therapeutic diagnostic, prognostic and/or preventative purposes.

Example 47: Regulation of Transcription The cell surface localization of 24P4C12 and its ability to regulate DNA methylation indicate that it is effectively used as a modulator of the transcriptional regulation of eukaryotic genes. Regulation of gene expression is confirmed, e.g., by studying gene expression in cells expressing or lacking 24P4C12. For this purpose, two types of experiments are performed.

WO 2004/050828 PCT/US2002/038264 In the first set of experiments, RNA from parental and 24P4C12-expressing cells are extracted and hybridized to commercially available gene arrays (Clontech) (Smid-Koopman E et al. Br J Cancer. 2000. 83:246). Resting cells as well as cells treated with FBS, pheromones, or growth factors are compared. Differentially expressed genes are identified in accordance with procedures known in the art. The differentially expressed genes are then mapped to biological pathways (Chen Ketal. Thyroid. 2001. 11:41.).

In the second set of experiments, specific transcriptional pathway activation is evaluated using commercially available (Stratagene) luciferase reporter constructs including: NFkB-luc, SRE-luc, ELK1-luc, ARE-luc, p53-luc, and CRE-luc.

These transcriptional reporters contain consensus binding sites for known transcription factors that lie downstream of wellcharacterized signal transduction pathways, and represent a good tool to ascertain pathway activation and screen for positive and negative modulators of pathway activation.

Thus, 24P4C12 plays a role in gene regulation, and it is used as a target for diagnostic, prognostic, preventative and/or therapeutic purposes.

Example 48: Involvement in Tumor Progression The 24P4C12 gene can contribute to the growth of cancer cells. The role of 24P4C12 in tumor growth is confirmed in a variety of primary and transfected cell lines including prostate, and bladder cell lines, as well as NIH 3T3 cells engineered to stably express 24P4C12. Parental cells lacking 24P4C12 and cells expressing 24P4C12 are evaluated for cell growth using a well-documented proliferation assay (Fraser SP, et al., Prostate 2000;44:61, Johnson DE, Ochieng J, Evans SL. Anticancer Drugs. 1996, 7:288). Such a study was performed on prostate cancer cells and the results are shown in figure 28. The growth of parental PC3 and PC3-24P4C12 cells was compared in low and 10% FBS. Expression of 24P4C12 imparted a growth advantage to PC3 cells grown in 10% FBS. Similarly, expression of 24P4C12 in NIH-3T3 cells enhances the proliferation of these cells relative to control 3T3-neo cells. The effect of 24P4C12 can also be observed on cell cycle progression. Control and 24P4C12-expressing cells are grown in low serum ovemight, and treated with 10% FBS for 48 and 72 hrs. Cells are analyzed for BrdU and propidium iodide incorporation by FACS analysis.

To confirm the role of 24P4C12 in the transformation process, its effect in colony forming assays is investigated.

Parental NIH-3T3 cells lacking 24P4C12 are compared to NIH-3T3 cells expressing 24P4C12, using a soft agar assay under stringent and more permissive conditions (Song Z. et al. Cancer Res. 2000;60:6730).

To confirm the role of 24P4C12 in invasion and metastasis of cancer cells, a well-established assay is used. A non-limiting example is the use of an assay which provides a basement membrane or an analog thereof used to detect whether cells are invasive a Transwell Insert System assay (Becton Dickinson) (Cancer Res. 1999; 59:6010)). Control cells, including prostate, and bladder cell lines lacking 24P4C12 are compared to cells expressing 24P4C12. Cells are loaded with the fluorescent dye, calcein, and plated in the top well of a support structure coated with a basement membrane analog the Transwell insert) and used in the assay. Invasion is determined by fluorescence of cells in the lower chamber relative to the fluorescence of the entire cell population.

24P4C12 can also play a role in cell cycle and apoptosis. Parental cells and cells expressing 24P4C12 are compared for differences in cell cycle regulation using a well-established BrdU assay (Abdel-Malek ZA. J Cell Physiol.

1988, 136:247). In short, cells are grown under both optimal (full serum) and limiting (low serum) conditions are labeled with BrdU and stained with anti-BrdU Ab and propidium iodide. Cells are analyzed for entry into the G1, S, and G2M phases of the cell cycle. Alternatively, the effect of stress on apoptosis is evaluated in control parental cells and cells expressing 24P4C12, including normal and tumor prostate, colon and lung cells. Engineered and parental cells are treated with various chemotherapeutic agents, such as etoposide, flutamide, etc, and protein synthesis inhibitors, such as cycloheximide. Cells WO 2004/050828 PCT/US2002/038264 are stained with annexin V-FITC and cell death is measured by FACS analysis. The modulation of cell death by 24P4C12 can play a critical role in regulating tumor progression and tumor load.

When 24P4C12 plays a role in cell growth, transformation, invasion or apoptosis, it is used as a target for diagnostic, prognostic, preventative and/or therapeutic purposes.

Example 49: Involvement In Angiogenesis Angiogenesis or new capillary blood vessel formation is necessary for tumor growth (Hanahan D, Folkman J. Cell.

1996, 86:353; Folkman J. Endocrinology. 1998 139:441). Based on the effect of phsophodieseterase inhibitors on endothelial cells, 24P4C12 plays a role in angiogenesis (DeFouw L et al, Microvasc Res 2001, 62:263). Several assays have been developed to measure angiogenesis in vitro and in vivo, such as the tissue culture assays endothelial cell tube formation and endothelial cell proliferation. Using these assays as well as in vitro neo-vascularization, the role of 24P4C12 in angiogenesis, enhancement or inhibition, is confirmed.

For example, endothelial cells engineered to express 24P4C12 are evaluated using tube formation and proliferation assays. The effect of 24P4C12 is also confirmed in animal models in vivo. For example, cells either expressing or lacking 24P4C12 are implanted subcutaneously in immunocompromised mice. Endolhelial cell migration and angiogenesis are evaluated 5-15 days later using immunohistochemistry techniques. 24P4C12 affects angiogenesis and it is used as a target for diagnostic, prognostic, preventative and/or therapeutic purposes.

Example 50: Involvement in Adhesion Cell adhesion plays a critical role in tissue colonization and metastasis. The presence of leucine rich and cysteine rich motifs in 24P4C12 is indicative of its role in cell adhesion. To confirm that 24P4C12 plays a role in cell adhesion, control cells lacking 24P4C12 are compared to cells expressing 24P4C12, using techniques previously described (see, Haier et al, Br. J. Cancer. 1999, 80:1867; Lehr and Pienta, J. Nail. Cancer Inst 1998, 90:118). Briefly, in one embodiment, cells labeled with a fluorescent indicator, such as calcein, are incubated on tissue culture wells coated with media alone or with matrix proteins. Adherent cells are detected by fluorimetric analysis and percent adhesion is calculated. This experimental system can be used to identify proteins, antibodies and/or small molecules that modulate cell adhesion to extracellular matrix and cell-cell interaction. Since cell adhesion plays a critical role in tumor growth, progression, and, colonization, the gene involved in this process can serves as a diagnostic, preventative and therapeutic modality.

Example 51: Detection of 24P4C12 protein in cancer patient specimens To determine the expression of 24P4C12 protein, specimens were obtained from various cancer patients and stained using an affinity purified polyclonal rabbit antibody raised against the peptide encoding amino acids 1-14 of 24P4C12 variant 1 and conjugated to KLH (See, Example 10: Generation of 24P4C12 Polyclonal Antibodies.) This antiserum exhibited a high titer to the peptide (>10,000) and recognized 24P4C12 in transfected 293T cells by Western blot and flow cytometry (Figure 24) and in stable recombinant PC3 cells by Western blot and immunohistochemistry (Figure 25). Formalin fixed, paraffin embedded tissues were cut into 4 micron sections and mounted on glass slides. The sections were dewaxed, rehydrated and treated with antigen retrieval solution (0.1M Tris, pH1 0) at high temperature. Sections were then incubated in polyclonal rabbit anti-24P4C12 antibody for 3 hours. The slides were washed three times in buffer and further incubated with DAKO EnVision+ T M peroxidase-conjugated goat anti-rabbit immunoglobulin secondary antibody (DAKO Corporation, Carpenteria, CA) for 1 hour. The sections were then washed in buffer, developed using the DAB kit (SIGMA Chemicals), counterstained using hematoxylin, and analyzed by bright field microscopy. The results showed expression of 24P4C12 in cancer patients' tissue (Figures 29 and 30). Tissue from prostate cancer patients showed expression of 24P4C12 in the Stumor cells and in the prostate epithelium of tissue normal adjacent to tumor (Figure 29).

Generally, expression of 24P4C12 was high in all prostate tumors and was expressed mainly around the cell membrane indicating that 24PC12 ismembrane associated in prostate tissues.

All of the prostate samples tested were positive for 24P4C12. Other tumors that were positive for 24P4C12 included heart, skeletal muscle, liver, brain, spinal cord, skin, adrenal, lymph node, spleen, salivary gland, small intestine and placenta. None demonstrated any expression of 24P4C12 by immunohistochemistry. Normal adjacent to tumor tissues were also suited to determine the presence of 24P4C12 protein by immunohistochemistry. These included, breast, lung, colon, ileum, bladder, kidney and pancreas. In some of the tissues from these organs there was weak expression of 24P4C12. this expression may relate to the fact that the samples were not truly normal and may indicate a precancerous change. The ability to identify malignancy in tissue that has not undergone obvious morphological changes is an important diagnostic modality for cancerous and precancerous conditions.

These results indicate that 24P4C12 is a target for diagnostic; prophylactic, prognostic and therapeutic applications in cancer.

Throughout this application, various website data contact, publications, patent applications and patents are referenced. (Websites are referenced by their Uniform Resource Locator, or URL, addresses on the World Wide Web).

The present invention is not to be limited in scope by the embodiments disclosed herein, which are intended as single illustrations of individual aspects of the invention, and any that are functionally equivalent are within the scope of the invention. Various modifications to the models and methods of the invention, in addition to those described herein, will become apparent to those skilled in the art from the foregoing description and teachings, and are similarly intended to fall within the scope of the invention. Such modifications or other embodiments can be practiced without departing from the true scope and spirit of the invention.

Throughout this specification the word "comprise", or variations such as "comprises" or "comprising", will be understood to imply the inclusion of a stated element, integer or step, or group of elements, integers or steps, but not the exclusion of any other element, integer or step, or group of elements, integers or steps.

Any discussion of documents, acts, materials, devices, articles or the like which has been included in the present specification is solely for the purpose of providing a context for the present invention. It is not to be taken as an admission that any or all of these matters form part of the prior art base or were common general knowledge in the field relevant to the present invention as it existed before the priority date of each claim of this application.

WO 2004/050828 WO 204/00828PCTIUS2002/038264

TABLES:

TABLE 1: Tissues that Express 241140 2: a. Malignant Tissues Prostate Blad der Kdney Lung Colon Ovary Breast Uterus Stomach TABLE fl: Amino Acid Abbreviations SINGLE LETTER THREE LETTER FULL NAME Phe phlenylalanine Leu leucine S Ser serine Y Tyr tyrosine C Cys cysteine W Trp tryptophan P Pro praline H His hisfidine Q Gin glutamine R Arg arginine lie isoleucine M Met methionine T Thr threonine N Asn asparagine K Lys lysine V Val valine A Ada alanine D Asp aspartc acid E Glu glutamicacdd G Gly glycine W¥O 2004/050828 PCTIUS2002/038264 TABLE III: Amino Acid Substitution Matrix Adapted from the GCG Software 9.0 BLOSUM62 amino acid substitution matrix (block substitution matrix). The higher the value, the more likely a substitution is found in related, natural proteins. (See world wide web URL ikp.unibe.chlmanuallblosum62.html) A C D E F G H I K L M N P Q R S T V W Y.

4 0 -2 -1 -2 0 -2 -1 -1 -1 -1 -2 -1 -1 -1 1 0 0 -3 -2 A 9 -3 -4 -2 -3 -3 -1 -3 -1 -1 -3 -3 -3 -3 -1 -1 -1 -2 -2 C 6 2 -3 -1 -1 -3 -1 -4 -3 1 -1 0 -2 0 -3 -4 -3 D -3 -2 0 -3 1 -3 -2 0 -1 2 0 0 -1 -2 -3 -2 E 6 -3 -1 0 -3 0 0 -3 -4 -3 -3 -2 -2 -1 1 3 F 6 -2 -4 -2 -4 -3 0 -2 -2 -2 0 -2 -3 -2 -3 G 8 -3 -1 -3 -2 1 -2 0 0 -1 -2 -3 -2 2 H 4 -3 2 1 -3 -3 -3 -3 -2 -1 3 -3 -1 I -2 -1 0 -i 1 2 0 -1 -2 -3 -2 K 4 2 -3 -3 -2 -2 -2 -1 1 -2 -1 L -2 -2 0 -1 1 -1 1 -1 -1 M 6 -2 0 0 1 0 -3 -4 -2 N 7 -1 -2 -1 -i -2 -4 -3 P 1 0 -i -2 -2 -1 Q -1 -1 -3 -3 -2 R 4 1 -2 -3 -2 S 0 -2 -2 T 4 -3 -1 V 11 2 W 7 Y WO 20041050828 WO 204100828PCT/US20021038264 TABLE IV: HL.A Class 11111 MotifslSupermotifs TABLE IV HIA Class I Supermotifsl~otifs SUPERMOTIF POSITION POSITION POSITION 2 (Primary Anchor) 3 (Primary Anchor) C Terminus (Primary Anchor) Al TIL VMS FWY A2 LIVMATQ IVMATL A3 VSMATLI RK A24 YFWIVLMT FIYWLM B7 P VILFMWYA B27 RHK FYLWMIVA B44 ED FWYLIMVA B58 ATS FWYLIVAA B62 QL1VMP FWYMIVLA Al TSM Y Al DEAS Y A2.1 LMVQfAT VLIMAT A3 LMVISATFCGEJ KYRHFA All VrMLISAGNCDF KRYH A24 YFWM FLIW A*31101 MVTALIS RK Ak3301 MVALFIST RK A*6801 AVTMSL1 RK 63*0702 F LMFWYAIV B3*3501 P LMFWYIVA 851 P LIVFVVYAM B35301 P IMFWYAL V 2*5401 P ATIVLMFWY Bolded residues are preferred, italicized residues are less preferred: A peptide is considered motlf-bearng if it has primary anchors at each primary anchor position for a motif or supermotif as specif ed in the above table.

TABLE IV HLA Class 11 Supermotif 1 6 9 W, F, Y, VI,L AV,1, L, P,C, S,T A, V,1, LC, S,T,M, Y WO 2004/050828 WO 204/00828PCT/US2002/038264 TABLE IV I-LA Class 11 Motifs MOTIFS 1 'anchorl1 2 3 4 5 P'anchor6 7 8 9 DR4 preferred FMYLiVW M T I VSTCPALIM MH MH deleterious Wv R WDE DRi preferred MFLIVWY PAMQ VMATSPUIC M AVM deilterious C CH EID CWD GDE D DR7 preferred MFLIVWY M W A IVMSACTPL M IV deleterious C G GRID N G R3MOTIFS Il*anchorl1 2 3 1l'anchor 4 5 1 anchor 6 Motif a preferred LIVMEY D Motif b preferred LIVMFAY DNQEST KRH DR Supennotif MFLI/WY VMSTACPLI Italicized residues indicate less preferred or 'tolerated' residues TABLE IV HLA Class I Supermotifs POSITION: 1 2 3 4 5 6 7 8 C-terminus

SUPER-

MOTiFS Al 10 Anchor 10 Anchor TIL VMS FWY A2 1' Anchor 10 Anchor LIVMATQ LIVMAT A3 Preferred 10* Anchor YEW YEW YEW P 1' Anchor VSMATLI (415) (3T5) RK deleterious DE (315); DE P A24 1' Anchor 10* Anchor YEWIVLMT F[YWVLM 87 Preferred FWY 11' Anchor FW'( FWA' 11'Anchor LIVM P

VILFMWYA

deleterious DE DE G QN DE P(516); (415) G(415); A(315); B27 1* Anchor I 'Anchor RHK FYLWMIVA B44 1'Anchor 11'Anchor ED

EWYLIMVA

B58 1' Anchor 10* Anchor ATS FWYLIVAA B62 1' Anchor 10 Anchor OLIVMP FWYMIVLA Italicized residues indicate less preferred or 'tolerated" residues WO 2004/050828 WO 204/00828PCTIUS2002/038264 TABLE IV HLA Class I Motifs POSITION 1 3 4 5 6 7 8 9 Cterminus or C-terminus Al preferred GFYWV l'Anchor DEA YFW P DEQN YEW I *Anchor 9-mer STM y deleterious DE RI-KLIVMF A G A Al preferred GRHK ASTCLIVM 1 0 Ancior GSTC ASTO LIVM DE 1 Anchor 9-mer DEAS Y deleterious A RHKIDEPYFWV DE PQN RHK PG GP Al preferred YFW I *Anchor DEAQIN A YFWQN PASTC GDE P 1PAnchor STM

Y

mer deleterious GP RHKGLIVNI DE RHK QONA RHKYFW RI-K A Al preferred YFW STCLIVM l 0 Anchor A YEW PG G YEW l 0 Anchor DEAS y mer deleterious RHK RHKDEPYFW P G PRHK QN A2.1 preferred YFW I 0 Anchor YEW STC YEW A P 1 *Anchor 9-mer LMIVQAT

VLMAT

deleterious DEP DERKH RKH DERKH POSITION: 1 2 3 4 5 6 7 8 9 C- Terminus A2.1 preferred AYFW 1 0 Anchor [VIM G G EYWNL 1 0 Anchor [MI VQAT vim VLIMAT mer deleterious DER DE RKHA P RKH DERK RKH

H

A3 preferred RHK I 0 Anchor YEW PRH-KYF A YFW P l 0 Anchor LMVISATECGD W

KYRHFA

deleterious DEP DE All preferred A 1 Anchor YFW YEW A YFW YFWV P 1*Anchor \/TLMISAGN CD

KRYH-

F

deleterious DEP A G A24 preferred YFWVRHK 1 0 Ancor STO YEW YEW 1 *Anchor 9-mer YFWM

ELIW

deleterious DEG DE G QNP DERH G AQN

K

A24 Preferred 1 'Anchor P YFWP P 1 0 Anchor YFWAA

FLIW

mer Deleterious GDE QN RHI< IE A QN DEA A310 Preferred RHK l'Anchor YEW P YEW YFW AP 1'Anchor 1 MYTALIS

RK

Deleterious DEP DE ACE IDE DE DE A330 Preferred I *Anchor YEW AYFW 1 0 Anchor 1 MVALFIST

RK

DeleteriousGP DE A680 Preferred YFWSTC l'Anchor YFWLIV YEW P 1 *Anchor I AVTAISLI M RK deleterious OP DEG RHK A B070 Preferred RHKFWY l 0 Arichor RHK RHK RHK RHK, PA 1 *Anchor 2 P [ME WYAI

V

deleterious DEONP DEP DE DE GDE QN DE WO 2004/050828 PCT/US2002/038264 POSITION 1 2 3 4 5 6 7 8 9 C terminus or C-terminus All preferred GFYW l 0 Anchor DEA YFW P DEOIN YFWV l 0 Anchor 9-mer STM Y deleterious LJE RHKLIVMP A G A Al preferred GRHK ASTCLIVM 1 0 Anchor GSTC ASTC IVM DE I 'Anchor 9-mer IDEAS Y deleterious A RHKDEPYFW DE PON RHK PG GP B350 Preferred FWYLIVM l 0 Anchor FWY FW 1 Anchor 1 P LMFWYIV

A

deleterious AGP G G B51 Preferred LIVMFWY I'Anchor MWY STC EWY G FWY i*Anchor P LIVFWY'A

M

deleterious AGPIDER DIE G DEON GDE

HKSTC

2530 preferred LIVMFWY I 'Anchor MWY STC FWY LIVMFW FWY l 0 Anchor 1 P Y IMFWYAL

V

deleterious A$3PQN G RHKQN DE B540 preferred FWY 1 0 Anchor FWYIVM LIVM ALIVM FWYA I*Anchor 1 P P ATIVLMF deleterious GPQNDE GDESTC RI-KDE DE QNDGE DE WO 2004/050828 WO 204/00828PCTIUS2002/038264 TABLE IV Ounaof Hsues Overall Dhenotvaic freauencies of HLA-snnn,1vn~ in diffpmnl ~thnir nnn~i~iinn~ ____specificity Phenoty ic frequeno upertype osition 2 CJ!Termirnu Caucasian Black a anese Chinese Hispanic veracl 7 pILMVFWY3.2 5.1 7.1 43.0 49.3 49.5 3 ILMVST RK 37.5 2.1 5.8 2.U7 4-3.1 44.2 ____ILMVT AILMVT 4.8 9.0 F2.4 45.9 43.0 42.2 4 F (WIVLMTtF ACsWLM) 23.9 8.9 8.6 40.1 38.3 40.0 B44 E (Dy, -WYLIMV !3.0 1.2 2.9 39.1 .9.0 37.0 1 TI(LVMS) FtWY 47.1 16.1 1.8 14.7 6.3 5. 2 27 RHK 2LWI 8.4 6.1 133 13.9 5.3 23,4 62 -QL (IVMP) Y,*(MIV 12.6 .8 86.5 5.%4 11.1 18.1 868 ATS :W LV 10.0 .1 .6 9.0 .9 10.3 TABLE IV Calculated population coverage afforded by different HLA-supete combinations HLA-supertypes Phenotypic frequency Caucasian J. lcs apanese Chinese ipnc vrg 33.0 86,1 87.5 8.4 86.3 86.2 A3 and 137 9.5 98.1 100.0 99.5 9.4 9.3 AA3, 87, A24, 844 39.9 99.6 100.0 9.8 9.9 99.8 and A, A3, B7, A24, B44, Al, B27, B62, and B 58 Motifs indicate the residues defining supertype specificites. The motifs incorporate residues determined on the basis of Published data to be recognized by multiple alleles within the supertype. Residues within brackets are additional residues Iso predicted to be tolerated by multiple alleles within the supertype.

Table V: Frequently Occurring Motifs Name avg Description Dotential Function Nlucleic acid-binding protein functions as ransciption factor, nuclear location z-C2H-2 34% Zinc finger, C21-2 type )robable Cytochrome b(N- membrane bound oxidase, generate cytochrome _b N 68% terminal)/b6/petB superoxide domains are one hundred amino acids long and include a conserved Ig19% Immunoglobulin domain intradomain disulfide bond.

tdem repeats of about 40 residues, Dach containing a Trp-Asp motf.

:unction in signal transduction and 18% WD domain, G-beta repeal rotein interaction 'ay function in targeting signaling PDZ 23% PDZ domain -olecules to sub-memibranous sites LRR 28% Leucine Rich Repeat hort sequence motifs involved in interactions )nserved catalytic core common to Doth serinelttireonine and tyrosine )rotein kinases containing an ATP jPkinase 23% Potein kinase domain inding site and a catalytic site WO 2004/050828 PCTIUS2002/038264 3leckstrin homology involved in ntracellular signaling or as constituents PH 16% PH domain )f the cytoskeleton 3040 amino-acid long found in the ,xtracellular domain of membrane- EGF 34% EGE-like domain bound proteins or in secreted proteins Reverse transcriptase (RNA-dependent DNA Rvt 49% polymerase) Cytoplasmic protein, associates integral nk 25% Jk repeat -ernbrane proteins to the cytoskeleton NADH- neinbrane associated. Involved in Ubiquinonelplasloquinone Droton translocation across the Oxidoredgql 32% (complex various chains Tembrane Iacium-binding domain, consists of al 2 'esidue loop flanked on both sides by a 2thand 24% EF hand 12 residue alpha-helical domain Retroviral aspartyl kspartyl or acid proteases. centered on VP 79% protease icatalytic aspary residue xtracellular structural proteins in volved -n formation of connective tissue. The Collagen triple helix repeat sequence consists of the G-X-Y and the Collagen, 42% (20 copies) olypeptide chains forms a triple helix.

Located in the extracellular ligandbinding region of receptors and is about 200 amino acid residues long with two pairs of cystaines involved in disulfide Fn3 20% lFibronectin type Ill domain 3nds 3even hydrophobic transmembrane -egions, With the N-terminus located 7transmemrbrane receptor axtracellutarly while the C-terminus is 7tm-1 19% (rhodopsin family) ypasi. Signal through G proteins Table VI: Motifs and Post-translational Modifications of 24P4C1 2 N-glycosylation site 29 -32 NIRSO (SEQ ID NO: 48) 69 -72 NSTG (SEQ ID NO: 49) 155-158 NMVTV (SEQ ID NO: 197-200 NDTT (SEQ ID NO: 51) 298- 301 NLSA (SEQ ID NO: 52) 393- 396 NISS (SEQ ID NO: 53) 405 -408 NTSC (SEQ ID NO: 54) 416-419 NSSC (SEQ ID NO: 678- 681 NGSL (SEQ ID NO: 56) Protein kinase C phosphorylation site 22 -24 SfR 218 -220 SvK 430 -432 SsK 494-496 TIR 573 575 SaK 619 -621 SgR Casein kinase 11 phosphorylation site 31 34 SCTD (SEQ ID NO: 57) 102 105 SVAE (SEC ID NO: 58) 119 -122 SCPDE (SEQ ID NO: 59) 135 -138 1VGE (SEQ ID NO: 304 -307 SVQE (SEQ ID NO: 61) WO 2004/050828 WO 204/00828PCT/US2002/038264 Tyrosine kinase pliosplorylation site 6 -13 RDEDDEAY (SEQ ID NO: 62) N-myristoylation site 72 -77 GAYCGM (SEQ ID NO: 63) 76 -81 GMGENK (SEQ ID NO: 64) 151 -156 GVPWNM (SEQ ID NO: 207-212 GLIDSL (SEQ ID NO: 66) 272-277 GIYYCW (SEQ ID NO: 67) 287- 292 GASISQ (SEQ ID NO: 68) 349- 354 GQMMST (SEQ ID NO: 69) .449- 454 GLFWTL (SEQ ID NO: 467-472 GAFASF (SEQ ID NO: 71) Amidaion site 695 698 IGKIK (SEQ ID NO: 72) Leucine zipper pattern 245 266 LFILLLRLVAGPLVLVLILGVL (SEQ ID NO: 73) Cystelne-rich region 536 547 CIMCCFKCCLWVC (SEQ ID NO. 74) Table VII: Search Peptides Variant 1, 9-mers, 10-mers, 15-mero (SEQ ID

MGGKQRDEDD

PRQVLYPRNS

PEDPWTVGKN

CF'PWTNVT PP VLSLLFI LLL

AYQSVQETWL

TPVLLLICIA

CLMCVFQGYS

DIPTFPLISA

FKCCLWCLE(

FGKLL'vVGGV

MCVDTLFLCF

EAYGKPVKYD

TGAYCGMGEN

EFSQTVGEVF

AL PG ITNDTT

RLVAGPLVLV

AALIVLAVLE

YWAIVTALYLA

SKGLIQRSVF

FIRTLRYETG

Fl KFLNRBAY

GVLSFFFFSG

LEDLERNDGS

PSFRGPIKNR

KDKPYLLYFN

YTKNRNFCLP

IQQGISGLID

LI LGVLGVLA

AILLLMLIFL

TSGOQYVLW

NLQIYGVLGL

SLAFGALIL'

IMIAIYGKNF

RI PGLGKDFK

LDRPYYMSKS

SCTDVICCVL

TFSCILSSNI

GVPWNMTVI T

SLNARDISVK

YGIYYCWEEY

RQRIRIAIAL

AS NISS PGCE

FWTLNWVLAL

LVQIARVILE

CVSAKNAFML

SPHLNYYWLP

LLKILGKKNE

NO:

FLLPILGYIV

I SVAENGLQC S LQQELCPS F I FEDFAQSWY

RVLRDKGASI

LKEASKAVGQ

KVPINTSCNP

GQCVLACAFA

YI DI-KLRGVQ

LMRNIVRVVV

IMTSILGAYV

APPDNKKRK<

VO IVAWLYGD PT PQVCVS Sc

LLPSAPALGR

WI LVALGVAL

SQLGFTTNLS

MM'STMFYPLV

TAHLVN SSC P S FYWAFHKPQ NPVARC IMCC

LDKVTDLLLF

TASGFFSVFG

Variant 3: S -rer s GRCFPWTNITPPALFGI (SEQ ID NO: 76) LGRCFPWTNITPPALPGIT (SEQ ID NO: 77) PSAPALGRCFPWTNITPPALPGITNDTTI (SEQ ID NO: Variant 9 -ier s VLEATLLLVLIFLRQRI (SEQ ID NO: 79) 1O-mersq AVLEAILLLVLIFLRQRIR (SEQ ID NO: ALIVLAVLEAILLLVLIFLRQRIRIAIAL (SEQ ID NO: Variant 6: 9-mer s GYSSKGLIERSVFNLQI (SEQ ID NO: 82) QGYSSKGLIPRSVFNLQTY (SEQ ID NO: 83) WO 2004/050828 LMCVFQGYSSKGLIPRSVFNLOIYGVLGL (SEQ ID NO: 84) Variant 7 9-mers SWYWILVAVGQM4STM (SEQ ID NO: 1O-iners QSWYWILVAVGQMMSTMF (SEQ ID NO: 86) FEDFAQSWYWILVAVGQW9ISTMFYPLVT (SEQ ID NO: 87) Variant 8 9-mer s NYYWLPIMRNPTTPTGIIVFQTSTLGAYV (SEQ TD NO: 08) -me rs LNYYWLPIMRNPITPTGHV'QTSIIrGAYVI (SEQ ID NO: 89) -me rs EIS2ELNYYWLPIMRNPITPTGHVFQTSILGAYVIASGFF (SEQ ID NO: Variant 9 9-mers YWAMTALYPLPTQPATLGYVLWASNI (SEQ TD NO: 91) lO-mers AYWAMTALYPLPTQPATLGYVLWASNIS (SEQ ID NO: 92) LLICIAYWAMTALYPLPTQPATLGYVLWASNISSPGCE (SEQ ID NO: 93) PCT/US2002/038264 WO 2004/050828 Tables VIII XXI: PCT/US20021038264 Table VIII.V-HLA.A1 .9mers.

24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino adds, and the end position for eacn peptide is the start position plus eight.

Startubsequen Score] 58 [YGDPRQVLY ]JN o 662 CVDTLFLCF I 25.000] 77 MGENKDKPY i 11.250 594 VTDLLLFFG 16.250 69811 KNEAPPDNK F4.500I Table VII7-V1-HLA-AI.9mers- 24P4C1 2 Each peptide is a portion of SEQ 10 NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Start Subsequence jfl 593 KVtDLLLFF ]j 0.500 321 AILLLMLIF 0.500 1736 ICCVLFLLF 3 0.500 50 VVGIVAVVLY .0 186 1 NVTPPALPG ]j 0.500 609 GVGVLSFFF j 0.500 287] GASISOLGF 6.556 F 187 1 VTPPALPGI ]J a 500 668j[ LCFLEDLER IW050] I LLUFL 1 0.500 272 GIY CWF1 0.500 521 YIDHKLRGV 253 [VAGPLVLVL 0 500 r-3-9[ GCEKVPINTj 0.450 r338 IALLKEASK 1 4-400] F135] TVGEVFYTK 10.400 349 GQMMSTMFY O75 18 SSCPEDPWT ][T=W VQETWLAAL 11270 i r FKSPHLN 0.250 1 ARDIGVKIF jr- 2507 F7072] PPDNKKRKK 0.250 L64ii IMTSILGAY 0.250 [71 NGSLDRPYV (0150] 513 QIARVILEY IF 02507 1F4831 PTFPLISAF 1 0. 2_5 1120 II CPEDPWTVG 1[ 0.25] [129 KNEFSQTVG 11 0.225 r VGEVFTKN 1121 17 0 FLLPSAPAL II OlO0 [147] FCLPGVPWN 0.200 393 NISSPGCEK 0.200 4674VLAGAFASF ME [1T]I VILYIDHK 110.200 4 ]I CVFQGYSSK II 0.200 3II ISSPGCEKV i 0.150 1 610 11 VGVLSFFFF J0.125 [-6071 VTFVLLLIC ]10.125 1567 MTVITSLQQ 1 0.125 L 77 I NNGSLDRPY 10125 498 1HTGSLFGA 0.125 172] LPSAPALGR 0.125 195 F ITNDTTIQQ ]F1Ti 452 WTLNWVLAL 0.125 353 STMFYPLVT 0.125 443 QIQYGVLGLF 0.100 543 CCLWCLEKF 10100 207 GLIDSLNAR 10100 407[SCNPTAHLV 0.100 180 RCFPWTNVT 354 TMFYPLVTF 0F100 Table VIIIV3-HLA-AI *9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 9 amino adds, and the end position for each peptide is the start position plus eight.

Start ubeene Score WO 2004/050828 PCT/US2002/038264 Table VIII-V3-HLA-Al -9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position lus ei t Start j Subsequence 11 Score 87l IPPALPGI [0500 8 1!NITPPALPG l[ 0.500 LI IRCFPWTNIT 0.100 7 7 WTNITPPAL ta0.0S I 77 TNITPPALP 110.001 GRCFPWTNI .001 7-37 0 P 0000 715 PWTNITPPA ii 0.000 7 74] ITP 0.000 Table VIIIV5-H LA-A -9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eiht.

[tar II Subsequence I LL]I VLEAILLLV [4.500 I ALLLVLIF 0.500 [8I LVLIFLRQR 1 0.100 7 LLVLIFLRQ 11 50 [i3i] EAILLLVLI J[&020J LsVLIFLRQRI 10.0101 E 2 lI LEAILLLVL j 0003 Table VIII-V-HLAAI -Smers-1 24P4C12 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start psto lsegt IStart IISubse uence Score r GLIPRSVFN 1[0.200 SKLP 0. 07 5 [Au KGLIPRSVF 0.025 LU] LIPRSVFNL 0.005 F-3I] SSKGLIPRS 10003 F-471 SKGLIPRSV [o001 F -q FPRSVFNQI 76637 a ]8I11 IPRSVFNLQ 0.000 l GYSSKGLIP 0.000 Table VIII-V7-HLA-Ai-gmers- 24P4C12 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the .start position plus eight.

FStart IStart][I Subseqluence I[ Score I I VAVGQfRIMST it0X)50 IT LVAVGQMMS I[0fl50 I 1 AVGQMMSTM 0.010 1 5 1 ILVAVGQMM 0.010 Liii WILVAVGQM] 05Th 1 YWILVAVGQ j 0.001 I SWYWILVAV lF0T1 2 IIWYWILVAVG I0.000~ Table VIII-VB-HLA-A-9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight Start] Subsequence jf Score 11 ii PITPTGIVF IFi]00 l9 j] FQTSILGAY j[0.075 20_ QTSILGAYV 10050] 17 ]L HVFQTSILG 0.050 12 ][ITPTGHVFQ II 5] 8 1MRNPITPTG 1L 9..0.i 4 1WLPIMRNPI J[50Th L. i1 LPIMRNPIT ]0JJ05] Table VIIIVHLA*AI-9mers- I i 24P4C1 2 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

start ubsequence It Score 10r VFQTSILGA It0.003 10 NPITPTGHV 0003 15[ TGHVFQTSI It 0003 F RNPITPTGH 0.003 1 14 ir PTGHVFTS It 0.003 I F3 IMRNPITPT UK I 3 I1[YWLPIMRNP ILt000T 7Y][YYYT~YLPIMRN !000 I 6[ F1IMRNPITP Table VIII-V-HLA-AI-Smers- Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start pcsition plus ei ht, Start I Subsequence c re ILilt PITPATLGY ii F MTALYPLPT4I7I h j[ IATLGYVLWA II 0.125] F-8 YPLPTQPAT]F -050 5 TALYPLPTQ 0020 2 WAMTALYPL F 527 r 16 T[GYVLWAS 0010 F -6 ALYPLPTP lj0.010 1 1311 QPATLGYVL r i7it LGYVLWASN 00 I 11 PLPTQPATL 0.002 14- jPATLGYLW 0X102 lul ITQPATLGYV F3 ATALYPLP 0.001 r18 iI GYVLWASNI 0.001 7 LYPLPTQPA 0.001 Lii YWAMTALYP II T555 WlO 2004/050828 PCT/US20021038264 Table IX-V1-HLA-A-1Omers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

StartI Subsequence j Score 594 VTDLLLFFGK 1125.000 32 11 CTDVICCVLF 25.000 120 j-CPEDPWTVGK 9.000 518 ILEYIDHKLR II 9.000 Table IX-V1-HLA-All0mers- S24P4CI2 Each peptide is a portion of SEQ ID NO; 3; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

75- If Subsequence ]core 609 GVGVLSFFFF 0500 353 1STMFYPLVTF i asoo 464 ]fVLAGAFASFY 0.500 322 If ILLLMLIFLR 0.500 351[ VICCVLFLLF 0.500 606 V WGGVGVLSF 0.500 521 1 YIDHKLRGVQj[ 0.500 662-1 CVDTLFLCFLI 0.50 661 MCVDTLFLCF 0.500 265 11 VLGVLAYGI Y ]f 0 49 If IVGIVAWLY f 0.500 667 FLFLEDLER 1 0.5001 APZJ SCNPTAHLVN If 5oo0 7-165 ELCPSFLLPS f.500 7777 MGENKDKPYL 7 7547Jf CLEKFIKFLN If-5: 33771 ILKAK .0 F 512 If VQjARVILEY lf 0.7 M I689 K SLLKILGKK 305JVQETWLAALIf P.?Z Zi 1 IKYDPSFRGPI if 0.5 J 6IGMGENKDKPY]J .50~ 75-577 RNAYIMIAIY 0.250 75-907 VLDKVTDLLL Ifoio 677 SLOPY 0.250 57 IFMLLMRNIVR I 0.250 18711 VTPPALPGIT II Q250] 7 463I CVLAGAFASF I 0200] F-5-1671 RVILEYIDHK 1f 0.200 774 02YCGMGENKDKj200 [1 I[ GAYCGMGEN~lI 200] 423 0IMCVFQGYSSKI .200] 6217 RIPGLGKDFK II FLLPSAPALG 1 70 F217i SLNARDISVK 11 0200] 1-61 ISLQQELCPSFII 0.2001 253 VAGPLVLVLI 0200] Table IX-Vi HLA-Al-1Omers.

24P4C12 Each peptide is a portion of SEQ 10 NO: 3; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Start IfSubsequence ore 186 il NVTPPALPGI 0.200 618 f FSGRIPGLGK F 0.150 [173 IPSAPALGRCF ii 0T1] r11 sscPEoPwTv ]10150] 125 IIGKNE FSQ1I 0.1251 ~676 IIRNNGSLDRPYIJ 0.125]1 608 IGGVGVLSFFF II .125] 286 f KGASISQLGF IiF 0.125] 80 II2NKDKPYLLYF I0.12 1 VTFVLLLIC I 5 f 1961 ITNDTIQGI 7-98 DTQQGISG ]0125 293 ]1 LGFTTNLSAY iFO-125] 7271I1YGInCWEE] 0A25 382 0.125 i 467GAFASAFI 0T 487I LISARTLR I 01001 650 II\IASGFFSVF II -5:T0 [6 I VLYPRNSTGA] 571&] 7272 7 WWEY .0 7333 7 IIILL j.0 [i]621VLSFF FFSGR 0.100 IOFCLPGVPWNM TO100 1 6f DISVKIFEDF 21 0.100 1 [T If IIVAWLYGPRIJ 0.ThW [326 MLIFLRQRIR II Ti1 ["54W f CLWCLE<FIK]J 0_100] Table IX-V3HLA*AI-10mers.

24P4CI2 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

WlO 2004/050828 PCT/US20021038264 Start Subsequence Score r0 1 ITPPALPGI 0250 9 NITPPALPGI 0.200 3 RCFPWTNTP 0.050 8 TNITPPALPG 0.013 7 WTNITPPALP 0.005 FPWTNITPPA U01 2 GRCFPWTNIT 0,001 1 LGRCFPWTNI 0.000 SPWTNITPPAL 0_000 41 CFPWTNITPP 0.000 4 SSKGLIPRSV 0.002 9 1hIPRSVFNLQI 0.001 I QGYSSKGLIP 0.001 8 LIPRSVFNLQ 0001 Table IX--HLA-AI-l0mers- 24P4C12 Each peptide is a portion of SEQ 10 NO: 15; each start position is specified, the length of peptide is 10 amino acids, and the and position for each peptide is the start position plus nine.

tart[ Subsequence Score] 7 AVGQMMSTMF 10.100 6 1ILVAVGQMMS 0.050 7 LVAVGQMMST 0.0501 8f VAVSQMMSTM 0.010 VVILVAVGQMM 0.010 QSWYWILVAV 0.003 SffWWILVAVG 0.001 4 I LYW VAVGM 0 ONO 1 3 "YWILVAVGQ 1 NYYWLPIMRN 0.003 16 TGHVFQTSIL 0.003 1771 GHVFQTSILG 0.003 6 LPIMRNPITP 0.001 8 I[ IMRNPITPTG 0.001 7 71 PIMRNPITPT 0.000 E YYWLPIMRNP 0.000 Table X.V1 HLA-A201 9mers* 24P4012 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Start iiSubs quence ~ore! WO 2004/0O50828 PCTIUS2002/03826I Table X-V1-HLA-A0201.9mers- 24P4CI2 Each peptide is a portion of SEQ NO: each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the WO 2004/050828 PCTIUS2002/038264 226 AQSWYWILV [U81 452 WTLNWVLAL 1 426 FQGYSSKGL 9.963 554 FLNRNAYIM 9.370 642 MTSILGAYV 9.032 164 QELCPSFLL 8.914 G93 KILGKKNEA 8.846 251 RLVAGPLVL 8.759 501 SLAFGALIL 8.759 487 LISAFIRTL -729 Table X-Vi-HLA-A0201.9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the startiiuse Start Susqec Sor I 536 CIMCCFKCC 114802 I 1 FILLLRLVA ]14.767 357 YP[VTFVLL 114.510 Table X-V3-HLA-A0201-9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start osiion lus ei t.

Start Subsequence Score F6 WTNITPPAL 1.365 E97 ITFPALPGI 0.567 2 RCFPVVTNIT 0.074 8 NITPPALPG 0.010 -411 FPWTNITPP 0.009 1 §HCF WThI 0.002 7 ij TNITPPAL 0000 r 5 P VVTNITP 3 0 0000 Table X-V5HLA-A0201-9ners- 24P4C12 Each peptide is a portion of SEQ ID NO: 11; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start positon plus eight.

Start Subsequenc Score ij ILLLVLIFL 11699 1774 9 \/LIFLRRI 173 1Th! VLEAILLLV 6]1 LLLVLIFLR l.251 2 LEAILLLVL 0.666 7 LLVLIFLRQ 0.048 WO 2004/050828 PCT/US20021038264 Table X-V7.HLAA0201-9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Start] Subsequence IlEe 11 WYWILVAVG 10.000 3 1YWILVAVGQ 0.000 Each peptide is a portion af SEQ ID NO: 19 each start position is specified, the length of pepflde is 9 amino acids, and the end position for each peptide is the start positon plus eight.

Start Subsequence 1Score 2 WAMTALYPL 12 TQPATLGYV 1.5 7 ATLGYVLWA l3 16 TLG YLWAS 11.2 YPLPTQPAT [.828 9 PLPTQPATL [5.470

MTALYPLPT

[IiY __iTI~GwL 00571 VT Cj ALYPLPTQP 0.46 F31 AMTALYPLP 001 17 LGYWWASN 0.0041 ]I TALYPLPTQ 0.002 W I GYLSNl 0.0301 7 II LYPLPTQPA 0.01 F 107 LPTQPATLG 0.001 I YWAMTALYP D.CI 0 14I PATLGYVLW 0.00 [i I PTQPATLGY 0.000 L 148 18.3 41 71 FILLFIL(I 641 IMTSILGAYV 66 69 7 546 WCLEKFIKFL 82 171 597 LLLFFGKLLV 82 12.5 598 LLFFGKLL I IL~II46 I 11 1138.5 665] TLFLC L DL f 241 VLSLLFILLL 4 .71 M49 P08.5FFS 101 433 1 GLIRSVFNL M84.94 2444 503 ILTLVQIARV 71.8 48 232

ILVALGVALV

72 11LLFILGYIVV 33 ALLIKEASKAV 1 42 243 449 GLFWLNWVLy 51 244 LLFILLLRLV E7.2 42 11.7 243 GLLFILILLR L 9 364 LLLICIAY:WA: 7.

68 48 170.9 23 2 RLVAGPLVLV 37 4- 321 AILLLLIFIL 88 1 YLLYFNIFSC 579 MLLMRNIVRV 603 I KLLWGGVGV 309 WLAALIVLAV Table X-VHLAA0201-9mers- 24P4C12 WlO 2004/050828 PCTIUS2002/038264 able XI-V-HLA-A0201 ers- 1 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

start 1 usqec 56 IWLYGDPRQVL

I

239 111HLFI 540 3 QMMSTMFYPL 1" 162 86 I LLYFNIFSC I 33 365 LLICIAYWAM i259 LVLILGV LGV 8 33 s2 LQQELCPSFL c7 580 LLMRNIVRVV 825 947 CILSSMIISV 1.21 F F517 VILEYIDHKL 57 1.9 FLNRNAYIMIII 19 686 YMSKSLLKIL 69 44 56.15WG 133 SQTVGEVFYT .4 1.79 F43B SVFNLQYGV F 231 WIL ALGVAL 99 9.13 F235 ALGVALVLSL f 1 41 441 NLQIYGVLGLf"I 6 GMCVDTLFL .8 F325 1LMLIFL R I 73 UK] 1CCFCC 1.29 Table XIVI.HLA-A0201 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Start II Subsequence Scor WO 2004/050828 PCT/US2002/038264 FQTSILGAYV 17.4 WLPIMRNPIT 14.

1 1 4 713 ITPTGHVFQT 23471 7 PIMRNPITPT 0.1921 [1871 HVFQTSILGA 126 21 11 QISILGAYVI 0.059 S11 RNPITPTGHV 0.0591 16 11TGHVFQTSIL 10.057 4 YLPIMRNPI 0.025 PTGHVFQTSI 0.012 F8 IMRNPITPTG 0.007 r 141 TPTGHVFQTS 0.001 VT] LNYYWLPIMR 001 F-1271 PITPTGHVFQ O.OUJ ii NPITPTGHVF 0.000 6 [PIMRNPITP 110,000 Y NWLPIMRN 0.000 I ]GHVFQTSILG 10.000 3 YYWLPIMRNP 0.000 S19 I11 OTSILGAY 0.000 9 VMRNPITPTGH jffE Table XI-V7-HLA-A00 jonr- 24P4C12 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

Start Subsequence PcoR WILVAVGQMM 1.62 17 QSWYWILVAV -7 I LVAVGQMMST I.51 8 VAVGQMMSTM 0.270 6 ILVAVGQMMS 0.127 F:i AVGQMMSTMF 0.007I 411 YWILVAVGQM 0.001 371 WYWILVAVGQ 0.000 1 SWYWILVAVG 0.000 Table XIV-HLAAA0201.0mers- 24P4C122 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

[Start Subsequence jScore able XI-V6-HLA-A0201 1 24P4C12 WO 2004/050828 WO 204100828PCTIUS2002/038264 QPALG 0.00 11 LPTQPATLGY 0.00 IMTALYPLPTQ 0.00 1 7-611 TALYPLPTQP 0_00 1 774T-1 QPATLGYVLW 0.00 1 [ii!AYWAMTALYP 10.000 GYVLWVASNIS 000 Table XII-Vl-HLA-A3-9mers- 24P4C11 2 Eao) peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eiht 7Stant I Subsequence IscRe 272I GIYYCWEEY 6.0001 _ii~ MMSTMFYFL 540 L 470 ASFYWAFHK 450 [i 4-i49 GLFWVTLNWV 14.500J 786] LLYFNIFSC 1.05 445] GVLGLFWTL 3.6451 66 GMCVDTLFL 3.6001 [633) l-LYWLPI 3600] 1 7] CLWCLEK 13.600 24~11 VLSLLFILL 3600 42 I LLFILGYIV 30001 393 1!NISSPGCEK 3000 3251 LMLI1FLRQR 2.700~ 322L ILLLMLIFL 2.70 23 -ALVLSLLFI 2.700 641[ IMTSILGAY 2.7001 15981 LLFFGKLLV 20001 2601 VLILGVLGV 11.8001 I513 IIQIARVILEY 11100 609~] GVGVLSFFF 1T-Boo 5371 IMCCFKCCL 1 VVGIVAWLY 1[tEo 636 11YMSKSLLKI. 1.8001 21 RLVAGPLVL 1100I 593i] KTLLLFF 1.80 358~] PLVTFVLLL 1.620 5441 CLWCLEKFI 1.600] 689 [KSLLKILGK 1. 350] 525 1[KLRGVQNPV 1.350 170 1 FLLPSAPAL 1.350o 64~7 11CLEKFIKFL 11.350 1597] LLLFFGKLL _375 36 LLICIAYWA 1__ 6506 I ALTLVQI 11.350 14 AR1 CPGVPWNM I~ Table X1II.VIHLA.A3-9Mers- 2424CI 2 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, tihe length of peptide is 9 amino acids, and the end position for each peptide is the start position plus ei ht.

Start Subsequence IScorel 501[_ SLAFGALIL 1.2001 1562 CVDTLFLCF 1.2001 F[349] GQMMSTMFYJ 1.0801 [A4-43 QIYGVLGLF 1 012] 31 I AILLLMLIF 0.900] 55 I VLOKVTDLL 0.900 326T II RQRI 0.9001 E268 if LAYGI1i IC 0,9001 r 107 ][GLQCPTPQV 0.9001 r613 LSFSGR 0.900 3181[ VLEA LLLM 0.900 -232 ILVLGVAL090 518 [LEY'IoHKL1 0.9001 4 52 TLNWVL 0.8101 56 DLLLFFGKL 07291 645 ][ILGAYVIAS 07201 258 11VLVLILGVL 0.608 49 IWGIVAWVL 06081 41 FLLFILt3YI 06081 54~ VAWLYGDPR 000 665 11i TLFLCFLED 060 95iI ILSSNIISV 060 457 VLALGQCVL 001 282 ]~VLRDK;ASIl 0600O PJFLNRNAYIM 0.600 1 -9 VLFLLFILG 110600] 315 ][VLAVLEAIL 10.600 68WLPIMTSIL[60 434~ 1LIQRSVFNL 0540 [612 l[VLSFFFFSG 0.540 L611 [GVLSFFFFS, 0.486 647] GAY VIASGF 0.450 58C] LLMRNIVRV 0.450 LLLICIAYW 0.450 56 IAIYGKNC 0.450 237 GVALVLSLL 0.:405 -IL CVLFLLF I L 0405 WO 2004/050828 WO 204100828PCTIUS2002/038264 Table XI-Vi-HLA-A3.9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptidle is 9 amino acids, and the end position for each peptide is the 2041 GISGLIDSL 045 045 37[AVLEAILLL 0.5 240 LLLFL 045 668~ LCFLED LER 0.400 38 VILWASNISS 0FO.4000 4891 SAFIRTILRY ]0.400 Table XII-V3-HLA-A3-9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the Table Xll-V5-HLA-A3-9mers- Each peptide is a portion of SEQ ID) NO: 11; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the -start position plus eiht.

Start I Subseuencejcore 54 ILVLIF 180 4ij LLLV 0900 F 9ll LFLRQR 027 F 2]I LLVL 0.00 2i LEAILLLVL 004 FTable XII.V6-HLA-A3-Smers- 24P4C12 Each peptidle is a portion of SEQI ID NO: 13; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the Str SubseqenceA e 7P I LIPRSVFNL i54 F 6 Gj RSVFN 0135 K IKGRLSVF FI0.013~ Table Xll-V8-HLA-A3-9mers- 24P4CI2 Each peptide ii; a portion of SEQ I D NO: 17; each start position is specified, the length of peptidle is 9 amino acids, and the end position for each peptide is the start position plus eight.

Startj Subsequence Score 4 ]j LPIMRNPI 10.600 7 IIMPITPT 1.2 Table XII-V5-HLA.A3-9mners- 24P4C12 Each peptide is a portin of SEQ I0 NO: 11; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptidle is the start position plus eigh 6 I.LLLVLIFLR P7.00 Table XlII.74LA.A3.9mers- 24P4C12 Start ISubse uence S[corc Each peptidle is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptidle is the start Rositonlus eight.

WO 2004/050828 PCT/US2002/038264 Table XII.VI-HLAA-lmers- 24P4C12 Each peptide is a portion of SEQ ID NO 3; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start osidon us nine.

Start c r 7.00 322 ILLLMLIFLR 0 584 NIVRWVLDK 7.00 230 43 3 GLIQRSVFNL 3 r-0 262 ILGVLGVLAY 24.00 00 272i GIYYCWEEYR M 0.0 464 VLAGAFASFYf8.00 13.50 665 TLFLCFLED L 7 0 13.50 516F RVILEYIDHK Mo 86 LLYFNIFSC I 135 7

LLPSAPALGR

0.0 578 FMLLMRNIVR 20 76 GMGENKDKPY 9.000 5974j VTDLLLFFGK 9.000 350t QMMSTMFYPL 8.100 667 FLOFLEDLER 8.000) 56 WLYGDPRQVL 6.757 333 RIRIAIALLK 6.000 I609 ]GVGVLSFFFF 5.4001 [4T VLSLLFILLL 5.400 5611 1AYGKF 45001 239 ALVLSLLFIL 4.050 49 IWGIVAWLY 050 r378 YLATSGQPQY .000 441 NLQIYGVLGL 3.600 235 1[ALGVALVLSL 3.600 S598 LLFFGKLL 3.000 621 fRIPGLGKDF 3.000 337[ L AIALLKEASK .11 362 FVLLLICIAY 1.8001 650 VIASGFFSVF 1.800 606 WGGVGVLSF 1.800 507 I LILTLVQIAR 1 1800 1) FLRQRIRIAI 1.800 Li] VLEAILLLML 118001 L24] GLGKDFKSPH 11800 [3079 WLAALIVLAV 1.800 1 3 12 ALVLAVLEA .118001 WO 2004/050828 WO 204/00828PCT/US2002/038264 Table XIII*V1-HLA-A3-10mners- 24P4C 12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is amino acids, and tihe end position for each peptide is the start position plus nine.

Sat Subsequence l i 275 11YCWEEYRVLR 0.900 232 I ILVALc3VALV 0.90 325 IILMLIFLRQRI 0.90 463] CVLAGAFASF 10.900 525 IIKLRGVQNPVA 090 5061 ALVOlIAh.900 fTable XIII.V3-HLA-A3-1 Omers- I 24P4C12 Start FSubsequence IScore] Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine. I [T-9]LNITPPALPGI FO.13 5]1 FPWTNITPPA 110.015 _L I RCFPWTNITP 10.0031 ID:t.. ITPPALPGIT 002 7zi WTNITPPALP 0.2I F-1 LGRCFPWTfNi 0.0011 2?i GRCFPWTNIT 0. 011 8 1 TNITPPALPG 0.000 [I-I1IP-WTNIT-PPAL 0.000 [Table XIII-V5-HLA-A3-10mers- Eah 244C 12 Eahpepfide is a portion of SEQ ID NO: 11; each start position is specified, the length of peptide is amino acids, and the end position for each peplide is the srt position plus nine.

It~r Subse( gence Score] 8 I LLVLIFLRQR 2 1 VLEAILLLVL 180 VLIFLRQRIR 060 AILLLVLIFL 0.4051 F-7 LLLVLIFLRQ 0.2701 Fli1l1 AVLEAILLLV 01 F-971.~ LVLIFLRQRI 10.0901 [4 EAILLLVLIF o.075A III[ LEAILLLVLI I0.03 Each peptide is a portion of SEQ 10 NO: 13; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine. I [tart Subsequence ficorel 8 L1I VFNLQ .0j F4i IISKGLIPRS 0.0003 L7I SSKGLIPRSV 110.000l Table XIII*V7-HLAA3-l0mors- 24P4C12 _Subseenc Score Each peptide is a portion of SEQ ID NO: 15; each start position is specified, tne length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine. I 91 AVQMTF 020 611 ILVAVGQMMS 0.120 11 WILVAVGQMM 110451 4 1 YWILVAVGQM 10.0001 1 Ws'WILVAGQ joo FTable XIII-V3-HLA1-A-lM-1ers. I 24P4C12 [Start E Subse uence ffj jTable Xlll-V8-HLA-A3-1 Diers. I Table XllI-V8-HL-A-l-1mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptide is 10 amino acids, and thle end position for each peptide is the start Position pus nine.

Sta8rt ScorI E 1871 HVFQTSILGA 10.300 WlO 2004/050828 PCTIUS2002/038264 Table XlI-V8-HLA-A3-10mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

Start Subequence 51 WLPIMRNPIT] 0.1001 21 1[ QTSILGAYVI 0.09 1 LNYYWLPIMR 1 .oe 13 JF ITPTGHVFIT 0.045 11 NPITPTGHVF ]F3X 8 IMRNPITPTG 0.030 II PTGHVFQTSI 0.0091 F j FQTSILGAYV 0.006 7 PIMRNPITPT 0.003 14 TPTGHVFQTS 0.003 19 II VFQTSILGAY 0.003 4 J1 YWLPIMRNPI 0,001 6 ]1 LPIMRNPITP 0.001 16 TGHVFQTSIL 0.0011 2 NWLPIMRN 0.000 9 J1 MRNFITPTGH Ioj0 12 PITPTGHVFQ 0.000 1711 GHVFQTSILG 0.0C1 RNPITPTGHV 11D0001 3 YYWLPIMRNP 0000 Table XIII-V9.HLAA.1 Omers.

24P4C12 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

Start I Subsequence Score T] ALYPLPTQPA 2.2501 AMTALYPLPT 0.600 II LPTQPATLGY 0.080 [13 71 QPAThGYVL 0.054 1I ATLGYVLWAS JNZo] I TLGYVLWASN 0.020 9 YPLPTQPATL 0.013 i LGY7LWASNI 0.009 Table Xll-WV9HLA-A31Omers- 24P4C12 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

[Start ce 5 PATLGYVLWA 0.004 10 TPATLG 0.003 71 YWAMTALYPL F0003 [571 MTALYPLPTQ 0.002 14 I QPAILGYVLW 0.002 [12] PiQPGY1 0.001 6 1 TALYPLPTQP 0.000 WAMTALYPLP 0.000 L1 AYWAMTALYP 0.000 F 1-9 GYVLWASNIS 0.000 j LYPLPTQPAT 0.000 Table XIV-Vt*HLAAI1O1-9mers- 24P4C12 start Subs. ue Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus nine.

=FTVGEVFYTK 74000 I IVRWVLDK 14000 CVFQGYSSK 4.000 [550 11 YIMIAIYGK 11 te00 YYMSKSLLK 1.600 F&721 KCCLWCLEK I1.20 6-901 SLLKILGKK 0.600 517 VILEYILI-K 0.600 [73( AYCGMGENK 0.400 [393[ NISSPGCEK 0.400 [207 GLIDSLNAR 0.360 [523 LLLMLIFLR 0.360 338 IALLKEASK 0300 [579 MLLMRNIVR 0,240 2431 SLLFILLLR 0240 [622 11 GJGKDFK 0. 200 able XIV.VI*HLA-AI I F9mers-] 24P4C12 Start1Subsequence 689[ KSLLKILGK 0.0] 1i RVILEYIDH 0.180 -i09] GVGVLSFFF 0.180 FPLISAFIR 0.180 -44-61 GVLGLFWTL 0.180 [i6i] GVLAYGIYY 0.180] [273 IYCWEEYR 0.160 5 ILTLVQIAR I o]6 P6Iq LCFLEDLER .6 698 KNEAPPDNK 0.120 4701 ASFYWAFHK 0.120 F-931 KVTDLLLFF 0.120 70-1 APPDNKKRK O 10 0 TDLLLFFGK 0.090 [38] CVLFLLFIL 0.0901 240 LVLSLLFIL 10.090 [5 J VAWLYGDPR 1.80 [1721 LPSAPALGR F 3491 GMMSTMFY 10.072 F 3347 IRIAIALLK 0.060 [745I LWCLEKFIK 0.060 F5-77 GKNFCVSAK 0. 0§6 E317 AVLEAILLL 0.060 699 NEAPPDNK< 0.060 SI 51I GVPWNMTVI 0.060 371 GVALVLSL 0 1 S57 LVLVLILGV 0.06j F20 7 PMRGPIK .6 575 KNAFMLLMR 0.048 F 212 LNAROISVK [359 1LVTFVLLLI .4 16 PVKYPS 0.04 F3047 SVQETwAA I F-()74 F6191 SGRIPGLGK JIFO040 0 j GIVAWLY 0.040j F681 LDRPYMSK 0.040 r6671 CVDTLFLCF 10.040 j7 DEDDEAYGK 0.036 83 1 KPYLLYFNI W0.036 147 GYIWGIVA 0.036 r-25171 RLVAGPLVL 0.036 3 II GQPQYVLWNA 0.036 WO 2004/050828 WO 204/00828PCTIUS2002/038264 ffable XIV.V-HLAAI1OI-9mers3 24P4C12 satSubsequence ~je 49 [VGAWL 000 314! [9yILAA 10-0301 r-76 WLQV 10.0301 1589 VVLDKTDj 0.3I r4521 WTLNVVVLAL 031 [141 11 YTKNRNFCL 0.3 4981 HTGSLAFGA 0.3 LVVGGVGVL I0.030 [362 l[FVLLLUCIA 0.0301 16,171! GVLSFFFFS 0.027 F-137l GEVFYTKNR 0.0271 [-STi[ AIYGKNJFCV 0211 if 27721] GIYYCWEEY 0.04j 11 ThRQLYPR JFT4 I 1-4 111 GLMVCVFQGY 021 467 1 GAFASFYWA 00241, I449 IIGLFWVTLNV ]024 1660 IIGMCVDTLFL -0-4 1496 IIRYHTGSLAF 0.0241 1511 ]j LVQIARVIL 000 r-2-18 SVKEDFA 0.20 1 233 1 9yAGVALVJ 0.020 r-27 SFRGPIKNR 0.20 I2i] 7 GMGNK 10.0201 L-4i14L LVNSSCPGL 000 F 2-5 2]1 LVAGPLVLV 0.2 I 7) CVSAKNAFM 10.0201I I "34771 AVGQMMSTM 0.0201 F 53-4 ARCIMCCFK 00201 15271 RGVQNPVAR 10.0181 [5471 DVICCVLFL 008 [i6-93[ KILGKKNEA 181 F1I1[ GQCVLAGAF 10.18 1=1 KQDDEA ]1 F-107 DEAYGKPVIK 10,018] Ii442]1 LQIYGVLGL .18 15981 LLFKLLV 001 F4 1 LLFILGYIV 016] 24411 LLFILILLIRL 10.016] Ii[E[ -LIFLRQRIR o1.ois] Irable XIV-V5-HLA-AI I 01-9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 11; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Ss-tart][ Subsequence Scie F6 1 LLLVLIFLR 0.3601 j1[ LVLIFLRQR7] 0060 F4 1 AIILLLVILIF 7 0.0121~ 5 1 ILLLVLIFL 10.0121 F-171 VLEAILLLV kOOS I.8 F~1 9 ULQR .06 F-7 7 LLVLIFLRQ 0.01 I-27 1 LEAILLLVL 0.01 M3 Ii EAILLLVLI 0.0011 Table XIV-V6.HLA-AII101 -Diners- 24P4C 12 Start Subsequencej Score) Each peptide is a portion of SEQ 1D NO: 13; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the 1Pstart positionplsigt Trable XIV-V6-HLA-AI1OI-9mers- L 24P4C12 35ir ISubsequence Scr LIVFNL002 2T YSKGIP 10.008 1 1[I SSGLIPI1 0.002 7f L IIS FN 0.2 8 1l IPRSVFNLQ 10-0001 1) PRSVFLQI 0.0001 1SSKGLIPRS IF0001 Sable XIV.V7.HL-A.AIio19mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peplide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Fs jiaSubsequence Scr) =311 ILVAVGQMM oool F41 WILVAVGQM 0F.006) L~ILVAVGQMMS. O004 I1 1 AVGQMMST 001 171SWYWILVAV ooo 3 -YWILVAVGQ 000 Sable XIV-VB.HLA-AI 101 .9mers) 24P4C12 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

[t a rt I[ Subsequence coe F11 NYYWLPIMR 030 1 20 1 QTSILGAYV 000 IF 1 HFT I .70081 1 19 11 FQTSILGAY 006 18-7 VFQTSILGA 004 M I1 WLPIMRNPI FO. 004 ~Io NPITPTGHV 003 WO 2004/050828 WO 204/00828PCTIUS2002/038264 Table XIV.V8-HLA-AI 101 -9mers- 24P4C1 2 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

2 YYWLPIMRN 0.002 9 RNPITPTGH 0.001 -12-11 ITPTGHVFQ 16 J1 GHVFQTSIL ~0 -1311 TPTGHVQ 6T 117[ PITPTGHrVF 10.000 7[ IMRNPITPT 1.0 5 LPtMRNP IT 0.000 ]j TGHVFQTSI 001 7671 PIMRNPITP 0.I 0 F-1I7 PGHFQrS 1~ oo 8T1 MRNPITPTG I.00 1 YWLPIMRNP ocoo IT able XlV-V9-HLA-AI 101-9mers- 24P4C12 Each pepfide is a portion of SEQ tD NO: 19; each start position is specified, the lengt of peptide is 9 amino acids, and the end position for each paptide is the start position plus eight.

IStarti Subsequence IEe 000 ~GYVLWASNI 9i .27 WAMTALYPL L12 IfTQATGY W0g [7 1I LYPLPTQPA 10, 04 [IY -jI QPATLGYVL 10.004 MTALYPLPT 0.2 VF-117 PTQPATLGY 0.2 6 j ALYPLPTQP 00 1 ITLGYVLWAS 00 [IIAMTALYPLP 1.0 L .9 1 PLPTQPATL 0.00 =81 YPLPTQPAT 000 Table X-IV- A-AI101 -9mers1 24P4C1 2 j Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start ps itinpus eight.

Start 11Subsequence ~je 14 7TPATLGYVW0.000 17 1LGYVLWASN 10.000! 1 YWVAMTALYP ]EaoI Table XV-VI-AI 11 OI-lners- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 10 amino acids, and the, end position for each peptide is the start position plus nine.

Fstar IL Subsequence Icoe F-51][ RVILEYID-K 319.0001 14 VTDLLLFFGKH i3.000l F-]1"5[7 QTVGEVFYTK 3.000l F-544]1 CLWVCLEKFI1(j 2.400_ r62711[ RIPGLGKDFK r5579 AYIMIAIYGK EE.00 I 7271[ GAYCGMGENK 1.00 [6801 SLDRPYYMSK 080 [4-69 FA.SFYWAFHK 060 22IIGIYYCWEEYR 040 [-427[ GYSSKGLIQR 040 F35771 AIALLKEASK 040 [i5371 IVAWVLYGDPR 10 ISLNARDISVK 11040 t I LLMLFR F 4237 MCVFQGYSSK 030 F5077I LILTLVQIAR 0,24 F-57781 FMLLMRNIVR 0.4I [1120 11 CPEDFWTVGK 020 F53 VARCIMCCFK 0.200 1151I KPVKYDPSFR 0.8 1 2643 GVLGVLAYGI 0.18 1609]] GVGVLSFFFF I 0.180 Table XV-V1 -Al 101-1 Omers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

I64] PYYMSKSLLK 160 -671FLOFLEOLER 10.160 [1171[ LLPSAPALGR 0.60 f-6978[ KNEAPPDNKK 1110.1201 F-84 TEPLISAFIR 010 1 27 1 GVALVLSLLF 10.10 r174311 YCGMGE NKDK0.0 I 6891[ KSLLKILGKK 0.9 SfYVIASGFS IRVLRE)KGASI 0I90 61211 VLSFFFFSGR 0.080 F4877I1 LISAFIRL 0.080 1 275 11 YCWEEYRVLR 110.0801 WO 2004/050828 WO 204/00828PCTIUS2002/038264 IfTable XV-VI -Al 101I-i mers- 24F4C112 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

Sat Subsequence ]j5co~r 157T] TVITSLCQEL 0.0301 463. 1 CVLAGAFAGF 000 1588I WVLDI(VTDL0.3 700W] EAPPDNKKRK [0.030 31 ~IT VIA VLEAIL 0,030 456i VNVLALGQCVL 0.0301 I257 ][LVLVLILGVL 0.0301 34 DL II VLFLLI 1611 j[GVLSFFFFSG0.2 159 GDPQVLYPR 004 12201 KIFEFAQSWJ 0.0241 65.4[ GFFSVFGMCV 0.2 58 LEKFIKFLNR 10.24 [i-4677 GAFASFYWAF O0024 [i6774 LERNNGSLDR 0.94 [347][ AVGQMMSTMF 0 .020 [566] YGKNFCVSAK 0.020] 353 TMFPLVTF 0.0201 585] IVRVWVLDKV 0.020] 707 APPDNKKRKK 0.020] 304] SVQETWVLAAL 0.2M [380[ ATSGQPQYVL 0.02 76-27. CVDTLFLCFL 0.0201 LV4141( L 0.020 YDPSF RRGPiK 0.020 71-1761 CVSSCPEDPW 0.020] 718671I NVTPPALPGI 00 20 7642 II SGAYI 0.020 512 1 I J IEY 0.018 E4781 KPQDlPTFPL001 7477L GYIVVGIVAW 008 461]j GQCVLAGAFA 008 KFLRNEDAY 10.0181 163II QQELCPSFLL0.1 Table XV-VI-AIIOI-lormers.7 [24P4C1 2 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptidle is the start position plus nine.

[Start ISubseq uec re l0mers-24P4C12 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the .start position plus nine.

rs-ta rt Subsequence Soe i~ ~rNITPPALPGI 004 F 57] FPWTNITPPA 1W004] r13] RCFPWTNITP 110.0021 7u [WTNIPPALP IM01 r-107i ITPPLGT 001 r--47 CFPWTNITPP o]ooo FL1]1 LGRCFPWTNI 10.00 fTNITPPALPG 0.00 GrC2PTNIT 0.000 M PW TNIPAL i .000j F Table WV45*ILA-A11 101.

l0mers-24P4C12 Each peptide is a portion of SEQ ID NO: 11 each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptidle is the start position plus nine.

[star ubuence Scre LEAILLVLI 0.0011 ITable XV.V6-HLA-A1 101.- 11lrmers-24P4C12 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptidle is 10 amino acids, and the end position for each peptide is the start oosition Dlus nine- Table XV..V7-HLA-AIIOI1- I Omers-24P24C12 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peplide is the start position plus nine.

Start 11 Sboune1Score WlO 2004/050828 PCTIUS2002/038264 Table XV-V7-HLA-A1101- 10mers-24P4C12 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the L artpi nine.

[Start Subsequence 3 ITQATLGYVL 0.012 7 ALYPLPTQPA j.ooa W I LPTQPATLGY 0.0041 1 YPLPTQPATL 0.003 1-671 ATLGYVLWAS 0.0031 f14 1 PATLGYVLW 10.002 1191 GYVLWASNIS 0.002 1 AYWAMTALYP 0.002 F12 PTQPATLGYV 0.001 MTALYPLPTQ 0.0011 AMTALYPLPT 0.001 l LYPLPTQPAT 0.000 18 LGYVLWASNI 0.000 S PATLGYVLWA 0.0000 117 TLGYVLWASN f 911 WAMTALYPLP o.ooo 2 YWAMTALYPL 0000 TALYPLPTQP 0.000 Li=J II ATLG I.000 Table VIM-HLA-A24-9mers- 244C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acds, and the end position for each peptide is the start position plus eight.

[Start Subsequence Score '17 1 200 356H FYPLVTFVL 0 57 11 LYGDPRQVL F496 RYHTSLAF 00 00 6471 AYVIASGFF 8 F- 00 E7 LYFNIFSCI .0 I I2i QYVIWASNI j700 Table XVI--HLAA24-9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the slartoson Pi t.

[Start Subseuence Scre 5 0 3 A F G A L I L T L 0 784I1 PYLLYFNIF "06 ,7540[ jFCLWCL 0.0 20.00 0.0 06 Tars.2HLA-1: MWes 4412 WO 2004/050828 PCT/US2002/038264 Table XVIVi-HLA-A24-gmers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and tie enid position for each peptide is the start osition lus elht.

Start 154 [NMTATSL 8.400 3111AALIVLAYL 8.400 26t LILGVLGVL 8.400 440 FNLQIYGVL 8.400 234 VALGVALVL 8.400 883 RPYYMSKSL 8.000 333 RIRIAIALL .000 596 LLLFFGKL 7.920 76[ LYPRNSTGA FT50O 3281 IFLRQRIRI 7.500 7377 AVLEAILL 7.200 125 GPLVLVLIL 7.200 F38 CVLFLLFIL I7200 2401 LVLSLLFIL I.EoI 232 ILVALGVAL 7.200 [5W1 WLDKVTDL 7.200 170 FLLPSAPAL 17,200 357 YPLVTFVLL 7.20O [23j LGVALVLSL .0 [621 RIPGLGKDF E.01 58 6.336 30751 VQETWLAAL 6.000 157 KPVKYOPSF 6.000 547]1 CLEKFIKFL 6.000 F597 LLLFFGK LL 6.0001 IYGKNFCVS 6.000 134] DVICCVLFL 6.000 13087 TWLAALIVL 6.000 [184 WTNVTPPAL 6.000 316 LAVLEAILL 6.000 200 TIQQGISGL 6.000 r 635 NYYWLPIMT 6.000 1TI[ F1TKNRNFC 6.000 673 IRNGSL 6.000 F44271 LQIYGVLGL 6000 F414 LVNSSCPGL 6.000 444[ IYGVLGLFW 6.000 Table XVIVI-HLA-A24-Smers.

24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start psto lsegt Starts Score 452 WTLNWVLAL 7R F242 LSLLFLLL 16.000 60VLGGVGVL 16.0000 WLPIMTSIL 6.000 511 LVQIARVIL 6.000 163 QQELCPSFL 6.000 291 SQLGFTTNL 6.000 434 LIQRSVFNL ff000 432 KGLIQRSVF L6.000 22_5 FAQSWYWIL 6.000.

3221ILLLMLIFL 6.000 [77 Ej KVTDLLLFF 5.760 24 I LSLILFILL7 76 i2-5 31 LVLVL 5.760 1TI GVALVLSLL 5.600 [281 SWYWILVAL 5.600 [4i ]LLRL AGPL 5600 35 1I VICCVLFLL 5600 3] 11CTDVICCVL 5600 590 ]VLDKVTDLL 5.600 f217 ISVKIFEDF 5.0401 224 DFAQSWYWI .000 [614[ SFFFFSGI 5000 274 LYYCWEEYRV 5.000 F6-3]6 YYWLPIMTS 5.0001 1370 AYWAMTALY 5.000 573 SAKNAFMLL 4.800 351] MMSTMFYPL 4.800 315 VLAVLEAIL 4.8001 100 IISVAENGL 4.800 204 GISGLIDSL 4.800 687 MSKSLKIL 4.800 I"44~ LLFILLLR L 4.8001 499 TGSLAFGAL 4.800 Table XVIV3-HLA-A24.9mers- P!!2410402 6" WTNITPPAL 2 RCFPWTNIT 1I GRCFPWTNI .1 3 CFPWTNITP 0.075 7 TNITPPALPJ0.01 5 PWTNITPPA 0014 8 NITPPALPG 0.012 =4Jj FPWTNITPP 0.010 Table XVI.V5-HLA-A24-Smers- 24P4C12 Each peptide is a portion of SEQ ID NO: 11; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start p t.

[Start 5 ILLLVLIFL 8.400 W I LLLVLIF 3.600 I VIFLRQRI 2.160 TI3 EAILLLVLI 1.800 I27 LEAILLLVL 0.480 1 VLEAILLLV 0.210 7.-l LLVLIFLRQ 0.025 6:i LLLVLIFLR 8 1 LVLIFLRQR 0.015 Table XV-V6HLA-A24-9mers- 24P4Ci2 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start positio plus eight.

Start Subsee Score 57[_ KGLIPRSVF 6.000 I LIPRSVFNL 6.000 GYSSKGLIP 0.500 WO 2004/050828 WO 2O4IOO82~PCT/US20021038264 Table XVl-V6-HLAA24-9mers- 24P4C12 Each pepile is a portion of SEQ ID NO: 13; each start position is specifed, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Start Susuence Rii] 6~i G~PSVPN 0.1 37 SSKGLIPRS 012 8 IPRSVFNLQ 0.020 :4i SKGLIPRS [j '01 4 21 1-KLIR 0-.0101 9 RVNQ Table XVI-V7-HLA-A24-9mers- 24P4C1 2 FEach peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptda is the start oosition plus eiaht, Table XVI-V8.HILA-A24-9nlers- 24P4C12 Each peptide is a portion of SEQ ID NO: 17; each start position is specifie, the length of peptide is 9 amino acids, and the end position for each peptide is the start posiion plus eight.

Start [Subsequence 11coel 11i PITPTGHVF 0.O240 10L! NPITPTGHV I_0.1 501 57 1 LPIMRNPIT olo iiil FQTSILGAY 010 27 QTSILGAYV I0.1201 [1211 IMRITPT 0.1001 F-37 F -YWLIMRNP 0.0251 14~~i PTGHVFQTS 1[-.2 Fl 1-21 ITPTGHVFQ 0.015G 1711 HVFQTSILG 10.010 ~T3 RNPITPTG 0.003 6 1PIMRNPITP 0FO.002 Table XVJ-V9-HLA-A24-9mers- 24P4C1 2 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the Tab leXVII-VI-HLA-A2410mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Start Subsequence IScore] 356 [jPLVFVL 60.01 370 AYQSQETL Pcool6 87 LYFN(FSCIL 0 01 140~ FYTKNRNFCL G000] 2741 YYCWEEYRVL 200.1 11 12000 370 AYAIIALr 0 0 11KYDPSFRGPI 12001 169 2~~S P~LI 000 47S VFNQYSGL 200 18 11 GYVIWASNI [=7j LYTQPA 19000 2 IIWAMTALYPL 16.000 [13 if QPATLGYVL I4.800.

PLPTQPATL 10600 111 YPLPTQPAT 0.180 1513 ATLGYVLWA 0.150 j16 31TLGYVL WAS ~.4 17111 LGYVLWASN 60120 MTALYLPr 0.00 1113 PTQPATLGY 0.018 WO 2004/050828 WO 204/00828PCT/US2002/038264 Table XVII-VI -HLA-A24-1 lOeS- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

[start If Subse uence IScorel j[LYPRNSTGAY I7.500! [U53f KFLNRNAYIM 750 254 Mf AGLVLVLfL.20 F34 ifSVQETWLAAL 720 I~ F-237 LVA 7.2g0) 637 if YLPIMTSI 750 I~.[LQQELCPSFL 720 [~FF ALVLSLLFIL 7.200 F-3i8 VLEAILLLML 720 F314711 IVLAVLEAIL 17.2001 F-3771 CCVLFLLFIL 17.200 r 5461 WCLEKFIKFL 720 F-35701 QMMSTMFYPL 720 F-97 NIISVAENGL 7.200 I231 QGISGLIDSL J7.2001 F243 SLLFILLLRL 7I00 rI2297 WYWILVALGV 17.0001 F-11SCTD)VICCTV 6.2 r 44 11 NLQIYGVLGL 6.0009 r357] YPLVTFVLLL 6.000 -760471!LLWGGVG VL. 6.000 Icj TIVOIARVIL 16.000] 5967 DLLLFFGKLL I6.0001 F-6887 VWVLDKVDL F431GLIQRSVFN 6-000 [-6W579[FGMCVDTLFL I .O 6I CIVLI 6.0001 '1 413 HLVNSSCPGL :6.0001 F2907 sQGTN 6.00 F-3211 AILLLMLIFL :6.0001 [7567 LAVLEAILL 16.0001 5 ~i LYGDPRQVLY 16.00 fM I FS ISS I f 77 ]1 F13H 6.000QECPFL I ThQ iS 6.000 ITable XVII-Vi-IILA-A24-10ors1 I 24P24C12j Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide, is 110 amino acids, and the end position for each peptide is the start position plus nine.

F S ubsequece Wcre F KPYLLYFNIF 15.7601 10] LAALIVLAVL 15.6001 F23371[ LVALGVALVL 5.600 F 565f IYGKNFCVSA 5.000 279 jfEYRVLRDKGA 635 1f NYYWLPIT .0 273fIYCEYR .0 I444 IiIYGVLGLFWT 5.000 68 11I YMSKSLLKIL F-5671 WLYGDPRQVL 4.800 2351 IALGVALVLSL 14.800 -252 11 LVAGPLVLVL ]4.8001 4491 GLFWVTLNWVL 1 4. 8001 FRO 27 LAGAI TL 400 [EIE5- LGKDFKSPHL 14.800) [1I[HTGSLAFGAL 4.800) VSAKNAFMLL 4.8001 F5421 KCCLWCLEKF ]4.400) F442 ]1 LQIYGVLGLF 4.200 iL~ 1CIAYWAMTAL 4.000) 24 )VLSLLFILLL 4.00 WO 2004/050828 WO 2O4IOO82~PCT/US20021038264 Table XVII.V3-HLA-A2410mers- 24P4C1 2 Each peptideo is a porton of SEQ ID NO: 7; each start position is specified, the length of peptidle is amino acids, and the end position for each peptidle is the start position plus nine.

Start Subsequence FPVVINITPPA 0.4 4[ CFPWTNITPP 0O.075 3 I RCFPWTNITP 0.02 871Zl TNITPPALPG 0.015 71 WTNITPPALP 0.015 2r GRCFPWTrNIT 10.012 Table XVII.V6.H-LA.A24-110mers-] 24P4C12 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptidle is the start position plus nine.

Start Subsequence Score 67[ KGLIPRSVFN 10.300 5j[ SKGLIPRSV 0EE0 4[SSKGLIPRSV 010 i[YSSKGLIPRS 010 IQGYSSKGLIP 10.0101 101 PRSVFNLQY 0.01 Table XVII-V7-HLAA24-l0mers-] 24P4C12j Each peptide is a portion of SEQ I D NO: 15; each start position is specified, the length of pepide is amino acids, and the end position for each peptide is the start pstin plus nine.

785Til Subsequence 1IScorel L..971 AVGQMMSTF200 5j[ WILVAVGQMM 1,2601 7TyWILVAVGQM

JE

=8 IVAVGQMMSTM 10,7501 13I WYWILVAVGQ 070 6 1 ILVAVGQMMS015 Ki1J SYIVV 010 7-FL QIMT010 E211 77VGL FTable XVII-V8-HLA-A24-l0mers- 24P4C12 Each peptde is a portion of SEQ ID NO: 17; each start position is specified, the length of pepfide is 10 amino adids, and the end position for each pepfide is the start position plus nine.

[tlartj Subsequence Scor [=1E NYYWLPIMRN 5.00 F 167 TGHVFQTSII 74.000 r iNPITPTGHVF 3.000Q 14 YWLPIMRNPI 12.1601 1 19i VFQTSILGAY 1.050j 1_21 QTSILGYI 100 F-31 YYWLPIMRNP 070 F -10[ RNPITPTGHV 030 R NPIT oio 1I13 1 ITPTGHVFQT 11.1501' 4 ]J EA LLVL 3 [.600 9 LVLRQRI 2160 1 =i AL LLV0.5 3~l LEAILLLVLI 0.120 17 11 LLV IFR 0.02 6 ILLLVUIFLR 0.018 j VL RQRR :F.15 81 iL~FR 0,1 Table XVII-V6-HLA-A24-1 Omers- 24P4C12 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the -start position plus nine.

Start Subsequence Scre 7Z GLIPRSVFNL 7.20 9 1PPSVFNIQ IF 1000 2 GYSSKGLIPR 0.O500 Table XYII-V9-HLA-A24-1 Diners-I 24P4C12 Each peptide is a portion of SEQ ID NO: 19; each start position is specifi ed, the length of peptidle is 10 amino acids, and the end position for each peptide is the start position plus nine.

[aaff P1 Tiieuence hs-core 7317 G LWS I g 9.000 [T7 ffLPPQPII750 71-3]1 TQPATLGYVLM720 2 YPLPTQPATL [18 Jj YVLASNI 1.0001 [Z I PAATALYPJ 000 [T IALYLTQPLJ 0.1441 MALPP17] TLGYVLWASN 0.1 201 I14 QPATLGYVLW 10.1001 WlO 2004/050828 PCT/US20021038264 Table XVIIV9-HLA-A24-lOmers- 24P4C12 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

Start Subsequence Score PLPTQPATLG 0.002 Table XVlII*V-HLA.BT.S9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Start Subse uence Score 240 LVLSLLFIL 20] 6 LWGGVGVL 20.00 34 DVICCVLFL 587 WLDKVTDL 347 AVGQMMSTM 15.00 00 SAKNAFMLL 2.001 0.0 uII3 VAGPLVLVL 1.0 12.001 3791IAYWAMTAL 12 1 225 FAQSWYWIL 2 NAR701SVKI IARVIEYI 2.00 514I3RVEYlE 0 12.00 1 WNMTVITSL 1.0 00 3 If LAVLEAILL 1.0 100 234 VALGVALVL 0 396 SPGCEKVP 18000 83 1 KPYLLYFNI 8.000 406 TSCNPTAHL 16.000 Table XVIIIV1HLA-B7-9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Start Score 241 VLSLLFILL 14.000 236 LGVALVLSL 4.000 440 FNLQIYGVL 4.000 184 WTNVTPPAL 4.000 597 i LLLFFGKLL 4.000 583 1 RNIVRVVVL 4.000 275 71 YCWEEYRVL 4.000 170 FLLPGAPAL 4.000 596 DLLLFFGK L 4.000 282 it VIROKOASI (4.000 158 II VITSLQQEL i537f1 IMCCFKCCL 4.000 7660 GMCVDTLFL 4.

7457 VLALGQCVL k 499 TGSLAFGAL 4.000 7-6671 YPRNSTGAY 4000 1417i YTKNRNFCL 555 LNRNAYIMI 4.

4 FQGYSSKGL 4.000 244] LLFILLLRL 4.000 242I LSLLFILLL 487j LISAFIRTL 4.000 ENKDKPYLL 4,00 351 11MMSTMFYPL 14.00 442- 1 LQIYGVLGL 4.000 200i TIQQGISGL 14.000 434] LIQRSVFNL 14.000 750-1 SLAFGALIL 14.000 722 ILLLMLIFL 4.000 7251 RLVAGPLVL 4.000 204 GISGLIDSL 4.000 75727 VSAKNAFML 4.000 F687 MSKSLLKIL 4.000 100 IISVAENGL f.0 [T2 ILVALGVAL 4.000 302[ YQSVQETWL 4.000 1 VICCVLFLL 4.000 WO 2004/050828 [Table XVIII.V1 *HLA-B7-9mers- L 24P4C12 [Each peptide is a portion of SEQ ID NO. 3; each start position is Ispecified, the length of peptide is 9 amino acids, and the end positionl for each peptide is the start position plus eight.

StartfSbeeneior GIKNSCT.00 482 ffIPTFPLI,9A r300 343 11 EASKAVGQM 149] I LPGVPWNMT =300 -581]I LMRNIVRVV 115211 VPWNMTVIT 2.000 531_ 2PVA0C00 188] TPLPI 200 112 f TPQVCVSSC 200 I DPRQVLYPR i200 52 If KLRGVQNPV- 2.Z000I 314]I IVLAVLEAI ii2,000! 1671 CPSFLLPSA 2.000 151 ]1 GVPWNMTVI 2.0001 192I LPGITNDTT 2.0001 359 f LVTFVLLLI 2.0001 252J1 LVGPVLV 150 491 1 FIRTLRYHT =1.50 530]1 QNPVARCIM 150 2391 ALLSLF 120 305 j Table XVIII-V3-HLA*B7.9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Sat Subsequence Score NITPPALPG .1 C-FPWVTNITP-- 0.00 SPWTNITPPA_0.00 PCTIUS2002/038264 Fable XVIII-V6-HLA-BT-9mers.

24P4C12 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the startposition plus eight, rGYSSKGLIP 0001 1 1-VLIFLRQRI J10.6001 4- 2 fLAILLLVL 0.40601 ,17 VLEIt ILLL V 0.0601 7:T7F LLVLIFR? 0.010] trable XVIII-V6-HLA-B.9mers- 2474IC12I Each peplide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 9 amino acids, and t end position for each peptide is the L start position plus eight.

ra I Subsequence Sgire 771 ILIPRSVFNL 400 781[ IPRSVFNLQ 200 757I KIRSF 04] 757iGLPSFN0O0 7 1411 SGLPRV .00 WO 2004/050828 PCT/US2002/038264 ~jHVFQTSILG 0.050 19 FQTSILGAY 10.020 18 VFQTSILGA 0.010 9 7jRPITPTH 0.01 F 61__jRNPjTPJ10.003 7271 YYWLPIMRN 1 111 PITPTGHVF IM0 F147 PTFQTS EK 78i MRNPITPTG =ai N'YWVLPIMR JE.01 -A-87-10mers- C12 11 WO 2004/050828 WO 204100828PCTIUS2002/038264 [Table XIX.VI -HLA-07-1 Omers-1 24P4C1 2 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptde is amino acids, and the end position for each peptide is the start position plus nine.

Str 1Subsequence S~cor SLLFILLLRL 400 L LLL4000: [-449]1 GLFWTLNWVL 4.000 [i162-1 LQQELCPSFL 400 F271QSWYNILVAL 4.00 F-49-1 TLFGAL7 4.-000 F-4871 YIWVGIVAWL 4000, I EU If LLWVGGVGVL I F-1-97 LPGVPWNMTV 4.000 I VLILGVLGVL 4L.000 4931I RTHTGSL HK 24 ]1 LLLVAGPL 4. 00 0 1231 ~fj WLALGVAL 4. 000 1 5I GSLAFGALIL I~ 54 67 J[ WLEKFIKFL 4.0 12411 VLSLLFILLL 4.00 I539 1I CCFKCCLWCL =4.000 I445 1[ YGVLGLFWTL 14.000 F307 7[ ETWLAALIVL 14.000 I 435 IQSFLI 4000 7721 VSAKNAFMLL 400 I3 FI1 GLIQRSVFNL 4.00 5~.1 VILE-YIDHKL 1S9 I TTIQQGISGL [F i SCTDVICCVL 4.000 [17- F34311 IFKVGM 3000 34 AfKVQMT 3.000 F581]L LMRNIVRVVV 3.000 573]r SAKINAFMLLM Ir~ ASGFFSVFGM j0OQ IVPINTSCNPT 2.000 F182 i1 FPWVTNVTPPA O 07 r5-28 IGVQNPVI 2.00 [2817] PRVLRDGSI 200 16 Table XIX-Vl-lILA-B7-10mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptde is 10 amino acids, and the end position for each peptide is the start position plus nine.

Stairt r1 Subsequence 1Iscore, Table XIX-3-HL11A-B7-lOmers- 24P4C1 2 Each peptide is a portion of SEQ ID NO: 7, each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Start jSubsequence i -or 171 LGRCFPV/TNI 116.500 Table XIX-V6-HLA-B7-10mrs- 24P4C12 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

St~r Subsequence r-corc IPEFNLQI_8000 71 GIPRSFNL 14.000 1~J SSKGLIPRSV 0.0 :]6J1 KGLIPRSVFN 0.0O20 TaI XI 5 H A -137- lom e rs Table XIX-V7-HLA-B7.I0mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the .start position plus nine. WO 2004/050828 WO 204/00828PCTIUS2002/038264 W Star Subsequence Score! .1VAVGQMMSTM f 3. 000 L7.. IfAVGQMMST E I =9 1 IQSWLVA 10.3001 4 YWILVAVGQM 0.1o00 6 I ILVAVGQMMSII 2J F-27 YWILAVGJ 0.001 L~(WYWILVAVGQ 0.001 Table =4XIX A-~1711mers- Each peptide is a portion of SEQ ID NO: 17;- each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus nine, SatSubsequence Ioe [6iT][ ~QTSIL 14.0001 HVF1QISILGA 10.4001 111 NPITPTGHV 10.3001 FQTSILGAYV 0.2001 17is] ITPTGHVFQT 10.100 -1 IMRNPITPTG 10.100 WLPIMRNPIT 10.100, F-7 .YWLPIMRNPI 10.060 L7 PIMRNPITPT 0. 045 PTGHVFQTSI 10.040 1= 1~ LNYYWLPIMR 10o.0*1 2 NYYWVLPIMRN 10.0031 19I ~T IEG I~ PTGHF 0.0 01 LET RNPTPTGH 0.00 7_j[ ALYPLPTQPA 040 [11Z 11 ATLGY 0.400 QPAL GYLW 10.400 FI 27 YTALYPL 1 0.400 Ha 181 LjYVLWSN 0.400 F 4 1 J[ YPLPT0300 F 371 WATALYPL rf,00 1iF67 ATLGYVLWAS 0.060 -7 1 TALYPLPTQP 0.03 15 I PATLGYVLA0.3 [12 j[ TLGYV 0.020 17 T GYVLWASN]0.2 5~I MTALYPLPTQ0.1 8al LYPLPTQPAT0.0 [T~AYWAMTALYP .00 ialGYVLWASNIS0.2 I E] PLPTQPATLG ]0.0027 WO 2004/050828 PCTIUS2002/038264 I Table XX-VI-HLA-B35-9meres- I 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position pluseight.

Start Subsequence IS 311 1 AALIVLAVL JIa00 1275 YCWEEYRVL aooo 253 VAGPLVLVL 3.000 651 IASGFFSVF 3.000 647 GAYVASGF 3.000 225 FAQSWYWIL 3.000 R74E GRC3o 2347 VALGALVL 3.000 3679 lAYWAMTAL 3.000 F141[ YTKNRNFCL 3.00 1 4947 TLRYHTGSL 3.000 F678[ NGSLDRPYY 3.000 1249 LLRLVAGPL 3000 11171 VSSCPEDPW 2.500 I 282] VLRDKGASI 2.400 F-287 KNRSCTDVI 2.400 317 AVLEAILLL 2.000 266 LGVLAYGIY 2.000 363 VLLLICIAY 2.000 267 GVLAYGIYY 2.000 GPIKNRSCT 2.000 415 VNSSCPGLM 2.000 VVGIVAWLY 2.000 589 WLDKVTDL 2.000 272 GIYYCWEEY 2.000 188 TPPALPGIT 2.000 432 KGLIRSV 1L0 152 VPWNMTVIT 2.000 192 LPGITNDTT 2000 531 NPVARCIMC 12.000 583 RNIVRVVVL 12.000 366 LICIAYWAM 12.000 546 WCLEKFIK F 2.000 I554 FLNRNAYIM I12.000 513 QIARVILEY 2.000 h927 FSCILSSN I 2.000 I QNPVARCIM 2.000 Table 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Start ubsquence cr 133 STVGEVFYT120 LVAGPL 2.000 409 NPTAHLVNS 2.000 [347 AVGQMMSTM 2.000 634 LNYYWLPI 2.000 62171 RIPGLGKDF 2.000 [643 TSILGAYVI2.000 [110 IPTFPLISA 2.000 [841 ]J IMTSILGAY 12,000 [677 NNGSLDRPY 2.000 F-421 GLMCVFQGY 2.000 [ii6 LQQELCPSF LPGVPWNMT 2.000 [263 LGVLGVLAY 2.000 [167 IICPSFLLPSA 145 II CLPGVPWNM 2.000~ [571 CVSAKNAFM 2 F TFQVCVSSC 2.000 384 QPQYVLWAS j.0o [500 1 GSLAFGALI 2.000 9 11 GQMMSTMFY 2.000 F 65371 SGFFSVFGM 2.00 0 FT47 KQRDEDDEA 1.800 1 560 7GMCVDTLFL r [Thy I RSCTDVICC Irl.500 L ii SSKGLIQRS 1.600 Table XX-V3-HLA-B35-Omers- 24P4C12 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

[Start Subsequence_ Score [1 WTNITPPAL 1.000 24P4C12 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 9 amino addsiand the end position for each peptide is the start position plus eight.

Start Ii Subsequence IIScore WO 2004/050828 PCT/US2002/038264 7= 1LIPRSVFNL 1.000 8 IPRSVFNLQ 0.6001 6 GLIPRSVFN 0.100 4 SKGLIPRSV 0.020 F- 9 1 PRSVFNLQI 0.004 I..li GYSSKGLIP o.ooi Table XX-V7-HLA-B35-9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

[itar-iJ Subsequence PiR F 8 11 AVGQMMSTM 2.000 11 ILVAVGQMM 2.000 4 71WILVAVGQM 2.000 J VAVGQMMST0.300 SLVAVGQMMS U.

SWYWILVAV 0.020 YWILVAVGQ o.oo0j 2 I WYWILVAVG 0.001 Table XX-V-HLA-235-9mrs- 24P4C12 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start osition us eight.

Start Subs.quence Score f- 1-071 NPITPTGHV R 4,000 F-131 TPTGHVFQT 2.000 -I 9 71 FQTSILGAY 2.000 Fi-571 LPIMRNPIT 12.0001 TGHVFQTSI 0.400 I T7 WLPIMRNPI 0.400 7 1IMRNPITPT 0.3001 QTSILGAYV 0.200 1I1( PITPTGHVF 0.100 -1671 GHVFQTSIL 0.100 1=1 RNPITPTGH 0.020 12 [i YWLPIMRN l10nici Table XX-V8-HLA-535-9mers.

24P4C12 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Startore F18 7 1 VFTSILGA ]10,010 [14[ I1 TGVFQjS 0.010 F17 HVFQTSiLG 10.010 F 127 ITPTGHVFQ 0.010 S PI MRNPITP 0,001 1 37 YWLPIMRNP 0.001 8 MRNPITPTG 0.001 1 NYYWLPIMR 0.001 Table XXV9MHLA-B35-9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight Start Subsequence score 13 QPATLGYVL 2 WAMTALYPL 3.000 i YPLPTQPAT 2.000 Ii itI PTQPALGY )0.2001 I T TQPATLGYV 110.200 10 7 LPTQPATLG 0.200 I PATLGYVLW 110.1501 15 1 ATLGYVLWA 10.100 L±.4i1 MTALYPLPT o.iool F-16 TLGYLWAS 01001 11711 LGYVLWASN l0.1001 I 9 J PLPTQPATL 0.100 F GYVLWASNI 0.040 5 TALYPLPTQ 0.030 1 711 LYPLPTQPA 0.010 3 AMTALYPLP I.010 j96j ALYPLPTQP 0.010 YWMAMTALYP 10.001] Table XXl.V1.HLA-35-10mers- 24P4C12 Each peplide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Start Subsequence Kcr 478 KPQIPTFPL 0T'§b 00 68 0.00J 23 KPYLLYFNIF fN 0 683 RP MSKSLL1J] 00 24 IKQRDEDDEAYIF 00 123 DPWTVG7KNEF 488 f]IPTFPLISAF 625 II~KSPHL .00 357 YPLVTFVLLLOn0 .00 213 NARDISVKIF 0.0 573 SAKNAFMLLM 00 346 KAVGQMM1 12.000 00 79 ENKDKPYLLY F F]0 662 ASGFFSVFGM 1 488 ISAFIRTLRY 1 6.000 0 1 175 APALGRCFPW 100 0.0 630 KSPHLNYYWL100 6192 LPGITNDTTI0 551 Fl LNRNAY 6251 331 RQRIRIAIAL 6.00 343 EASKAVGQMM 00 7U07 DPRQVLYPRN 601 66 7 _000C 6 369 IAYWAMTALY 6 .0001 WO 2004/050828 WO 204/00828PCTIUS2002/038264 rTable XXI-V1 -HLAB35-l Omers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is amino acids, and the end position [or each peptde is the start position plus nine [Startl[ SujbsequenceScw F3961 SPGCEKVPIN 2.000 LGVLAYGIYY 12.0001 1 40211 VPINTSCNPT r2.0001 [378 IYLATSGQPQY I200 1 36571~ LLICIAYWAM 200 2 9_3 GFTTNLSAY I2.000I 262 [ILGVLGVLAY. 112.0001 F2861 KGASISQLGF 2.0001 5297[ VQNPVARCIM 2I00 491 WGVLY 2. =Ol 147 JIFCLPGVPWNM 11.00, 265~ VLGVLAYGIY 304__ SVQETWLAAL 44 VLAGAFASFY 200 F207[ DPSFRGPIKN 200 r6 61 ][MCVDTLF-LCF 2.0001 192 FSL SSNII 00 [512 VQIRVILEY99 j182] _I P J.I6 j639][LPIMTILGA FCVSAKNA 200 4931 RTLRYHTGSL 112.00 -633 HLNYYWLPIM F53L1 NPVARCIMCC 200 622[ IPGLGKIDFKS ]2.0001 [4~1 FPLISAFIRT 200 542 I( CCL-WCLERF 200 589 J WLDKVTDLL 200 F-517]1 VILEYIDHKL 200 F41471 LVNSSCPGLM 2.00 L-3474 ASKAVGQMMS [1.50 F LAGAFASFYW -46 I SAYQSVQETW [.0 -6691 FGMCVDTLFL 11500 LF315]I VLAVLEAILL o LE II S-SCPEDPWTV l[-500 WO 2004/050828 WO 204/00828PCTIUS2002/038264 [Tbe XXI-V5-HLA-B35-1 Omers- 24P4C1l2 Each peptide is a portion of SEQ ID NO: 11; each start position is specified, the length of peptide is amino acids, and the end position for each peptid e is the start position plus nine, [start][ Subso uence Scor 1~ 10 VLFRRI l LjLLVLIFLRQ7R7010 Table XXI-V6.HLA-B35-l Omers.1 I24134C12 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

rStart 1 Subsequence Score F9i I IPRSVFNL 1.000 II- [j SSKGLIPRS 1050 1 GLIPRSVFN 0.200 i 5 'i SKGLIPRSVF7M 1 I 1071f PRSVFNLQ1Y 0.2 F-87 1 LIPRSVFNL 0. 010 r1 1 QGYSSKGLIP If0.010 [Table XX4V7-1-11A1335-10mers- 24P4C12 Each peptide is a portion ot SEQ ID NO: 16; each start position is specified. the length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

F1IIt Sbequen e cr 8- ]I AVQMSM 16.000 f-51 IIwiLVAVGQMM 2.000 F-9 1 I GMSTMF 110- ,=LI QSWYWILVAV][1.00 Table XXI-V7-HLA-B35-l Omers.

24P4C1 2j Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Start I Subsequence Scoe 6 ILVVGQM 0.100 71 LVAVGQMMST o0.io 2 IISWYWILVAVG 0.001] 7311 'WWILVAVGQEi Table XXI-V8-HLA-B35-i Omers- 24P54C12 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peplide is 10 amino acids, and the end position for each peptide is the start position plus nine.

~Subse uence Soa [TFII QTSILAYV 20.4001 ~~1I NPITPTGH V [IF 7611 LIPTP 0.200 F211 TFQTSILY 0.00 F] 11 QTSILGAYI F F1311 RIT GHV7 [F 6 WjLPIMRN PITP oio r 197 1 TVFQTSI 0.40 1 :i IMRNPIT 0.30 F-4 NYWLPI-MRNl .1 !T 87] 1IMRNPITPT ooo LiF LNYYWLPIMR 0.1 3 YYWLPIMRNP 0.00 12T 1 PITPTGH Q 0.0 I9 1 MRNPITPTGH 09.001 [Table XXI-V9-HLA-B35-l0mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptidle is 10 amino acids, and the end position for each peptide is the start position plus nine FStart ISubsequence Score 271 YWAMTALYPL [1.92 HT 13 TQPATLGYVL I[i.q F-7 YPLPTQPATL 18I FT 147 QPATLGWVLW 050 [F1/ LPTQPATLGY [.200 16 J1 ATLGYVLWAS o1i7 F 4II AMTALYPLPT Io~i6E 811 LYPLPTQPAT 010I [17 77 TLGYVLWASN ]ME100 F7]ALYPLPTQPA 0.100 F--3 1 WVAMTALYPLP 10.050 I F12 PTQPATLGYV 0.20 TALYPLPTQP 1 l] 57FMTALYPLPTO 10.010 F15 ]I PATLGYVLWA 0.010 I 1 1 AYWAM1TALYP 110.0101 WO 2004/050828 WO 204100828PCTIUS2002/038264 Tables XXII.XLIX: 7ableXXlI-Vl-HLA-Al-9Ifles- 24P4C12 Each peptide is a portion of SEQ ID NO: 3: each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pas 123456789 score NKDKPYLLY 34 58 YGDPRQVLY 33 222 FEDFAQSVW 26 QRDEDDEAY 77 MfGENKDKPY 263 LGVLGVLAY 24 489 SAFIRTLRY 23 513 QIARVILEY 23 628 DFKSPHLNY 22 LFLLFILGY 21 267 GVLAYGIYY 21 363 VILLLICIAY 21 421 GLMCVFQGY 21 WVGIVAWLY 318 VLEAILLLM 629 FKS PHLNYY 133 S 1VGEVFY 19 437 RSVFNLIY 19 662 CVDTLFLCF 19 11 EkYGKPVKY 18 370 AYWVAMTALY 18 1 B KYDPSFRGP 17 32 CTDVICCVL 17 66 YPRNSTGAY 17 277 WEEYRVJLRD 17 379 LATSGQPQY 17 594 VTDLLLFFG 17 165 ELCPSFLLP 16 353 STMFYPLVT 16 398 GCEKVPINT 16 552 IKFLNRNAY 16 590 V LIDKVIDILL 16 678 NGSLDRPYY 16 TableXXII-V3-H LA-Al -9mers- 24P4C12 Each peptide is a portion of SEQ I D NO: 7; each start position is specified, the length of peptide is 9 amino acids, and the end position for each pelpfide is the start position plus eight.

Pos '123456789 score WO 2004/050828 WO 2004050828PCTIUS2002/038264t TableXXll.V3.HLA-A1 .9mers- 24P4C1 2 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 8 NITPPALPG 11 9 ITPPALPGI 6 WTTNITPPAL 6 3 cFPWrN!TP -9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 11; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 1 VLEAILLLV 20 7 LLVLIFLRQ 10 TableXXll-V6-HLA.A1-9mers- 24P4C1l2 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 2 YSSKGLIPR 12 1 GXYSSKGLIP 7 3 SSKGLIPRS 7 8 IPSVFNLQ 7 9 P3RSVFNLQI 7 6 GLIPRSVFN 5 TableXXIl-VI-HlLA-Al -9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score IL VA VGMM 5 3 'rWILVAVGQ 4 7 VAVGQMMST 4 6 LVA\/GQMMS 3 1 SWYWILVAV 2 2 WWLVAVO 2 TableX(XIl-V8-HILA-AI.9mers- 24P4C 12 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptidle is the start position plus eight.

Pos 123456789 scor 0 19 FQTSILGAY 16 14 PTGHVFQTS 11 12 ITPTGH'LFQ 8 18 VqFTSILGA 7 20 QTSILGAYV 7 TableXXI*VSHLA-A1*9mers- 24P4C1 2 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 11 PITQPATLGY 31 15 ATLGYVIDWA 16 TableXXIII-VI -H-LAA0201gn'ers-24P4C1 2 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide, is the stard position plus eight.

Pos 123456789 score TableXXIl-VI-HLA-A0201- 9nmers-24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino adids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 260 VLILGYLGV 31 244 LLFILJLRL 29 580 LLMRN!1VRV 29 95 ILSSNIISV 28 204 GISGLIDSL 28 261 LILGVLGVL 28 322 ILLLMLIFL 28 506 ALILCVQl 28 170 FLLPSAPAL 27 252 LVAGPLVLV 27 449 GLFWVTLNWVV 27 487 LISAF!RTL 27 604 LLV'VGGVGV 27 45 ILGYIWGI 26 232 ILVALGVAL 26 233 LVALGVALV 26 315 VLAVLEAIL 26 501 SLAFGALIL 26 521 YIDHKLRGV 26 42 IiFILGYIV 1107 GLQCPTPQV 200 TIQQGISGL 211 SLNARDISV 239 ALVLSLLFI 257 LVLVL.ILGV 258 VLVLIlJGVL 282 VLRDKGASI 317 AVLEAILLL 457 VLALGgCVL 598 LLFFGKLLV 650 VIASGFFSV 686 YMSKSLLKI 41 FLLFILGYI 24 WO 2004/050828 TableXXllI-VI -HLA-AO201 9mers-241P4C112 Each peptide is a portion of SEQ ID NC: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 49 IVVGIVAWL 24 310 LAALIVLAV 24 311 AALIVLAVIL 24 333 R11RIAIALL 24 434 LIQRSVFNL 24 509 LTILVQIARV 24 525 KLRGVQNPV 24 564 AIYGKNFCV 24 581 LMRNIVRVV 24 596 DLLLFFGKL 24 605 LVVGGMGVIL 24 VICCV7LFLL 23 56 WLYGDPRQV 23 240 LVLSLLFIL 23 251 RLVAGPLVL 23 253 VAGPLVLVL 23 309 WVLAAL!'VLA 23 340 LLKEASKAV 23 358 PLVTFVLLL 23 494 TLRYHTGSL 23 518 ILEYIDHKL 23 547 CLEKEIKEL 23 589 WVLDKVTDL 23 590 VLDKViTDLL 23 597 LLLFF1*K[L 23 100 IISVAENGIL 22 241 LSILLIFILL 22 248 LLLRLVAGP 22 249 LLRLVAGPL 22 265 VLGVLAYGI 22 446 GVLGLEWTL 22 452 WTLNWVLAL 22 578 FMLLMRNIV 22 638 WLPIMTSIL 22 660 GMCVDTLFL 22 158 VITSLQQEL 21 187 VTPPALPGI 21 191 ALPGIINDT 21 237 GVALV7LSLL 21 247 ILLLRLVAG 21 313 LIVLAVLEA 21 314 IVLAVLEAI 21 442 LQIYGVLGL 21 507 LILTLQIA 21 537 IMCCFKCCL 21 599 ILFFGKLLVV 21 693 KILGK1SNEA 21 TableXXIII-VI -HLA-AC201- 9mers.24P4C1 2 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight, Pos 123456789 score 34 DVICCVLFL 20 38 CVLFLLFIL 20 44 FILGY!WVG 20 207 GLIDSLNAR 20 228 SWYWILVAL 20 234 VALGVALVL 20 236 LGVALVLSL 20 242 LSLLFILLL 20 319 LEAILLLML 20 326 MLIFLRQRI 20 339 ALLKEASKA 20 364 LLLIC!AYW 20 417 SSCPGLMCV 20 503 AFGALILTL 20 633 HLNYYWLPI 20 644 SILGAY VIA 20 673 IDLERNNGSL 20 690 SLILKIJLGKIK 20 48 YlVVGIVAW 19 245 LFILLLRLV 19 255 GPLVLVLIL 19 262 ILGVLGVLA 19 268 VLAYGIYYC 19 291 SQLGFTTNL 19 318 VLEAILLLM 19 323 LLLML!FLR 19 329 FLRQRIRIA 19 351 MMSTMFYPL 19 365 LLICIAYWVA 19 414 LVNSSCPGL 19 464 VILAGAFASF 19 544 CLWCLEKFI 19 617 FFSGR!PGL 19 666 LFLCFLEDL 19 86 LLYFNIFSC 18 231 WILVALEGVA 18 235 ALGVALVLS 18 243 SILLFILLLR 18 336 IAIALLKEA 18 355 MFYPLVTFV 18 369 IAY WAMIAL 18 380 ATSGQEQYV 18B 394 ISSPGCEKV 18 439 VIFNLQCIYGV 18 459 ALGQCVLAG 18 510 ILVOIARVI 18 511 LVQIARVIL 18 PCT/US2002/038264 TableXXI.VI-HLA.A0201 Smers-24P4C12 Eadh peptide is a portion of SEQ I D NO: 3; each start position is specified, the length of peptidle is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 514 [ARVILEYI 18 517 VILEY!DHK 18 583 RNIVRVWVL 18 602 GKLLWVGGV 18 645 ILGAYVIAS 18 46 LGYIVVGIV 17 128 GKNEFSQTV 17 154 WNMTV!TSL 17 177 ALGRCFPWT 17 184 WTNVTPPAL 17 213 NARDISVIKI 17 246 FILLLRLVA 17 289 SISQLGFTT 17 300 SAYQSVQET 17 305 VQETWLAAL 17 312 AILIVILAVILE 17 325 LMLIFLRQR 17 335 RIAIALLKE 17 354 TMFYPLVTF 17 359 LVTFVLLLI 17 453 TLNWVLALG 17 458 WVLALGQCV 17 502 LAFGAILILT 17 504 FGALILTLV 17 513 QIARVILEY 17 554 FLNRN8YIM 17 560 YIMIAIYGK 17 586 VRVWVLDKV 17 642 MTSILGAYV 17 658 VFGMCVDTL 17 31 GCTDVICCV 16 43 LEILGYIWV 16 64 VLYPRN4STG 16 90 NIFSCILSS 16 19 SCPEIDIEW 16 144 NRNFCLPGV 16 148 CLPGVPWVNM 16 161 SLQQELCPS 16 230 YWILVALGV 16 254 AGIPLVILVILI 16 308 TWLMLIVL 16 316 LAVILEAILL 16 320 EAILLLMLI 16 357 YPLVTEVLL 16 362 FV1.LL!CIA 16 373 AMIALYLAT 16 376 ALYLATSGQ 16 WO 2004/050828 TableXXIIl-VI-HLA-AC201 9mers-24P4C1 2 Each peptide is a portion of SEQ ID NO: 3; each start posi Von is specified, the length of peptidle is 9 amino acids, and tihe end position for each peptide is the start position plus eight, Pos 123456789 score 407 SCNPTAHLV 16 458 LALGOOVIA 16 637 YWLPIMVTSI 16 640 PIMTSILGA 16 52 GIVAWLYGD 15 141 YTKNRNFCL 15 225 FAQSWYWIL 15 250 LRLVA6PLV 15 264 GVLGVLAYG 15 275 YCWEEXRVL 15 366 LICIAYWAM 15 368 CIAY WAMTA 15 371 YWAMTALYL 15 374 MTALYILATS 15 406 TSCNPTAHL 15 433 GLIQRSVFN 15 443 QIYGVLGLF 15 491 FIRTLRYHT 15 573 SAKNAFMLL 15 657 SVFGMC VDl 15 663 VDTLFLCFL 15 TableXXlIl-V3-HLA-A0201 9mers-24P4C12 Each peptide is a porton of SEC ID NO: 7; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight- Pos 123456789 score 9 iTPPAJLPGI 22 6 WTNITPPAL 17 8 NITPPALPG 11 2 RCFPWT7NIT 10 TableXXlII-V5-HLA-A0201 9mers-24P4C12 Each peptide is a portion of SEQ ID NO: 11: each start position is specified, the length of peptide is 9 amino acids, and the end position for eacl3 peptide is the start position plus eight.

Pos 123456789 score ILLLVLIFL 28 rableXXIII-V5.HLA-A01- 9mers-44Cl2 Each pepfide is a portion Of SEQ ID NO: 11; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 1 VLEAILLLV 25 9 VLIFLRQRI 21 2 LEAILLLVL 20 6 LLLVL!FLR 19 3 EAILLLVLI 18 4 AILLLVLIF 18 7 LL\/LIFLRQ 13 8 LVLIFLRQR 13 TableXXI~ll-V6-HLA-A0201 9mears-24P4C12 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 2 YSSFKGLIPR 12 1 GYSSKG1LIP 7 3 SSKGLIPRS 7 8 IPRSVFRLQ 7 9 PRkSVFNLOI 7 6 GLIPRSVFN 5 TableXXIlIV7-HLA-A0201- 9mers-24P4C12 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptido is 9 amino acids, and the end position for each peptide is the start position plus eight, Pos 123456789 score I SWYPWILVAV 4 WILVAiLGQM 18 5 ILVAVGQMM 16 7 VAVGQ-MMST 13 8 AVGQMMSTM 12 6 LVAVGQMMS TableXYflI-V8-HLA-AO201- 9mors-24P24C1 2 PCT/US2002/038264 Each peptidle is a portion of SEQ ID NO: 17; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 4 WLPIMRNPI 19 7 IMRNPITPT 19 20 QTSILGAYV 1 7 10 NPITPIGHV 16 GHVFTSIL 12 15 TGHVFQTSI 11 18 VFQTSILGA 11 12 ITPTGHVFQ 5 LPIMRNPIT 9 13 TPTGHVFQT 9 TableXXIlI-V9HLA-A0201 9mers-24P34C1 2 Each peptide is a portion of SEQ I D NO: 19; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 9 PLPTOPATL 21 2 WAMTALYPL 15 ATLGYVLWA 6 ALYPLPTQP 16 12 TQPATLGYV 14 13 QPATLGYVL 14 16 TLGYVL WAS 14 5 TALYPLPTQ 13 4 MTALYPLPT 12 8 YPLPTQPAT 12 3 AMTALYPLP 11 TableXXIV-Vi -HLA-A0203- 9mners-24P4C112 Pos 1234567890 score NoResultsFound.

TableXXIV-V3-HLA*A0203- Simers-24P4C12 Pos 1234567890 score NoResultisFound.

TableXXIV-V5-HLA-A0203- 9mers-24P4C12 Pos 1234567890 score NoResultsFound.

WO 2004/050828 TableXXIV.V6.HLA-A0203- 9mers-24P4C12 Pos 1234567890 score NoResultsFound.

TableXXIV.V7-HLA*A0203- 9mers-24P4C12 Pos 1234567890 score NoResultsFound.

TableXXIV-Y8-HLA*A0203- 9mers-24P4C12 Pos 1234567890 score NoResultsFound.

TableXXIV-V9-HLA-A0203- 9mers-24P4C1 2 1234567890 score NoResultsFaund.

TableXXV--HL-A-A3*9mers- 24P Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptidle is the start position plus eight.

Pos 123456789 score 585 IVRWVLDK 29 424 CViFQGYSSK 27 64 VLYPRNSTG 26 135 'TVGEVFYTK 26 251 RLVAGPLVIL 26 506 ALILTLVQI 24 513 QIARVILEY 24 603 KLLWGGVG 24 690 SLLKILGKK 24 267 GVL[AYGIYY 23 282 VILRDKGASI 23 312 AGl VLAVLE 23 334 FIRIILLIK 23 102 SVAENGLQC 22 232 ILVALGVAL 22 247 ILLLRLVAG 22 443 QIYGVLGLF 22 464 VLAGAFASIF 22 516 RVILEYIDH 22 579 MLLMRNIVR 22 VVGIVAWLY 21 212 LNAIRDISVIK 21 281 R*VLRDKGAS 21 321 AILLLMLIF 21 338 IALLKEASK 21 339 ALLIKEASKA 21 376 ALYLATSGQ 21 TableXXV-Vl-HLA-A3-9mers- 24P Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456769 score 393 NISSPGCEK 21 517 VILEViIDH-K 21 593 KVTDLLLFF 21 619 SGRIPGLGK 21 621 RIP5GLGKDF 21 44 FILGYIVVG 20 56 WLYGDPRQV 20 243 SLLFILLLR 20 259 LVLILGVLG 20 347 AVGQMMSTM 20 363 VLLLICIAY 20 463 CVLAGAFAS 20 501 SLAFGALIL 20 606 WVGGVGVLS 20 689 KSLLKILGK 20 16 PVKYDPSFR 19 170 IFLLIPSAPAIL 19 186 NVTPPALPG 19 207 GLIDSLNAR 19 246 FILLLRLVA 19 249 LLRLVAGPL 19 260 VILGVLGV 19 262 ILGVLGVLA 19 298 NIL:SAYSVQ 19 317 AVLE-AILLL 19 333 RIRIAIALL 19 433 GLICIRSVFN 19 508 ILTLVQIAR 19 525 KLRGVONPV 19 560 YIMIAIYGK 19 588 'A'VL6KvT 19 604 LLWVGGVGV 19 605 LWGGVGVL-- 19 681 LDRIPYIMSIK 19 11 EAYGKPVKY 18 49 IWGIV AWIL 18 73 AYCGMGENK 18 220 KIFEIJFAQS 18 248 LLLRLVAGP 18 261 LILI3VLGVL 18 264 GVLGVLAYG 18 272 GIYYCWEEY 18 278 EEYRVLRDK 18 314 IVLAVLEAI 18 432 KGLIQRSVF 18 441 NLQIYGVLG 18 446 GVLGLF-wTL 13 PCT/US2002/038264 TableXXV-VI-HLA-A3-9mers- 24P Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 457 VLALGQCVL 18 564 AIY'GrQNFCV 18 587 RWVLDKVT 18 649 YVIASGFIFS 18 10 DEAYGKPVK 17 63 QVLYPRNST 17 121 PEDPWTVGK 17 177 ALGRCFPWT '17 211 SLNARDISV 17 233 LVALGVALV 17 235 AILGVALVILS 17 239 ALVLSLIFI 17 252 LVAGPLVLV 17 309 WLAALIVLA 17 335 RIAIALLIKE 17 365 LL!CIAYWVA 17 368 ClAY WAMTA 17 401 KVPINTSCN 17 421 GLMCVEMGY 17 456 WVVLALqCV 17 459 ALGQCVLAG 17 510 TLVQIARVI 17 542 KCCLWCLEK 17 562 MIAIYGKNF 17 580 LLMRNIVRV 17 583 RN! VIRVWL 17 644 SILGAY VIA 17T 657 SVFGMCVDT 17 662 CVDTLFLCF 17 26 PIKNRSCTD 16 34 DVICCVLFL 16 45 ILGYIWVGI 16 86 LLYFN1IFSC; 16 157 TVITSLQQE 16 165 ELCPSFLLP 16 237 GVALVLSLL 16 258 VLVLILGVL 16 289 SISQLGFTT 16 304 SVQETWLAA 16 323 LLLMLIFLR 16 364 LLLICIAYW 16 470 ASIFYW AIF-K 16 494 TLRYHTGSL 16 511 LVQIARVIL 16 554 IFLNRN AYIM 16 571 CVSAKNAFM 16 584 NIVWVLID 16 WO 2004/050828 WO 204/00828PCTIUS2002/038264I TableXXV-VI -HLA-A3-9mers- 24P Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptidle is 9 amino acics, and the end position for each peptidle is the start position plus eight.

Pos 123456789 score 673 DLERNNGSL 16 693 KILGKKNEA 16 698 KNEAPPDNK 16 DPSFRGPIK 15 48 YIWVGIVAW 15 58 YGDPROVLY 15 99 NIISVAENG 15 151 GVPWNMTVI 15 191 ALPGITNDT 15 231 WILVALGVA 15 234 VALGVALVL 15 257 LVLVLILGV 15 318 VLEAILLILM 15 322 ILLLMLIFL 15 327 L-lQl 15 329 FlJRQRIRIA 15 532 PVARCIMCO 15 589 WLDKTD 15 597 LILLIFFGKILL 15 598 LLFFGKLLV 15 622 IPGLGKDFK 15 645 ILGAYVIAS 15 651 IASGFFSVF 15 680 SLDRPYYMS 15 691 LLKILGKKN 15 7 DEDDEAYGK 14 42 LLFILGYIV 14 53 IVAWLYGOP 14 81 KDKPYLLYF 14 IllSIS 14 148 CLPGVPWVNM 14 171 LLPSAPALG 14 244 LLFILLLRL 14 311 AALIVLAVL 14 315 VILAVLEAIL 14 324 LLMLIFLRQ 14 326 Ml1l~R 14 337 AIALLIKEAS 14 359 LVTFVLLLI 14 370 AYWAMTALY 14 378 YLATS9PQ 14 388 VLWASNISS 14 453 TLNWVLALG 14 465 LAGAFASFY 14 487 LISAFIRTL 14 496 RYHTGSLAF 14 523 DHKLRGVQN 14 TabIeXX(V-VI .HLA.A3-9mers- 24P Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptd is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 527 RGVQNPVAR 14 528 GVQNPVARC 14 534 ARCIMCCFK 14 558 NAYIMIAIY 14 567 GKNFCVSAK 14 596 DLLLFFGKL 14 609 GVGVLSFFF 14 638 WILPIMTSIL 14 647 GAYVIASGF 14 665 TLFLCFLED 14 685 YYMSKSLLK 14 694 ILGKKNEAP 14 699 NEAPPIDNKK 14 701 APPDNKKRK 14 TableXXV-V3.HLA1-A--mers.

241141312 Each peptidle is a portion of SEQ I D NO: 7; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight Pos 123456789 score 8 NITFPALPG 17 TableXXV-V5-HLA-A3-9mners- 24P12 Each peptice is a portion of SEQ I D NO: 11; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptidle is the start position plus eight.

Pos 123456789 score 4 AILLLVLIF 21 8 LVLIFLRQR 20 5 ILLLVLIFL 16 6 LILLVILIFILR 16 1 VLEAILLLV 15 7 ILLVILIFLIRQ 14 9 VI-lFlER 14 TabIeXKV.V6-HLA.A3-9mers- 24P12 Each peptidle is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 6 GLIPRSVFN 22 5 KGJ~IPRSVF 18 7 LIPRSVFNL 11 TableXXV-W-HLA-A3-9mrers- 24P34C12 Each peptidle is a portion-of SEQ ID NO: 15; each start position is specified, the length of peptidle is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 8 AVGOMMSTM 5 ILVAVGQMM 19 6 LVAVGC MMS 4 WILVAVGQM 14 3 YWILVAVGQ 12 1 SWYWILVAV TableXXV.V8-HILA-A3.9mers- 24P12 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptidle is 9 amino acids, and the end position for each peptidle is the start position plus eight.

Pos 123456789 score I1I PIIPIGHYF 22 6 PIMRNPITP 16 4 WLPIMRNPI 12 9 RNPITPTGH 11I 1 INYYWEPIMIR 17 HVFQTSILG TablaXXV-V9HLA-3*9ers- 24P4C1 2 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 6 ALYPLPTQP 9 PLPTQPATL 18 11 PTQPATLGY 12 WO 2004/050828 16 TLGYVLWAS 12 TableXXVI-V1 .HLA-A26- Simers-24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptidle is the start position plus eight.

Pos 123456789 score 34 DVICCVLFL 35 49 IWGIVAWL 28 483 PTFPLISAF 28 605 LWGGVGVL 27 593 KVTDLLLFF 26 317 AVLEAILLL 25 592 DKVTDLLLF 25 135 EVFYTKNRN 24 240 LVLSLLFIL 24 589 WVLDKVTDL 24 38 CVLFLLFIL 23 237 GVALVLSLL 23 11 EAYGKPVKY 22 267 GVLAYGIYY 22 285 DKGASISQL 22 452 WTLNWVLAL 22 WIVAWLY 20 79 ENKOKPYLL 20 157 TVITSLQQE 20 263 LGVLGVLAY 20 446 GVLGLFWT7L 20 628 DFKSPHLNY 20 641 IMTSILGAY 20 662 OVOTIFLOF 20 236 LGVALVLSL 19 258 VLVLILGVL 19 307 ETWILMALIV 19 320 EAILLLMLI101 19 414 LVNSSCPGL 19 437 RSVFNLQIY 19 513 QIARVILEY 19 609 GVGVLSFFF 19 673 OLERNNGSL 19 32 CTDVICCVL 18 198 DTTIQQGIS 18 200 TIQQGISGL 18 204 GISGLIOSL 18 244 LLFILLLIRL 18 294 GFTTNLSAY 18 354 TMFYPLVTF 18 360 VTFVLLLIC 18 400 EKVPINTSC 18 511 LVQIARVIL 18 596 DLLLFFGKL 18 102 SVAENGLQC 17 TableXXVI-V -HLA-A26- 9mers-24P4C12 Each peptidle is a portion of SEQ ID NO: 3; each start position is specified, the length of peptidle is 9 amino acids, and the end position for each peptide is the start Position plus eight.

Pos 123456789 score 184 WTNVTPPAL 17 216 DISVKIFED 17 261 LILGVLGVL 17 358 PLVTFVLLL 17 438 SVFNLQIYG 17 442 LQIYGVLGL 17 443 QIYGVLGLF 17 467 LISAFIRTL 17 608 GGVGVLSFF 17 664 DTLFLCFLE 17 Tab leX)(VI-V3-HLA-A26-gmers- 24P11 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptidle is 9 amino acids, and the end position for each peptide-is the start position plus eight.

Pos 123456789 score 6 WTNITPPAL 17 9 ITPPALPGI 13 TabIeXXVI-V5-HLA-A26-9mners- 24P14CI2 Each peptidle is a portion of SEQ ID NO: 11; each start position is specifed, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 scor e 3 EAILLLVLI 19 4 AILLLVLIF 1B 8 LVUJFLRQR 15 2 LEAILLLVL 14 5 ILLLVLIFL 13 Table)OMIV-HIIA-A26-9mers.

24M1 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 9 amino acids, and tne end position for each peptide is the start position plus eight.

PCT/US2002/038264 123456789 score LIPRSVFNL 16 KGLIPRSVF 9 TableXXVI-V7-HLA-A26-9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 16; each start position is specified, the length of peptidle is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 8 AVGQMMSTM 12 6 LVAVGQMMS 11 4 WILVAVGQM 1 SWYWLVAV 8 5 ILVAVGQMM 6 2 WYWILVAVG 7 VAVGQMMST TableXXVI-V8-HLA-A26-9mers- 24P14C 12 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptidle is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 19 FQTSILGAY 11 PITPTGHVF 17 HVFQTSILG 16 GHVFQTSIL 13 20 QTSILG3AYV 14 PTGHVFQTS 9 TableXXVI-V9-HLA-A26-9mers- 24P4C1 2 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 11 PTQPATLGY 15 ATLGYVLWA 13 2 WAMTALYPL 12 13 QPAYLGYVL 4 MTALYPLPT 9 9 PLPTQPATL 9 TableXXVII-V1 -HLA-B0702gmiers-24412 WO 2004/050828 WO 204100828PCTIUS2002/038264t Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 255 GPLVLVLIL 23 357 YPLVTFVLL 23 683 RPYYMSKSL 21 149 LPGVPWNMT 20 396 SPGCEKVPI 20 482 IPTFPLISA 20 631 SPHLNYYWL 20 KPVKYDPSF 19 152 VPWNMTVIT 19 167 CPSFLLPSA 19 GPIKNRSCT 18 172 LPSAPALGR 18 83 KPYLLYFNI 17 188 TPPALPGIT 17 192 LPGITNDTT 17 57 LYGDPROVL 16 232 ILVALGVAL 18 253 VAGPLVLVL 16 479 PQDIPTFPL 16 503 AFGALILTIL 16 49 IWGIVAWL 15 120 CPEDPWTVG 15 175 APALGRCFP 15 189 PPALPGITN 15 234 VALGVALVL 15 251 RLVAGPLVL 15 381 TSGQPQYVL 15 406 TSCNPTAHL 15 583 RNIVRVVVL 15 617 FFSGRIPGL 15 DPSFRGPIK 14 34 DVICCVLFL 14 66 YPRNSTGAY 14 204 GISGLIDSL 14 236 LGVALVLSL 14 252 LVAGPLVILV 14 291 SQLGFTTNL 14 311 AALIVLAVL 14 317 AVLEAILLL 14 333 RIRIAIALL 14 351 MMSTMFYPL 14 419 CPGLMCVFQ 14 452 WTLNWVLAL 14 499 TGSLAFGAL 14 605 LWVGGVGVL 14 660 GMCVDTLFL 14 DPRQVLYPR 13 100 IIVAENGL 13 110 CPTPQVCVS 13 164 QELOPSELL 13 rableXXVIl-Vl-HLA-BO72- 9mers-24P4CI2 Each peptide is a portion or SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 170 FLLPSAPAL 13 182 FPWTNVTPP 13 228 SWYWILVAL 13 241 VLSLLFILL 13 249 LLRLVAGPL 13 261 LILGVLGVL 13 302 YQSVQETWL 13 319 LEAILLLML 13 358 PLVTFVLLL 13 369 lAY WAMTAL 13 371 YWAMTALYL 13 409 NPTAHLVNS 13 442 LQIYGVLGL 13 446 GVLGLFWTL 13 478 KPQDIPTFP 13 487 LISAFIRTL 13 494 TLRYHTGSL 13 501 SLAFGALIL 13 511 LVQIARVIL 13 590 VLDKVTDLL 13 622 IPGLGKDFK 13 651 IASGFFSVF 13 32 CTD\/ICCVL 12 78 GENKDKPYL 12 154 WNMTVITSL 12 184 WT-NVTPPAL 12 242 LSLLFILLL 12 244 LLFILLLRL 12 285 DKGASISQL 12 305 VQETWLAAL 12 308 1VJLAALIVL 12 315 VLAVLEAIL 12 322 ILLLMLIFL 12 356 FYPLVTFVL 12 373 AMIALYLAT 12 380 ATSGQPQYV 12 457 VLALGQCVL 12 525 KLRGVQNPV 12 547 CLEKFIKFL 12 572 VSAKNAFMIL 12 589 WLDKVrDL 12 591 LDKVTDLLL 12 626 GKDFKSPHL 12 658 VFGMCVDTL 12 701 APPDNKKRI( 12 28 KNRSCTO)VI 11 45 ILGYIVVGI 11 79 ENKOKPYLL 11 TableXXVII-Vl-HLA-BOT02- 9mners-24P4IC12.

Each peptidle is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position fbr each peptide is the start position plus eight.

Pos 12345637139 score 104 AENGLQCPT 11 107 GLQCPTPQV 11 109 QCPTPQVCV 11 112 TPQVCVSSC 11 123 DPWTVGKNE 11 163 QQELCPSFL 11 169 SFLLPSAPA 11 177 ALGRCFPWT I11 101 ALPGITNDT 11 237 GVALVLSLL 11 239 ALVLSLLFI 11 258 VLVLILGVL 11 262 ILGVLGVLA 11 275 YCWEEYRVL 11 310 LAALIVLAV 11 332 QRIRIAIAL 11 34 EASKAVGQM 11 354 TMFYPLVTF 11 384 QPQYVLWAS 11 414 LVNSSCPGL 11 426 FQGYSSKGL 11 434 LIQRSVFNL 11 44C FNLQIYGVL I1I 450 LFWTLNW'/L 11 464 VILAGAFASF I11 618 ILEYIE)HKL 11 531 NPVARCIMC III 537 IMCCFKCCL 11 571 CVSAKNAFM 11 573 SAKNAFMLL 11 574 AKNAFMLLM I11 596 DLLLFFGKL 11 597 LLLFFGKLL 11 599 LFFGKLLVV 11 638 WLPIMTSIL I1' 663 VDTLFLCFL 11 686 YMSKSLLKI 11 702 PPDNKKRKK 11I TableXXiI-V3-HLA-80702- 9mers-24P4C12 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptile is the start position plus eight.

Pos 123456789 score WO 2004/050828 WO 204/00828PCTIUS2002/038264 rableXXVI-V3-HLA-B0702- 9mers-24P4Cl2 Each peptide is a portion of SEQ 1I) NO: 7; each start position is specif ed, the length of peptidle is 9 amino acids, and the end position for each peptidle is the start position plus eight.

Pos 123456789 score 4 FPWTNITPP 12 6 WTNITPPAL 12 1 GRCFPWVTNI 10 2 RCFPWTrNIT 9 PWTNITPPA 9 9 ITPPALPGI 9 8 NITPPALPG 7 TableXXVII-V5-HLA-120702- 9mers-24P14C12 Each peptide is a portion of SEQ ID NO: 11; each start position is specified, the length of peptide is 9 amino acids, and the end position, for each peptidle is the start position plus eight.

Pos 123456789 score 2 LEAILLLVL 14 6 ILLLVLIFL 12 4 AILLLVLIF 11 1 VLEAILLLV 9 3 EAILLLVLI 9 9 VLIFLRQRI 7 TableXXVII-V6*HLA-B10702gmers.24P4C12 Each peptidle is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peplide is the start position plus eight.

Pos 123456789 score 8 IPRSVFNLQ 14 KGLIPRSVF 12 7 LIFRSVFNL 11 9 PRSVFNLQI 10 4 SKGLIPRSV 7 TableXXVII-V1-HLA-B0702* 9mners-24P4C1 2 Each peptidle is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptidle is the start position plus eight.

Pos 123456789 scor SWYWILVAV 9 IL VAVGQM~M 9 AVGQMMSTM 9 VAVGQMMST 8 WILVAV[SQM 7 TableXXVI1---A-10702-9mers- 24P4C1 2 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123466789 score 19 FQTSILGAY 20 11 PITPTGHVF 15 17 HVFQTSILG 15 16 GHVFQTSIL 13 20 QTSILGAYV 10 14 PTGHVFQTS 9 rableXXVIl-V9-HLA-80702-9mors- 24P4C1 2 Each peptide is a portion of SEQ ID NO: 19 each start position is specfied, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 13 QFATLGY/L 23 8 YPLPTOPAT 19 10 LPTQPATLG 14 15 ATLGYVLWA 13 2 WVAMTALYPL 12 7 LYFLPTQPA 11 9 FLPTQPATL 11I TableXXVIII-VI -HLA-B08-9meors Each peptide is a portion of SEQ ID NO: 3; each start position Is specified, the length of peptide is 9 amino adds, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 79 ENKDKPYLL 32 141 YTKNRNFCL 29 282 VLRDKGASI 29 573 SAKNAFMLL 26 249 LLRLVAGPL 23 494 TLRYHTGSL 23 26 PIKNRSCTD 22 329 FLRQRIRIA 22 TableXXVIIIIHLA-BO8-9mers Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptidle is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 589 WLDKVTDL 22 333 RIRIAIALL 21 583 RNIVRVWVL 21 591 LDKVTDLLL 21 626 GKDFKSPHL 211 687 MVSKSLLKIL 21 340 LLKEASKAV 474 WVAFHKPQDI 523 DHKLRGVQN 540 CFKCCLWCL 617 FFSGRIPGL 2 GGKQRDELJD 19 232 ILVALGVAL 19 255 GPLVLVLIL 19 631 SPHLNYYWL 19 694 ILCKKNEAP 19 139 VFYTKNRNF 18 170 FLLPSAPAL 18 241 VILSLLIFILL 18 247 ILLLRLVAG 18 268 VLVLILGVL 18 315 \/LAVLEAIL 18 322 ILILLMLIFL 18 357 YPLVrFVLL 18 457 VLALGQCVL 18 501 SLAFGALIL 18 514 IARVILEYI 18 518 ILEYIDHKL 18 .546 WCLEKFIKF 18 547 CLEKFIKFL 18 683 RPYYMSKSL 18 11 EAYGKPVKY 17 213 NARDISVKI 17 216 DISVKIFED 17 358 PLVTFVLLL 17 533 VARCIMCCIF 17 590 VLDKVTDLL 17 598 DLLLFFGKL 17 597 LLLFFGKLL 17 873 DLERNNGSL 17 691 LLKILGKKN 17 45 ILGYIVVGI 16 64 VLYPRNSTG 16 81 KDKPYLLYF 16 100 IISVAENGL 16 158 VITSLQQEL 16 204 GISGLIDSL 16 211 SLNARDISV 16 244 LLFILLLRL 16 WO 2004/050828 WO 204/00828PCTIUS2002/038264 TableXXVIII.Vl.HLA-B08-9mers Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptidle is the start position plus eight.

Pos 123456789 score 251 RILVAGPLVIL 16 253 VAGPLVLVL 16 338 IALLKEASF( 16 369 lAY WAMTAL 16 433 GILIQRSVIFN 16 551 FlKFLNRNA 16 638 WILPIMTSIL 16 702 PPDNKKRKK 16 VICCVLFLL 15 200 TIQQGISGL 15 225 FAQSWYWIL 15 234 VALGVALVIL 15 316 ILAVLEAILL 15 331 RORIRIAIA 15 396 SPGCEKVPI 15 434 LIQRSVFNL 15 487 LISAFIRTL 15 553 KFLNRNAYI 15 564 AIYGKNFCV 15 579 MLLMRNIVR 15 693 IKILGIKKNEA 15 TableXXVIIl*V3-H11A*B1108-9mers- 24P4G12 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 6 WTfNITPPAL 11 4 FPWTNITPP 8 1 GRCFPWTNI 7 9 ITPPALPGI 7 TableXXVlll-V5-B1108-9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 11; each start position is specified, the length of peptidle is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score ILLLVLIFL 18 3 EAILLLVLI 14 9 VLIFLRQRI 13 4 AILLLVLIF 12 TableXXVlll.8-B-108-9nlers- 24P4C12 Each peptide is a portion of SEQ I D NO: 11; each start position is specified, the length of peptidle is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 2 LEAILLLVL 10 6 LLLVLIFLR 3 TableXXVlll.V6-HLA-BO18-9mers- 24P4012 Each peptide is a portion of SEQ ID NO: 13; each start position is specif ed, the length of peptide is 9 amino acids, and the end positon for each peptide is the start position plus eight Pos 123456789 score 6 GILIPIRSVIFN 16 7 LIPRSVFNL 15 3 SSKGLIPRS 13 8 IPRSVFNLQ 13 1 GYSSKGLIP 11 9 PRSVFNLQI 8 TabieXXVIII.V7.HL1A.B308-9mers.

241P4IC12 Each peptide is a portion of SEQ ID NO: 15; each start positin is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 5 ILVAVGQMM 7 4 WILVAVGQM 6 7 VA\/GQMMST 5 1 SWYWILVAV 4 TableXXVIII.IIVS.HL-A.B108-9mers- 24P4C1 2 Each1 peptide is a portion of SEQ ID) NO: 17; each start position is specifed, the length of peptido is 9 amino acids, and the end position for each peptdle is the start position plus eight.

Pas 123456789 score 5 LPIMIRNPIT 15 4 WLPIMRNPI 12 16 GHVFQTSIL 11 11 PITPTGHVF 10 7 IMRNPITPT 8 13 TPTGHVFQT 7 TableXXVIII-V8.HLA-B08-9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight Pos 123456789 score 15 TGHVFQTS 7 Table)OW11III-V9-HL11A-1308- 9mers-24P4C12 Each peptide is a portion of SEQ D NO: 19; each start position is specified, the length of peptIde is 9 amino acids, and the end position for each peptide is the start position plus eight Pos 123456789 score 9 PLPTQPATL 16 13 QPATLGYVL 16 2 WAMIVTALYIPL 14 16 TLGYVLWAS 8 18 GYVLWASNI 8 8 YPLPTQPAT 7 TableXXIX-V1 -HLA-B1 510-9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptidle is the start position plus eight.

Pos 123456789 score 275 YCWEEYRVL 16 583 RNIVRVWVL 16 57 LYGDPRQVL 232 ILVALGVAL 253 VAGPLVLVL 381 TSGQPQYVL 487 LISAFIRIL 605 LWGGVGVL 49 IWGIVAWL 14 78 GENKDKPYL 14 100 IISVAENGL 14 170 FLLPSAPAL 14 184 WTN VIPPAL 14 200 TIQQGISGL 14 204 GISGLIDSL 14 251 RLVAGPLVL 14 357 YPLVTFVLL 14 369 lAY WAMTAL 14 457 VLALGQCVL 14 617 FFSGRIPGL 14 32 CTDVICCVL 13 WO 2004/050828 WO 204/00828PCTIUS2002/038264 TableXXIX-VI.HLA*B1 510*9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 79 ENKDKPYLL 13 228 SWYWILVAL 13 234 VALI3VALVL 13 255 GPLVLVLIL 13 261 LILGVLGVL 13 302 YQSVQETWL 13 308 TWLAALIVL 13 440 FNLOIYGVL 13 446 GVLGLFW-TL 13 499 TGSLAFGAIL 13 511 [VQIARVIL 13 518 ILEYIDHKL 13 537 IMCCFKCCL 13 547 CLEKFIKFL 13 572 VSAKNAFML 13 163 QQELCFSFL 12 237 GVALVLSLL 12 244 LLFILLILRL 12 258 VLVLILGVL 12 305 VQETWLAAL 12 311 AALIVLAVL 12 315 VILAVLEAIL 12 317 AVLEAILLL 12 322 ILLLMLIFL 12 356 FYPLVTFVL 12 371 YWAMTALYL 12 406 TSCNPTAHL 12 412 AHLVNSSCP 12 442 LQIYGVLGL 12 450 LFWTLNWVL 12 452 WTLNWVLAL 12 476 FHKPQDIPT 12 497 YHTGSLAFG 12 501 SILAFGAILIL 12 503 AFGALILTL 12 523 DHKLRGVQN 12 589 WVLDKVTDL 12 626 GKDFKSPHL 12 651 IASGFFSVF 12 658 VFGMCVDTL 12 660 GMCVDTLFL 12 673 DLERNNGSL 12 34 DVICCVLFL 11 88 YFNIFSCIL 11 141 YTKNRNFCL 11 154 WNMTVITSL 11 158 VITSLQQEL 11 164 QELCPSFLL 11 TableXXIX-VI -HILA.BI1 51 Q-9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 236 LGVALVLSL 11 241 VLSLLFILL 11 242 LSLLFILLL 11 286 DKGASISQL 11 291 SQLGFTTNL 11 319 LEAILLLML 11 332 QRIRIAIAL 11 333 RIRIAIALL 11 351 MMSTMFYPL 11 354 TMFYPLVTF 11 358 PLVTFVLLL 11 414 LVNSSCPGL 11 434 LIQRSVFNL 11 479 PODIPTFPL 11 494 TLRYHTGSL 11 590 VLDKVTDLL 11 591 LDKVTDLLL 11 631 SPHLNYYWL 11 684 PYYMSKSLL 11 35 VICCVLFLL 10 38 CVLFLLFIL 10 124 PWTVGKNEF 10 225 FAQSWYWIL 10 240 LVLELLFIL 10 249 LLRLVAGPL 10 316 LAVLEAILL 10 343 EASKAVGQM 10 418 SCPGLMOVF 10 426 FQGYSSKGL 10 477 HKPQDIPTF 10 483 PTFPLISAF 10 540 CFKCCLWCL 10 573 SAKNAFMLL 10 596 DLLLFFGKL 10 597 LLLFFGKLL 10 632 PHLNYYWLP 10 638 WLPIMTSIL 663 VDTLFLCFL 10 666 LFLCFLEDL 10 683 RPYYMSKSL 10 687 MSKSLLKIL 10 33 TDVICCVLF 9 36 ICCVLFLLF 9 217 ISVKIFEDF 9 347 AVGQMMSTM 9 432 KGLIQRSVF 9 461 GQCVLAGAF 9 607 VGGVGVLSF 9 TableXXIX-VI-HL-A-1I510-9mers- 24P4C12 Each peplide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptidle is the start position plus eight.

Pos 123456789 score 679 GSLDRPYYM 9 15 KPVKYDPSF 8 81 KDKPYLLYF 8 132 FSQTVGEVF 8 139 VFYTKNRNF 8 148 CLPGVPWNM 8 182 LQQELCPSF 8 174 SAPALGRCF 8 287 GASISQLGF 8 415 VNSSCPGLM 8 464 VLAGAFASF 8 468 AFASFYWAF 8 496 RYHTGSLAF 8 530 QNPVARCIM 8 570 FCVSAKNAF 8 608 GGVGVLSFF 8 609 GVGVLSFFF 8 647 GAYVIASGF 8 48 YIWVGIVAWV 7 69 NSTGAYCGM 7 214 ARDISVKIF 7 238 VALVLSILLF 7 318 VLEAILLLM 7 321 AILLLMLIF 7 366 LICIAYWAM 7 443 QlYGVLGLF 7 533 VARCIMCCF 7 546 WCLEKFIKF 7 554 FLNRNAYIM 7 562 MIAIYGI NF 7 571 CVSAKNAFM 7 574 AKNAFMLLM 7 593 -(VTDLLLFF 7 621 RIPGLGKDF 7 634 LNYYWLPIM 7 653 SGFFSVFGMv 7 TableXXIX-V3-HLA*BI 51 O-gniers- 24P4CI2 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 9 amino acids, and the and position for each peptide is the start position plus eight, Pos 123456789 score 6 WNITPPAL 13 WO 2004/050828 WO 204/00828PCT/US2002/038264 TableXXIX-V5-B151O9mers- 24p4C12 Each peptide is a portion of SEQ ID NO: 11; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 Score 2 LEAILLLVL 13 IILLLVILIFL 12 TableXXIX-V6-BI 510-9mers- 24P34C1 2 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptidle is the start position plus eight.

Pos 123456789 score 7 LIPRSVFNL 11 KGLIPRSVF 10 3 SSKGLIPRS 5 6 GLIPRSVFN 5 rableXXIX-V7-Bi5I0-9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptidle is 9 amino acids, and the end position for each peptidle is the start position plus eight.

Pos 123456789 score 8 AVGQMMSTM 9 4 WILVAVGQM 8 ILVAVGQMM 8 I SWYWILVAV 3 2 WYWILVAVG 3 3 YWILVAVGQ 3 6 LVAVGQMMS 3 TableXXlX-V8-B1 51 O-9mers- 24P4C12 'Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptidle is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 16 GHVFQTSIL 21 11 PITPTGHVF 10 13 QPATLGYVL 13 9 PLPTQPATL 12 2 WAMTALYPL 10 TableXXIX.V9.B1510-9mers-24P4C12 Each peplide is a portion ot SEQ ID NO: 19; each start position is specified, the length of peptide is 9 amino acids, and the end positon for each peptide is the start position plus eight.

Pos 123456789 score 13 QFATLGYVL 13 9 PLPTQPATL 12 2 WAMTALYRIL 10 TableX)O(-VI-HLAB2705-9mers.

24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 334 IRIAIALLK 28 332 ORIRIAIAL 25 675 ERNNGSLDR 24 214 ARDISVKIF 23 534 ARCIMCCFK 21 620 GRIPGLGKD 21 5 QRDEDDEAY 20 204 GISGILIDSL 20 446 GVLGLFVVrL 20 689 KSLLKILGK 20 251 RLVAGPLVL 19 424 CVFQGYSSK 19 436 QRSVFNLQl 19 483 PTFPLISAF 19 583 RNIVRVVVL 19 608 GGVGVLSFF 19 15 KPVKYDPSF 18 22 SFRGPIKNR 18 179 GRCFPWTNV 18 200 TIQQGISGL 18 207 GLIDSLNAR 18 234 VALGVALVL 18 244 LLFILLLRL 18 255 GPLVLVUIL 18 291 SQLGFTTNL 18 317 AVLEAILLL 18 330 LRQRIRIAI 18 333, RIRIAIALL 18 496 RYHTGSLAF 18 527 RGVQNPVAR 18 647 GAY VIASGF 18 668 LCFLEDLER 16 683 RPYYMSKSL 18 690 SLLKILGKK 18 49 IWVGIVAWL 17 78 GENKDKPYL 17 154 WNMTVITSL 17 TableXXX-VI.HILA-B32705-9mers- 24P4CI2 Each peptidle is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptidle is the start position plus eight.

Pos 1 23456789 score 237 GVALVLSLL 17 242 LSLLFILLL 17 261 LILGVLGVL 17 287 G3ASISQLGF 17 311 AALIVLAVL 17 338 IA[LKEASK 17 354 rMFYPLVTF 17 381 TSGQPQYVL 17 429 YSSKGLIQR 17 477 H-KPQDIPTF 17 503 AFGALILTL 17 516 RVILEYIDH 17 546 WCLEKFIKF 17 549 EKFIKFLNR 17 605 LWVGGVGVL 17 621 RIPGLGKDF 17 11 EAYGKPVKY 16 23 FRGPIKNRS 16 137 GEVFYTKNR 16 139 VFYTKNRNF 16 170 FLLPSAPAL 16 283 LRDKGASIS 16 285 DKGASISQL 16 321 AILLLMLIP 16 322 ILLLMLIFL 16 323 LLLMLIFLR 16 327 LIFLRQRIR 1 432 KGLIQRSVF 16 440 FNLQIYGVL 16 442 LQIYGVLGL 16 443 QIYGVLGLF 16 457 VLALGQCVL 16 508 ILTLVQIAR 16 517 VILEYIDHK 16 589 WLDKVTDL 16 617 FFSGRIPGL 16 626 GKDFKSPHL 16 699 NEAPPDNKK 16 10 DEAYGKPVK 40 LFLILFILGY 60 DPRQVLYPR 73 AYCGMGENK 81 KDKPYLLYF 124 PWTVGKNEF 212 LMARDISVK 217 ISVKIFEDF 228 SWtYWILVAL 236 LGVALVLSL 1s WO 2004/050828 WO 204/00828PCT/US2002/038264 TableXXX-I LA-B32705-9miers.

24P4C1 2 Each peptide is a portion of SEQ ID NO: 3;1 each start position is specified, the length of peptide is e amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 238 VALVLSLLF 15 243 SILLFILLLR 15 253 VAGPLVLVL 15 258 VLVLILGVL 15 308 TWVLAALIVIL 15 316 LAVLEAILL 15 369 IAY WAMTAL 15 461 GQCVLAGAF 15 470 ASFYWAFHK 15 518 ILEYIDHKL 15 542 KCCLWCLEK 15 543 OOLWCLEKF 15 547 CLEKFIKFL 15 567 GKNFCVSAK 15 579 MLLMRNIVR 15 586 VRVWVLDKV 15 593 KVTrDLLLFF 15 596 DLLLFFGKL 15 607 VGGVGVLSF 15 609 GVGVLSFFF 15 622 IPGLGKDFK 15 651 IASGFFSVF 15 684 PYYMSKSLL 15 .698 KNEAPPONK 15 34 DVICCVLFL 14 38 CVLFLLFIL 14 61 PRQVLYPRN 14 CGMGENKDI( 14 83 KPYLLYFNI 14 84 PYLLYFNIF 14 135 TVGEVFYTK 14 148 CLPGVPWNM 14 158 VITSLQQEL 14 162 LQQELCPSF 14 164 QELCPSFLL 14 232 ILVALGVAL 14 240 LVLSLLFIL 14 263 LGVLGVLAY 14 267 GVLAYGIYY 14 272 GIYYCWEEY 14 278 EEYRVLRDK 14 325 LMLIFLRQR 14 379 LATSGQPQY 14 418 SCPGLMCVF 14 434 LIQRSVFNL 14 437 RSVFNLQIY 14 450 LFWILN\NVL 14 452 WTLNWVLAL 14 TableXXX.VI.HLA-B2705-gmers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 464 VLAGAFASF 14 485 FPILISAFIR 14 487 LISAFIRTL 14 488 ISAFIRTILR 14 489 SAFIRTLRY 14 501 SLAFGALIL 14 513. QIARVILEY 14 515 ARVILEYID 14 552 IKFLNRNAY 14 556 NRNAYIMIA 14 668 NAYIMIAIY 14 560 YIMIAIYGK 14 575 KNAFMLLMR 14 585. IVRWVVLDK 14 595 TDLLLFFGK 14 613 LSFFFFSGR 14 6.43 TSILGAYVI 14 659 FGMCVDTLF 14 660 GMCVDTLFL 14 679 GSLDRPYYM 14 700 EAPPIJNKKR 14 701 APPDNKKRK 14 702 PPDNKKRKK 14 7 DEDfDEAYGK 13 36 ICCVLFLLF 13 172 LPSAIPALGR 13 241 VLSLLFILL 13 249 LLRLVAGPL 13 250 LRLVAGPLV 13 273 IYYCWEEYR 13 275 YCWEEYRVL 13 280 YRVLRDKGA 13 294 GFTTNLSAY 13 319 LEAILLLML 13 347 AVGQMMSTM 13 348 VGQMMSTMF 13 349 GQMMSTMFY 13 356 FYPLVTFVL 13 357 YPLVTFVLL 13 358 PLVTFVLLL 13 363 VLLLICIAY 13 492 IRTLRYHTG 13 495 LRYHTGSLA 13 506 ALILTLVQI 13 526 LRGVQNPVA 13 545 LWCLEKFIK 13 570 FCVSAKNAF 13 572 VSAKNAFML 13 TableXXX-VI.HILA-B12705-9miers* 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 582 MRNIVRVWV 13 590 VLDKVTDLL 13 592 DKVTDLLLF 13 610 VGVLSFFFF 13 637 YWLPIMTSI 13 64 AYVIASGFF 13 653 SGFFSVFGM 13 666 LFLCFLEDL 13 681 LDRPYYMSK 13 682 DRPYYMSKS 13 685 YYMSKSLLK 13 686 YMSKSLLKI 13 29 NRSCTOVIC 12 32 CTDVICCVL 12 33 TDVICCVLF 12 35 VICOVIFIL 12 57 LYGDPRQVL 12 58 YGDPRQVLY 12 79 ENKDKPYLL 12 80 NKDKPYLLY 12 93 SCILSSNII 12 100 IISVAENGL 12 121 PECPWTFVGK 12 132 FSQTVGEVF 12 144 NRNFCLPGV 12 151 GVPWNMTVI 12 163 QQELCPSFL 12 190 PALPGITND 12 193 PGITNDTTI 12 239 ALVLSLLFI 12 276 CWEEYRVLR 12 302 YQSVQETWL 12 305 VQE1VWLAAL 12 315 VLAVLEAIL 12 320 EAILLILMLI 12 328 IFLRQRIRI 12 343 EASKAVGQM 12 3711 YWAMTALYL 12 386 QYVLWASNI 12 393 NISSPGCEK 12 406 TSCNPTAHL 12 414 LVNSSCPGL 12 421 GLMCVFOGY 12 426 FQGYSSKGL 12 468 AFASFYWAF 12 490 AFIRTLRYH 12 500 GSLAFGALI 12 510 TLVQIARVI 12 WO 2004/050828 WO 204/00828PCTIUS2002/038264 TableXXX.VI-HLA-B2705.9mers- 24P4C1 2 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 519 LEYIDHKLR 12 537 IMCCFKCCL 12 540 CFKCCLWNCL 12 553 KFLNRI\AYI 12 557 RNAYINIIAI 12 562 MIAIYGKNF 12 591 LDKVTDLLL 12 597 LLLFFGKLL 12 614 SFFFFSGRI 12 619 SGRIPGLGK 12 628 DFKSPHLNY 12 631 SPHLNYYWL 12 634 LNYYWVLPIM 12 658 VFGMCVDTL 12 662 CVDTLFLCF 12 663 VDTLFLCFL 12 673 DLERMNGSL 12 687 MSKSLLKIL 12 TableXXX-V3-HLA-B2705-9mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 1 GRCFPWTNI 24 6 WTNITPPAL 11 TabIeXXX-V5-HLA-B27U5-9mners- 24P4C12 Each peptidle is a portion of SEQ ID NO: 11; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 4 AILLLVLIF 17 ILLLVLIFL 17 6 LLLVLIFLR .16 2 LEAILLLVL 14 8 LVLIFLRQR 14 3 EAILLLVLI 12 9 VLIFLRQRI 11 TableXXX-V6.HLA-B2705-9mers- 24P41 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peplide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 9 PRSVFNLQI 19 5 KGLIPRSVF 17 2 YSSKGLIPR 16 7 LIPRSVFNL 14 3 SSKGLIPRS 9 TabloXXX-V7-HLA-B2705- Slmers-24P4C12 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 8 AVGQMMSTM 13 4 WILVAVGQM 12 5 ILVAVGQMM 11 3 YWILVAVGQ 6 TableXXX-V8-HLA-B2705-Smers- 24P4C12 Each peptide is a portion of SEQ ID NO: 17-1 each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 16 GI-VFQTSIL 15 1 NYYWLPIMR 14 8 MRNPITPTG 14 9 RNPITPTGH 14 '11 PITPTGHVF 12 15 TGHVFQTSI 11 19 FQTSILGAY 10 2 YYWLPIMRN 8 4 WLPIMRNPI 7 7 IMRNPITPT 7 17 HVFQTSILG 7 TableXXX.M.LA-122705-9mers- 24P4CI2 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 18 GYVLWASNI 13 QPATLGYVL 13 2 WAMTALYPL 12 9 PLPTQPATL 12 11 PIQPATLGY 6 ALYPLPTQP 8 15 ATLGYVLWA 7 TableXXXI-VI.-LA*B2709- 9merse-24P4IC112 Each peptide is a portion of SEQ ID NO: 3: each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 332 ORIRIAIAL 23 179 GRCFPWTNV 22 250 LRLVAGPLV 21 214 ARDISVKIF 436 QRSVFNLQ 144 NRNFCLPG\/ 19 330 [RORIRIAI 19 582 MRNIVRVWV 19 586 VRWVLDKV 19 255 GPLVLVLIL 17 583 RNIVRVWL 17 251 RLVAGPLVL 16 683 RPYYMSKSL 16 78 OENKDKPYL 170 FLLPSAPAL 334 IRIAIALLK 446 GVLGLFWTL 620 GRIPGLGKD 647 GAYVIASGE 660 GMCVDTLFL 49 IWVGIVAWL 14 228 SWYVVILVAL 14 234 VALGVALVL 14 244 LLFILLLRL 14 317 AVLEAILLL 14 333 RIRIAIALL 14 452 WTLNWVLAL 14 602 GKLLWVGGV 14 626 GKDFKSPHL 14 679 GSLDRPYYM 14 23 FRGPIKNRS 13 34 DVICCVLFL 13 83 KPYLLYFNI 13 WO 2004/050828 WO 204100828PCTIUS2002/038264 TableXXXI.VI-HLA-B2709- 9merse-24P34C1 2 Each peptide is a portion of SEQ NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptidle is the start position plus eight.

Pos 123456789 score 107 GLQCPFPQV 13 204 GISGLIDSL 13 232 ILVALGVAIL 13 236 LGVALVLSL 13 237 GVALVLSLL 13 240 LVLSLLFIL 13 242 LSLLFILLL 13 253 VAGPLVLVL 13 291 SQLGFTTNL 13 311 MALIVLAVL 13 322 ILLLMLIFL 13 357 YPLVTFVLL 13 358 PLVTFVLLL 13 369 lAY WAMTAL 13 4.40 FNLQIYGVL 13 442 LQIYGVLGL 13 449 GLFWILNWV 13 496 RYHTGSLAF 13 500 GSLAFGALI 13 515 ARVILEYID 13 557 RNAYIMIAI 13 589 WLDKVTDL 13 KPVKYDPSF 12 38 CVLFLLFIL 12 ILGYIWVGI 12 56 WVLYGDPRQV 12 61 PRQVLYPRN 12 81 KDKPYLLYF 12 158 VITSLQQEL 12 164 QELCPSFLL 12 258 VLVLILGVL 12 261 LILGVLGVL 12 287 GASISQLGF 12 308 TWLAALIVL 12 316 LAVILEAILL 12 321 AILLLMLIF 12 328 IFLRQRIRI 12 355 MFYPLVTFV 12 371 YWAMTALYL 12 414 LVNSSCPGL 12 432 KGLIQRSVF 12 434 LIQRSVFNL 12 461 GQCVLAGAF 12 492 IRTLRYHTG 12 495 LRYHTGSLA 12 501 SLAFGALIL 12 503 AFGALILTL 12 506 ALILTLVQI 12 TableXXXI-Vi .HLA-B2709fmerse-24F4IC12 Each peptide is a portion of SEQ I D NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 518 ILEYIDHKL 12 553 KFLNRNAYI 12 593 KVTDLLLFF 12 595 DLLLFFGKL 12 597 LLLFFGKLL 12 605 LVVGGVGVL 12 606 GGVGVLSFF 12 621 RIPGLGKDF 12 637 YINLPIMTSI 12 656 LFLCFLEDL 12 684 PYYMSKSLL 12 5 QIRDEUDEAY 11 28 KNRSCTDVI 11 29 NRSCTDVIC 11 32 CTDVICCVL 11 41 FLIFILGYI 11 42 LLFILGYIV 11 46 LGYIWVGIV 11 67 PRNSTGAYC Ii 79 ENKDKPYLL 11 87 LYFNIFSCI 11 100 IISVAENGL 11 128 GKNEFSQTV 11 139 VFYTKNRNF 11 151 GVPWVNMTVI 11 184 VVTNVTPPAL 11 217 ISVKIFEDF 11 225 FAQSWYWIL 11 230 Y'INILVALGV 11 238 VALVLSLLF 11 239 ALVLSLLFI 11 249 LLRLVAGPL 11 257 LVILVILILGV 11 260 VLILGVLGV 11 280 YRVLIRDKGA 11 283 LRDKGASIS 11 285 DKGASISQL 11 297 TNLSAYQSV 11 310 ILAALIVLAV 11 314 IVLAVILEAI 11 319 LEAILLLML 11 351 MMSTMFYPL 11 354 TMFYPLVTF 11 381 TSGQPQYVL II1 386 QYVLWVASNI 11 427 QGYSSKGLI 11 480 QlDIPTFPLI 11 483 PTFPLISAF 11 TableXXX-VI -HLA-B270g- 9merse-24P4C1 2 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptde is 9 amino acids, and the end position for each peptide is the start positior plus eight.

Pus 123455789 score 509 LTLVQIARV 11 510 TLVQIARVI 11 511 LVQIARVIL 11 526 LRG\/QNPVA 11 534 ARCIMCCFK 11 537 IMCCFKCCL 11 564 AIYGKNFCV 11 572 VSAKNAFML 11 591 LDKVTDLLL 11 592 DKVTDLLLF 11 598 LLFFGKLLV 11 b99 LFFGKLLWV 11 609 GVGVLSFFF 11 614 SFFFFSGRI 11 617 FFSGRIPGL 11 631 SPHLNYYWL 11 634 LNYYWLPIM 11 643 TSILGAYVI 11 653 SGFFSVFGM 11 658 VFGMCVDTL 11 663 VDTLFLCFL 11 675 ERNNGSLDR 11 687 MSKSLLKIL 11 TableXXXI-V3-HLA-B2709- 9mners24F4C1 2 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 1 GRCFPWVTNI 22 6 WTNITPPAL 11 9 ITPPALPGI 11 TableXXXI-V5-B32709-9mers- 2434CI12 Each peptide is a portion of SEQ ID NO: 11 each start position is specified, the length of peptide is 9 amino adds, and the end position for each peptde is the start position plus eight Pos 123456789 score 4 AILLLVLIF 13 WO 2004/050828 WO 204/00828PCT/US2002/038264 ILLLVLIFL 13 2 LEAILLLVL 11 1 VLEAILLLV 10 3 EAILLLVLI 10 9 VLIFLRQRI 10 TableXXXl.VBHLA-B2709.

9mers-24P4C1 2 Each peptidle is a portion of SEQ ID NO: 13, each start position is specified, the length of peptidle is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 9 PRSVFMLQI 20 KGLIPRSVF 12 7 LIPRSVFNL 12 4 SKGLIPRSV 9 TableXXXI-VW.HLA-B2709- 9mers.24P4C12 Each peptide is a porton of SEQ ID NO: 15; each start position is specified, the length of peptide is 9 amino acids, and the end position far each pepfide is the start position plus eight.

Pos 123456789 score I SWYWILVAV 12 4 WVILVAVGQM 12 ILVAVGQMM 10 8 AVGQM.MSTM 9 TableXXXI-V8-H-LA-B2709- 9mers-24P4C12 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of Peptidle is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 16 GHVFQTSIL 14 8 MRNPITPTG 13 11 PITPTGHVF 10 NPiTPTGHIV 9 4 WLPIMRNPI 8 TGHVFQTSI 8 QTSILGAYV 8 TableXXXI-V9-HLA-B2709- 9rmers*24114C12 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptidle is the start position plus eight Pos 123456789 score 18 GYVLWASNI 14 2 WAMTALYPIL 11 13 QPATLGY'/L 11 9 PLPTQPATL 10 12 TQPATLGYV 8 TabloXXXII-Vi -HLA-B44O2- 9mners-24P41 2W Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptidle is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 164 QELCPSFLL 22 319 LEAILLLML 22 222 FEDFAQSWY 21 78 GENKDKPYL 20 306 QETWLAALI 20 483 PTFPLISAF 20 317 AVLEAILLL 19 332 QRIRIAIAL 19 503 AFGAILILTL 18 506 ALILTLVQI 18 552 IKFLNRNAY 18 58 YGDPRQVLY 17 170 FLLPSAPAL 17 214 ARIDISVKIF 17 242 LSLLFILLL 17 583 RNIVRVVVI 17 11 EAYGKPVKY 16 40 LFLLFILGY 16 48 YIVVGIVAW 16 81 KDKPYLLYF 16 121 PEDPWVTVGK 16 228 SWYWILVAL 16 253 VAGPLVLVL 16 254 AGPLVLVLI 16 311 MALIVLAVL 1l6 320 EAILILLMLI 16 321 AILLLMILIF 16 363 VLLLICIAY 16 382 SGQPQYVLW 16 452 WTLNWVLAL 16 480 QDIPTFPLI 16 487 LISAFIRTIL 16 489 SAFIRTLRY 16 617 FFSGRIPGL 16 TableXXXII-V1-HLA-84402gmers-24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start posifion plus eight.

Pos 123456789 score 629 FKSP-LNYY 16 699 NEAPPDNKK 16 34 DVICCVLFL 79 ENKDKPYLL 130 NEFSQTVGE 154 WNMTVITSL 204 GISGLIDSL 234 VALGVALVL 241 VLSLLFILL 263 LGVLGVLAY 278 EEYRVLRDK 294 GFTTNLSAY 354 TMFYPLV-TF 370 AYWAMTALY 399 CEKVPINTS 442 LQIYGVLGL 468 AFASFYWAF 477 HKPQDIPTF 499 TGSLAFGAL 513 QIARVILEY 547 CLEKFIKFL 66 YPRNSTGAY 14 80 NKDKPYLLY 14 84 PYLLYFNIF 14 93 SCILSSNII 14 104 AENGLQCPT 14 193 PGITNDTVI 14 223 EDFAQSWYW 14 239 ALVLSLLFI 14 244 LLFILLLRL 14 258 VLVLILGVL 14 261 LILGVLGVL 14 285 DKGASISQL 14 291 SQLGFTTNL 14 301 AYQSVQETW 14 305 VQETWLAAL 14 308 TWLMALIVL 14 316 ILAVLEAILL 14 322 ILILLMLIFIL 14 330 LRQRIRIAI 14 333 RIRIAIALL 14 356 FYPLVTFVL 14 357 YPLVTFVLL 14 358 PLVTFVLLL 14 364 LLLICIAYW 14 418 SCPGLMCVF 14 432 KGLIQRSVF 14 446 GVLGLFWTL 14 WO 2004/050828 WO 204/00828PCT/US2002/038264 TableXX)UI-V1 -HLA-B4402- 9mners-24P41 2 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 496 RYHTGSLAF 14 546 WC[EKFIKF 14 558 NAYIMIAIY 14 573 SAKNAFMLL 14 577 AFMLLMRNI 14 592 DKVTDLLLF 14 593 KVTDLLLFF 14 596 DLLLFFG(L 14 597 LLLFFGKJ.. 14 621 RIPGLGKDF 14 641 IMTSILGAY 14 643 TSILGAYVI 14 651 IASGFFSVF 14 662 OVOTLELCE 14 671 LEDLERNNG 14 678 NGSLDRPYY 14 QRDEDDEAY 13 7 DEDDEAYGK 13 32 CTDVICCVL 13 36 ICC VLFLLF 13 49 IWVGIVAWL 13 57 LYGDPRQV[ 13.

77 MGENKDKPY 13 87 LYFNIFSCI 13 137 GEVFYTKNR 13 146 NFCLPGVPW 13 174 SAPALGRCF 13 176 PALGRCFPW 13 184 WTNVTPPAL 13 187 VTPPALPGI 13 200 TIQQGISGIL 13 209 IDSLNARDI 13 213 NARDISVKI 13 232 ILVALGVAL 13 237 GVALVLSLL 13 238 VALVLSLLF 13 251 RLVAGPLVL 13 255 GPLVLVLIL 13 277 WEEYRVLRD 13 342 KEASKAVGQ 13 351 MMSTMFYPL 13 440 FNLQIYGVL 13 443 QIYGVLGLF 13 448 LGLFWTLrJW 13 461 GQCVLAGAF 13 466 AGAFASFYWV 13 501 SLAFGALIL 13 518 ILEYIDHKL 13 Tab~eXXXII-VI.HLA-B4402- 9mners-24P4C1 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 519 LEYIDHKLR 13 529 VQNFVARCI 13 543 CCLVtICLEKF 13 570 FCVSAKNAF 13 589 VVLDKVTDL 13 590 VLDKVTIJLL 13 605 LWGGVGVL 13 631 SPHLNYYWL 13 637 YVVLPIMTSf 13 648 AYVIASGFF 13 674 LERNNGSLD 13 687 MSKSLLKIL 13 33 TDV1CCVLF 12 35 VICCVLFLL 12 38 CVLFLLFIL 12 50 WVGIVAWLY 12 100 IISVAENGL 12 Q32 FSQTVGEVF 12 133 SQTVGEVFY 12 139 VFYTKNRNF 12 141 YTKNRNFCL 12 163 QQELCPSFL 12 217 ISVKIFEDF 12 221 IFEDFAQSW 12 236 LGVALVLSL 12 240 LVLSLLFIL 12 249 LLRLVAGPL 12 267 GVLAYGIYY 12 269 LAYGIYYCW 12 275 YCWEEYRVL 12 287 GAS[SQLGF 12 314 IVLAVLEAI 12 326 MLIFLRQRI 12 328 IFLRQRIRI 12 349 GQMMSTMFY 12 369 lAY WAMTAL 12 371 YWAMTALYL 12 406 TSCNPTAHL 12 421 GLMCVFQGY 12 426 FQGYSSKGL 12 434 LIQRSVFNL 12 437 RSVFNLQIY 12 450 LFVVTLNWVL 12 457 VLALGQCVL 12 464 VLAGAFASF 12 479 PQDIPTFPL 12 510 TLVQIARVI 12 511 LVQIARVIL 12 TableXXXII-VI.HLA-B4402- 9mners-24P54C1 2 Each peptide is a portion of SEQ I D NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 548 LEKFIKFLN 12 553 KFLNRNAYI 12 557 RNAYIMIAI 12 562 MIAIYGKNF 12 572 VSAKNAFML 12 591 LDI VTDLLL 12 607 VGGVGVLSF 12 608 GGVGVLSFF 12 610 VGVLSFFFF 12 630 KSPHLNYYW 12 638 WLPIMISIL 12 647 GAYVIASGF 12 658 VFGMCVDTL 12 659 FGMCVDTLF 12 660 OMOVOTLEL 12 663 VDTLFLCFL 12 666 LFLCFLEDL 12 673 DLERNNGSL 12 677 NNGSLDRPY 12 683 RPYYMSKSL 12 686 YMSKSLLKI 12 10 DEAYGKPVK 11 15 KPVKYDPSF 11 28 KNRSCTDVI 11 37 CCVLFLLFI 11 41 FLLFILGYI 11 45 ILGYIVVGI 11 117 VSSCPEDPW 11 124 PWTVGKNEF 11 151 GVPWVNMVI 11 197 ND1TIQQGI 11 201 IQQGISGLI 11 266 LGVLAYGIY 11 302 YQSVQETWL 11 359 LVTFVLLLI 11 361 TFVLLLICI 11 379 LATSGQPQY 11 381 TSGQPQYVL 11 436 QRSVFNLQl 11 444 IYGVLGLFW 11 465 ILAGAFASFY 11 474 WAFHKPQDI 11 484 TFPLISAFI 11 494 TLRYHTGSL 11 533 VARCIMCCF 11 538 MCCFKCCLW 11 540 CFKCCLWCL 11 614 SFIFFFSGRI I1I WO 2004/050828 WO 204/00828PCTIUS2002/038264 TabOXXXII-VI -HLA-54402- 9mers-24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptidle is 9 amino acids, and the end position for each peptidle is the start position plus eight.

Pos 123456789 score 626 GKDFKSPHL 11 628 DFKSPHLNY 11 684 PYYMSKSLL 11 19 YDPSFRGPI 10 83 KPYLLYFNI 10 88 YFNIFSCIL 10 158 VITSLQQEL 10 162 LQQELCPSF 10 225 FAQS WYWIt 10 272 GIYYCWEEY 10 315 VLAVLEAIL 10 348 VGQMMSTMF 10 386 OWVLWASNI 10 396 SPGCEKVPI 10 414 LVNSSCPGL 10 500 GSILAFGALI 10 514 IARVILEYI 10 537 IMCCFKCCL 10 544 CLWCLEKFI 10 555 LNRNAYIMI 10 609 GVGVLSFFF 10 TableXXX11143-141A-114402.

9mers-24P4C12 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptidle is 9 amino acids, and the end posifton for each peptide is the start position plus eight.

Pos 123455789 score 6 WTNITPPAL 13 9 ITPPALPGI 13 1 GRCFPWTNI 8 2 RCFPVITNIT 7 7 TNITPPALP 6 8 NITPPALPG 6 Tab~eXXXII*V5-HLA-B4402- 9mners-24P4C12 Each peptide is a portion of SEQ NO: 11; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score ILEAILLILVIL 23 EAILLLVLI 17 AILLLVLIF 17 ILLLVLIFL 14 VLIFLRQRI 12 TableXXXII-V6-HLA-B4402gmers-24P4C1 2 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 9 amino acids, and the end position for each pepile is the start position plus eight.

Pos 123456789 score 5 KGLIPRSVF 14 7 LIPRSVFNL 13 9 PRSVFNLQI1 1I 6 GLIPRSVFN 8 TableXXXII-V7-HLA-B14402* 9mers-24P4C12 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptidle is the start position plus eight.

Pos 123456789 score 1 SWVYWILVAV 6 3 YWILVAVGQ 6 8 AVGQMMSTM 4 4 WVILVAVGQM 3 2 VVYWILVAVG 2 TableXXXII-VS-HLA-34402- 9mers-24P4C12 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 11 PITPTGHVF 15 19 FQTSILGAY 14 4 WVLPIMRNPI 11 16 GHVFQTSIL 11 15 TGHIVFQTSI 8 TableXXXII-V9-HLAB4402*9mers- 241P4C12 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456769 score 11 PTQPATLGY 9 PLPTQPATL 14 2 WVAMTALYPL 13 14 PATLGYVLW 13 13 QPATLGYVL 12 18 GYVILWASNI 6 ALYPLPTQP 8 15 ATILGYVILWA 7 TableXXiII.V1.HL11A-115101grmers-24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 234 VALGVALVL 27 213 NAIRDISVIKI 46 LGYIWVGIV 24 83 KPYLLYFNI 24 311 AALIVLAVL 24 253 VAGPLVLVL 23 310 LAALIVILAV 23 357 YPLVTFVLL 23 369 lAY WAMTAL 23 474 WAFHKPQDI 23 514 IARVILEYI 23 683 RPYYMSKSL 22 254 AGPLVLVLI 21 255 GPLVLVLIL 21 320 EAILLLMLI 21 396 SPGCEKVPI 21 427 QGYSSKGLI 21 11 EAYGKPVKY 193 PGITNDTTI 316 LAVILEAILL 123 OPWTVGKNE 19 236 LGVALVLSL 18 314 IVILAVLEAII 18 599 LFFGKLLW 18 686 YMSKSLLKI 18 60 DPRQVLYPR 17 150 PGVPWNMTV 17 225 FAQSWYWIL 17 261 LILGVLGVL 17 269 LAYGIYYCW 17 300 SAYQSVQEI 1T 504 FGALILTLV 17 WO 2004/050828 TableXXXIII.V1 .HLA.B51 01- 9mers-24P4C12 558 NAYIMIAIY 17 573 SAKNAFMILL 17 651 IASGFFSVF 17 182 FPWTNVTPP 16 192 LPGITND.TT 16 328 IFLRQRIRI 16 355 MFYPLVTFV 16 359 LVTFVLLLI 16 458 LALGQCVLA 16 502 LAFGALILT 16 505 GALILTLVQ 16 510 TLVQIARVI 16 581 LMRNIVRWV 16 631 SPHLNYYWL 16 9 DDEAYGKPV 15 ILGYIVVGI 15 56 WLYGOPRQV 15 110 CPTPQVCVS 15 120 CPEDPWT7VG 15 151 GVPWVNMTVI 15 172 LPSAPALGR 15 224 DFAQSWYWI 15 275 YCWVEEYRVL 15 308 TWLAALIVL 15 336 IAIALLKEA 15 338 IALLKEASK 15 375 TALYLATSG 15 485 FPLISAFIR 15 529 VQNPVARCI 15 564 AIYGKNFCV 15 582 MRNIVRVWV 15 5S6 DLIIFFGKL 15 637 YWLPIMTSI 15 643 TSILGAYVI 15 647 GAYVIASGF 15 700 EAPPDNKKR 15 DPSFRGPIK 14 41 FLLFILGYI 14 43 LFILGYIVV 14 72 GAYCGMGEN 14 87 LYFNIFSCI 14 119 SCPEDPWTV 14 152 VPWNMTVIT 14 188 IPPALPGIT 14 190 PALPGITND 14 209 IDSLNARDI 14 230 YWILVALGV 14 238 VALVLSLLF 14 257 LVLVLILGV 14 409 NPTAHLVNS 14 411 TAHLVNSSC 14 450 LFWTLNINVL 14 465 LAGAFASFY 14 467 GAFASFYWA 14 482 IPTFPLISA 14 Tab~eXXXIII.VI.HLA-B51 01 9rners.24P114C1 2 499 TGSLAFGAL 14 509 LTLVQIARV 14 576 NAFMLLMRN 14 586 VRWVVLDKV 14 589 WLDKVTDL 14 602 GKLLWGGV 14 605 LVVGGVGVL 14 639 LPIMTSILG 14 701 APPDNKKRK 14 702 PPDNKKRKK 14 19 YDPSFRGPl 13 28 KNRSCTDVI 13 34 DVICCVLFL 13 54 VAWLYGDPR 13 66 YPRNSTGAY 13 112 rPQVCVSSC 13 149 LPGVPWNMT 13 174 SAFALGRCF 13 176 PALGRCFPWV 13 187 VTPPALPGI 13 189 PPALPGITN 13 201 IQQGISGLI 13 239 ALVLSLLFI 13 252 LVAGPLVLV 13 282 VLRDKGASI 13 285 DKGASISQL 13 293 LGFTTNLSA 13 322 ILLILMILIFL 13 330 LRQRIRA 13 340 LLKEASKAV 13 343 EASKAVGQM 13 356 FYFLVTFVL 13 361 TFVLLLICI 13 384 QPQYVLWVAS 13 478 KPQDIPTFP 13 487 ILISAFIRTL 13 489 SAFIRTIRY 13 500 IGSLAFGALI 13 506 ALILTLVQI 13 521 YIDHKLRGV 13 531 NPVARCIMC 13 553 KFLNRNAYI 13 555 LNRNAYINII 13 563 IAIYGKNFC 13 578 FMLLMRNIV 13 580 LLMRNIVRV 13 TableXXXlIl-V3-HLA-B51 01gmers.24P4C12 Each peptide is a portion of SEQ ID NO: 7; each start position is specifed, the length of peptidle is 9 amino acids, and the end position for each peptidle is the start position plus eight.

PCTIUS2002/038264 123456789 score FPWTNITPP ITPPALPGI 14 GRCFPWTNI 11 WVTNITPPAL 8 TableX(XXIII-V5-HLA-B5101-9mers- 24P4C12 Each peptidle is a portion of SEQ ID NO: 11; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 3 EAILLLVLI 22 5 ILLLVLIFL 14 2 LEALLLVL 13 1 VLEAILLLV 12 9 VLIFLRQRI 12 TableXXXIII-V6-HLA-B5101- 9mers-24P4C12 Each peptidle is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start posiion plus eight.

Pos 123456789 score 8 IPRSVFNLQ 16 7 LIPRSVFNL 12 9 FRSVFNLQI 12 5 KGLIPRSVF 11 4 SKGLIPRSV TableXXXIII.W-HILA-B115101- 9mers-24P4C12 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide Is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score I SINYWILVAV 14 7 VAVGQMMST 12 2 ~WWILVAVG 6 3 YWILVAVGQ 6 TableXXXIII.VB.HLA-B5101-Smers- 24P4C12 WO 2004/050828 WO 204/00828PCT/US2002/038264 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptidle is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score NPITPTGHV 21 TGHVFOTSI 18 13 TPTGHVFQT 14 4 WLPIMRNPI 13 LPIMRNPIT 13 TableXXXIII-V9-HLA*B51 01- 9lmers-24P24C12 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is 9 amino acids, and the end position for each peptide is the start position plus eight.

Pos 123456789 score 13 QPATLGYVL 20 2 WAMTALYPL 18 TALYPLPTQ 16 8 YPLPTQPAT 15 LPTQPATLG 14 12 TQPATLGYV 13 17 LGYVLWASN 12 9 PLPTQPATL 11 14 PATLGYWLW 11 18 GYVLWASNI 11 TableXXXIV.VI.HLA.AI -l0mers.

24P4C1 2 Each peptidle is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 221 IFEDFAQSWY 25 488 ISAFIRTLRY 25 39 VLFLLFILI3Y 23 58 YGDPRQ VLYP 23 79 ENKDKPYLLY 23 262 ILGVLGVLAY 23 512 VQIARILEY 22 627 KDFKSPHLNY 21 132 FSQTVGEVFY 20 266 LGVLAYGIYY 20 362 FVLLLICIAY 20 590 VLDKVTFDLLL 20 594 VTDLLLFFGK 20 318 VLEAILLLML 19 32 CTDVICCVLF 18 TableXXXIV.VI-HLA-Ai -10mers- 24P4C1 2 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234557890 score 49 IWGIVAWLY 18 378 YLATSG-QPQY 18 420 PGLMICVFQGY 18 464 VIAGAFASFY 18 10 D)EAYGKPVKY 17 57 LYGDPRQVLY 17 121 PEDPWTVGKN 17 265 VLGV/LAYGlY 17 271 YGIYYCVNEEY 17 276 CWEEYRVLRD 17 369 IAY WAMTALY 17 551 FIKFLNRNAY 17 80 NIKDKPYLLYF 16 348 VGfQMMSTMFY 16 676 RNNGSLDRPY 16 677 NNGSLDRPYY 16 4 KQRDEDDEAY 15 18 KYDFSFRGPI 15 65 LYPRNS-TGAY 15 76 GMGENkDKPY 214 ARDISVKIFE 15 293 LGFT-TNL-SAY 15 436 QRSVFN!LQIY 15 479 PQDIPTFPLI 15 557 RNAYIMIAIY 15 628 DFKSPHLNYY 15 640 IMTSILGAY 15 664 DTLFLCFLED 15 283 LliDKGA~ISQ 14 521 Y]IDHKLR4GVQ 14 673 DLERNNGSLD 14 141 YIKNRNFCLP 13 305 VQETWLAALI 13 382 SGQPQYVLWA 13 407 SCNPTAHLVN 13 518 iLEYIDH-KLR 13 547 CILEKFIKFLN 13 670 F!LEDLERNNG 13 680 SLDRPY)YMSK 13 7 DEDDEAYGKP 12 35 VICCVLFLLF 12 159 ITSLQQELCP 12 163 QQELCPSFLL 12 242 LSLLFILJLLR 12 618 F'SGRIPGL-GK 12 626 GjKDFKSPHLN 12 698 KNEAPPDNKK 12 TableXXXIV-V3-HLA-AI Omers- 24P4C12 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567690 score 10 ITPPALPGIT 3 RCFPWTNITP 9 7 WTNITPPALP 8 8 TNITPPALPG 6 9 NITPPALPGI 4 TableXXXIV-V5-HLA-A1- I Omers.24P4C12 Each peptide is a portion of SEQ I D NO: 11; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 2 VLEAILLLVL 19 7 LLLVLIFLRQ 1 AVLEALLL-V 9 TableXXXIV-V6-HLA-AI -l0mors- 24P4C1 2 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 10 PRSVFNLQIY I OGYSSKf3LIP 7 4 SSKGLIPRSV 7 9 IP"RSVFNLQI 7 TableXXXIV-V7-HLA-AI -lrners- 24P4C12 Each peptide is a portion of SEQG ID NO: 15; each start position is specified, the length of peptidle is 10 amino acids, and the end position for each peptidle is the start position plus nine.

Pos 1234567890 score 1 QLSWYWILVAV 4 2 SWVYWILVAVG 4 4 YWVILVAVGQM 3 5 WILVAVGQMM 2 6 ILAVGQMMS 2 WO 2004/050828 8 VAVGQMMSTM 9 AVGQMMSTMF PCTIUS2002/038264 TableXXXIV-V8-HLA-AIl0mers-24P4C12 Each peptide is a portion of SEQ ID NO: 17; each start position is specifed, the length of peptide is amino acids, and the end position for each peptde is the start position plus nine.

Pos 123456890 score 19 VFQTSILGAY 16 4 YWLPIMRNPI 7 13 ITPTGHVFQT 7 21 QTSILGAYVI 7 TableXXXIV.V9-HILA-AI-1 Omers.

AMUC1 Each pepide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 11 LPTQPATLGY 21 12 PTQPATLGYV 10 TableXXXV-V1 -HLA-A0201 I Omers-24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 235 ALGVALVLSL 29 44 FILGY!VVGI 28 232 ILVALGVALV 28 243 SLLFILLLRL 28 309 WLAALIVLAV 28 579 MLLMRNIVRV 28 244 LLFJLLLRLV 27 260 VLILGVLGVL 27 433 GLIQRSVFNL 27 508 ILTLVQIARV 27 580 LLMRNIVRWV 27 598 LLFFGKLLW 27 48 YIWVGIVAWL 26 94 CILSSNIISV 28 239 ALVLSLLFIL 26 241 VLSLLFILLL 26 251 RLVAGPLVLV 26 321 AILLLMLIFIL 26 Tab IeXXXV-VI -HLA-A0201l0mers-24P4C12 Each peptidle is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 10 amino acids, and the end position for eactr peptide is the start posibcn plus nine.

Pos 1234567890 score 441 NLQIYGVLGL 26 502 LAFGALILTL 26 517 VILEYIDHKL 26 603 KLLWVGGVGV 26 604 LLVVGGVGVL 26 45 ILGYIWVGIV 25 252 LVAGPLVLVL 25 304 SVE1WLAAL 25 312 ALl VLAVLEA 25 318 VLEAILLLML 25 486 PLISAFIRTIL 25 657 SVFGMCVDTL 25 665 TLFLCFLEDL 25 243 LLLRLVAGPL 24 259 LVLILGVLGV 24 310 LAALIVLAVL 24 339 ALLKEASKAV 24 597 LLLFFGKLLV 24 41 FLLFILGYIV 23 42 LLFILGYIWV 23 56 WLYGDPRQVL 23 231 WILVALGVAL 23 249 LLRLVAGPLV 23 .256 PLVLVLILGV 23 313 LIVLAVLEAI 23 315 VLAVLEAILIL 23 438 SVFNLQIYGV 23 459 ALGQCVLAGA 23 686 YMSKSLLKIL 23 99 NIISVAENGL 22 257 LVLVLILGVL 22 354 TMFYP!LVTFV 22 413 HLVNSSCPGL 22 449 GLFWTLNWVL 22 506 ALl LTL VQIA 22 510 TLVQIARVIL 22 513 OIARVILEYI 22 581 LMRNIVRVW 22 565 IVRV\PLDKV 22 590 VLDKVTDLLL 22 1S9 1TIQQGISGL 21 247 ILLLRLVAGP 21 253 VAGPLVLVLI 21 316 LAVLEA7ILLL 21 501 SLAFGALILT 21 505 GALILTLVQI 21 641 IMTSILGAWV 21 86 LLYFN!FSCI 20 TableX)O(V-VI-HLA-A0201l10mers-24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptde is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 95 ILSSNIISVA 191 ALPGITNDTT 238 VALVLSLLFI 261 LILGVLGVLA 314 IVLAVLEAIL 325 LMLIF!LRQRI 329 FLRQRIRIAI 350 QMMSTMFYPL 358 PAITFVLLLl 368 ClAY WAMTAL 393 NISSPGCEKV 554 FLNRNAYIMI 596 DLLLFFGKLL 645 ILGAYVIASG 649 YVIASGFFSV 34 DVlCC VLFLL 19 64 VLYPRNSTGA 19 85 YLLYFNIFSC 19 186 NVTPPALPGI 19 233 LVALGVALVL 19 264 GVLGVLAYGI 19 317 AVLEA!LLLM 19 327 LIFLR RIRI 19 335 RIAIALLKEA 19 351 MMSTMFYPLV 19 357 YPLVTFVLLL 19 363 VLLLICIAYW 19 364 LLLIC lAY WA 19 365 ILLICIAYWAM 19 380 AISGQPQYVL 19 457 VLALG-QCVLA 19 536 CIMCCFKCCL 19 588 VVVLDKVTDL 19 633 HLNYYWLPIM 19 644 SILGAYVIAS 19 39 VLFLLFILGY 18 157 TVITSLQOEL 18 203 QGISGLIDSL 18 208 LIDSLNARDI 18 240 LVLSLLFILL 18 246 FILLLRLVAG 18 262 ILGVLGVLAY 18 281 RVLRDKGASI 18 322 ILLLMLIFLR 18 332 QRIRIAIALL 18 360 '/TFVLLLICI 18 388 VLWASNISSP 18 448 LGLFWTLNWV 18 WO 2004/050828 WO 204/00828PCTIUS2002/038264 TableXXXV-VI -HLA-AC201l0mers-24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start Position plus nine.

Pos 1234567890 score 493 RTLRYHTGSL 18 525 KLRGV-QNPVA 18 589 WLDKVTDLL 18 616 FFFSGRIPGL 18 662 CVDTLFLCFL 18 685 YYMSKSLLKI 18 130 NEFSQTVGEV 17 143 KNRN4FCLPGV 17 148 CLPGVPWNMT 17 170 FLLPSAPALG 17 211 SLNARDISVI{ 17 227 QSWVYW!iLVAL 17 254 AGPILVLEVLIL 17 296 TTNLSAYQSV 17 324 LLMLIFLRQR 17 373 AMTALYLATS 17 481 DIPTFPLISA 17 546 WCLEKFIKFL 17 563 IAIYGKNFCV 17 582 MRNIVRVWVL 17 LFLLF!LGYI 16 108 LQCPTPQVCV 16 118 SSCPEDPWTV 16 169 SIFLILPSAPAL 16 200 TIQQG!SGLI 16 207 GLIDSILNARD 16 212 ILrNARDISVIKI 16 236 LGVALVLSLL 16 292 QLGFTTfNLSA 16 307 ETWLAALIVL 16 319 LEAIL!LLMLI 16 337 AIALLIKEASK 16 366 ILICIAYWAMT 16 405 NI-SCNPIAHL 16 451 FWTLNWVLAL 16 456 WVLALGQCVL 16 458 LALGQCVLAG 16 503 AFG3AL!LTLV 16 509 LTLVQIARVI 16 637 YWVLPIMTSIL 16 33 TDVICCVLFL 15 36 ICC\ILFLLFI 15 NIFSCILSSN 15 161 SLQQELCPSF 15 225 FAQSWYWILV 15 234 VALGVAILVILS 15 250 LRLVAGPLVL 15 284 RDKGASISQL 15 TableXXXV.V1.H1-11-AA020I I Cmers-24P4C1 2 Each peptide is a portion of SEQ ID NO: 3; eachi start position is specified, the length of peptide is 10 amino acids, and the end position for each peptidle is the start positon plus nine.

Pos 1234567890 score 323 LLLMLIFLRQ 15 340 LLKEASKAVG 15 378 YLATSGQPQY 15 379 LATSG- PQYV/ 15 430 SSKGL!QRSV 15 464 VILAGAIFASIFY 16 498 HTGSLAFGAL 15 520 EYIDHKLRGV 15 539 CCFKCCLWCL 15 601 FGKLLW'GGV 15 590 SLLKI!LGKKN 15 26 PIKN4SCTDV 14 30 RSCTDVICCV 14 37 CCVLFLLFIL 14 102 SVAENGLQCP 14 149 LPGVPWNMTV 14 153 PWNMTVITSL 14 162 LQQELCPSFL 14 165 ELCPSELLPS 14 171 LLPSAPALGR 14 177 ALGRCFPWTN 14 220 KIFEDFAQSW 14 273 IYYCWEEYRV 14 338 IALLKEASKA 14 353 STMFYPLVTF 14 370 AYWAMTALYL 14 395 SSPGCEKVPI 14 416 NSSCPGLMCV 14 445 YGVLGLFWTL 14 483 PTFPLISAFI 14 5CC GSLAFGALIL 14 571 CVSAKNAFML 14 577 AFMLLMRNIV 14 5S6 TDLLLFFGKL 14 6C6 VVGGVGVILSF 14 639 LPIMTSILGA 14 680 SLDRPYYMSK 14 693 IKILGIKKNEAP 14 694 ILGKKNEAPP 14 Tab~eXXXV-V3-HLA-A0201- 110mers.24P24C12 Each peptide is a portion of SEQ 1I) NO: 7; each start position is specified, the length of peptidle is 10 amino acids, and the end position for each peptide is the start pos tion plus nine.

Pos 1234567890 score 9 NITPPALPGI 23 10 ITPPALPGIT 12 Tab~eXXX(V-VS-HLA.A0201.

10Omers-24M11 2 Each peptidle is a portion of SEQ ID NO: 11; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 5 AILLLVLIFL 26 1 AVLEA!LLLV 26 2 VLEAILLLVL 3 LEAILLVLI 18 6 ILLILVILIFILR 18 8 LLVLIFLRQR 16 9 LVILIFILRQRI 16 7 LLLVL71FLRQ 10 VLIFLRQRIR 12 TableXXXV-V6-HLA-A0201 110mers-2441 Each peptide is a portion of SEQ 10 NO: 13; each start position is specified, the Length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 7 GLIPIRSVIFNIL 29 4 SSKGLIPRSV TableXXXV-V7-HLA-AU201Il10mers-24P4C112 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptidle is the start position plus nine.

Pos 1234567890 score 1 QSWYWILVAV 4 2 SWYWILVLAVG 4 4 YWILVAVGQM 3 5 WILVAVGQMM 2 6 IL VA VGMMS 2 8 VAVGQMVMSTMV 2 9 AVGQMM TMF 2 TableXXXV-V8-HLA-A020Ill10mers-24P4C12 WO 2004/050828 WO 204/00828PCT/US2002/038264 Each peptidle is a portion of SEQ ID NO: 17; each start position is specified, the length of pepide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Poe 1234567890 score 4 YWLPIMRNPI 15 WLPIMRNPIT 15 18 HVFOTSILGA 15 7 PIMRNPITPT 14 13 ITPTGHVFQT 14 8 IMRNPITPTG 13 21 QTSILGAYVI 13 FQTSILGAYV 12 PTGHVFQTSI 11 RNPITPTGHV 10 16 TGHVFQTSIL 10 12 PITPTGHVFQ 8 TableXX)CV-V9-HLA-A0201 I Oimers-24P4C112 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, tihe length of peptde is amino acids, and the end position for each peptide is the start position plus nine.

Poe 1234567890 score 9 YPLPTQPATL 20 2 YWAMTALYPL 19 7 ALYPLPTQPA 19 12 PTQPATLGYV 17 16 ATLGYVLWAS 15 4 AMTALYPLPT 14 MTALYPLPTQ 13 17 TLGYVLWASN 13 13 TQPATLGYVL 11 18 LGYVLWASNI 11 PATLGYVLWA 9 TableXXXVI.Vi .HLA-A0203.

1imers-24P4C12 Each peptidle is a portion of SEQ ID NO: 3; each start position is specif ed, the length of peptide is 10 amino acids, and the end position for each peptidle is the start position plus nine.

Pos 1234567890 score 303 Q SVQETWLMA 19 168 PSFLLPSAPA 18 330 LRQRIRIAIA 18 459 ALGQCVLAGA 18 461 dQCVLAGAFA 18 304 SVQETWLAAL 17 3 GKQRDEEDEA 10 TableXXXVI.V1-HLA-A0203l0mers-24P4C1 2 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptidle is 10 amino acids, and the end position for each peptde is the start position plus nine.

Pos 1234567890 score 46 LGYIWGIVA 10 64 VLYPRNSTGA 10 95 ILSSNIISVA 10 166 LCPSFLLPSA 10 182 FPWTNVTFPA 10 205 ISGLIDS-LNA 10 217 ISVKIFEDFA 10 226 AQSWYWILVA 10 230 YWVILVALGVA 10 245 -LFILLLRLVA 10 261 L!LGVLGVLA 10 279 EYRVLRDKGA 10 292 QLGFTTNLSA 10 302 YQSVQETWLA 10 308 TWLMLIVLA 10 312 ALIVLAVLEA 10 328 IFLRQRIRIA 10 335 R!AIALLKEA 10 338 IALLKEASKA 10 361 TFVLLL!ICIA 10 364 LLLICIAYWA 10 367 1ICIAYWAMTA 10 371 YWAMTALYLA 10 382 SGQ)PQYVLWA 10 403 P!INTSCNPTA 10 460 LFWTrLNWVLA 10 457 VLALGQCVLA 10 466 AGAFASFYWA 10 481 DIPTFPLISA 10 494 TLRYHTGSLA 10 497 YHTGSLAFGA 10 506 ALILTLVQIA 10 525 KLRGVQNPVA 10 550 KFIKFLNRNA 10 555 LNNYIMIA 10 565 IYGKNFCVSA 10 568 KNFCVSA-KNA 639 LPIMTSILGA 643 TSILGAYVIA 10 692 LjILGKK-NEA 10 4 KQRDEDDEAY 9 47 GYIWVGIVAW 9 65 LYPRNSTGAY 9 96 LSSNIISVAE 9 167 CPSFLLEPSAP 9 169 SFLLPSAPAL 9 183 PwTNVTPPAL 9 206 SGLIDSLNAR 9 TableXXXVI.VI-HLA*A0203* 10Omers-24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 218 SVLKIFEDFAQ 9 227 QSWYWILVAL 9 231 WILVALGVAL 9 246 FILLLRLVAG 9 262 ILGVLGVLAY 9 280 YRVLRDKGAS 9 293 LGFTTNLSAY 9 309 WLMLIVLAV 9 313 LIVLAVLEAI 9 329 FLRQRIRIAI 9 331 RQRIRIAIAL 9 336 IAIALLKEAS 9 339 AILLKEASKAV 9 362 FVLLLICIAY 9 365 LLICIAYWANI 9 368 CIAY WAMTAL 9 372 WAMTALYLAT 9 383 GQPQYVL WAS 9 404 INTSCNPTAH 9 451 FWTLNVLAL 9 458 LALGQCVLAG 9 460 L QCVLAGAF 9 462 QCVLAGAFAS 9 467 GAFASFYWAF 9 482 IPTFPLISAF 9 495 LRYHTGSI.AF 9 498 IHTGSLAFGAL 9 507 ILTLV -IAR 9 525 LRGVQNPVAR 9 551 FIKFLNRNAY 9 556 NRNAYIMIAI 9 566 YGKNFCVSAK 9 569 NFCVSAKNAF 9 640 P!MTSILGAY 9 644 SILGAY7VIAS 9 693 K!LGKKNEAP 9 TableXXX(VI-V3-HLA.AO203-10mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptidle is the start position plus nine.

Pos 1234567B90 score 5 FPWTNITPPA 6 PWTNITPPAL 9 WO 2004/050828 WO 204/00828PCTIUS2002/038264 TableXXXVI.V3-HLA-A0203-l0mers- 24P4C12 Each peptidle is a portion of SEQ ID NO: 7; each start positon is specified, the length of pepide is 10 amino acids, and the end position for each peptidle is the start position plus nine.

Pos 1234567890 score 7 WTNITPPALP 8 TableXXXVI-V5-HLA-A0203- 1 Omers-24F12 Pos 1234567690 score NoResultsFound.

TableXXXVI-V6-IILA-A0203- I Omers-24PACI 2 Pos 1234567890 score NoResultsFound.

TableXXX(VI-V7-4LA.AO2O3-1 Omers- 24P4C12 Each peptidle is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 1 QSWYWIVAV 9 2 SWVYWILVAVG 8 TableXXXVI.V8*IL111A.A0203-10mers.

24P4C1 2 Each pepide is a portion of SEQ ID NO: 17; each start positi is specified, the length of peptidle is 10 amino acids, and the end position for each peptidle is the start position plus nine.

Pos 12345671390 score 18 HVFQISILGA 10 19 VFQTSILGAY 9 FQTSILGAYV 8 TableXXXVI-V9.HLA-A0203-l0mers- 24P4C12 Each peptide is a portion of SEQ ID .NO: 19; each start position is specified, the length of peptide is 10 amino acids, and the end position for each pepfide is the start position plus nine.

Pos 1234567890 score 7 ALYPLPTQPA 10 ,PATL1GYVLWA 10 8 LYPLPT2PAT 9 16 ATLGYVILWAS 9 YPLPTQPATL 17 TLGYVLWVASN TableXXXVII.VI .HLA-A3-l0mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of pepide is 10 amino acids, and the end position for each peptidle is the start position plus nine.

Pos 1234567890 score 333 RIRIAIALLK 32 211 SLNARIDISVIK 30 .337 AIALLKEASK 28 516 RVILEYIDH-K 28 281 RVERDQASI 27 680 SLDRPYYMSK 27 464 V/LAGAFASIFY 25 584 NIVRVWVLDK 24 621 RIPGLIGKDFK 24 49 IVVGIVAWLY 23 463 CVLAGAJASF 23 233 LVALGVALVL 22 262 ILGVLGVLAY 22 376 ALYLATSGQP 22 443 QIYGVLGLFW 22 525 KLRGV NPVA 22 587 RWVVLDKVTD 22 603 KLLVVGGVGV 22.

56 WILYGIDPRQVIL 21 63 QVLYPRNSTG 21 177 ALGRCFPWVTN 21 564 AlYGKNFCVS 21 608 W\GGVGVLSIF 21 39 VLFLLFILGY 20 53 IVAWLYGD)PR 20 171 LLPSAPALGR 20 251 RLVAGPLVLV 20 252 LVAGPLV[VL 20 282 VLRDKGASIS 20 362 FV'LLLICI01AY 20 378 YLATSGQPQY 20 544 CLCLEKFIK 20 650 VIASGFFSVF 20 95 ILSSNIISVA 19 170 FLLPSAPALG 19 191 ALPGITNDTT 19 237 GVALVLSLLF 19 248 LLLRLVAGPL 19 260 VLI1LGVLGVL 19 261 LILGVLGVLA 19 298 NL AYISVQE 19 312 ALl VLAVLEA 19 314 IVLAVLEAIL 19 317 AVLEAILLLM 19 322 ILLLMLIFLR 19 TableXXXVII-Vi 24P4CI2 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position~ plus nine.

Pos 1234567890 score 340 LLKEASKAVG 19 347 AVGQMMSTMF 19 494 TLRYHTGSLA 19 605 LWV G-VIS 19 618 FSGRIPGLGK 19 645 lGAY VIASG 19 673 IJLERNNGSLID 19 6 RDEDiEAYGK 18 64 WLYPRSTGA 18 134 QTvGEVFYrK 18 231 WILVALGVAL 18 235 ALGVALVLSL 18 247 1ILL LRLVA G P 18 258 VLVLILGVLG 18 324 LLMLIFLRQR 18 456 WVLALgCVL 18 532 PVARCIMCCF 18 72 GAYCGMGENK 17 86 LLYFNIFSCI 17 161 SQQIECPSF 17 207 GLIDSLNARD 17 220 KIFEDFAQSW 17 232 ILVALGVALV 17 249 LLRLVAGPLV 17 257 LVLVLILGVL 17 264 GVLGVLaYGI 17 265 VLGVLAYGIY 17 292 QLGFTTNLSA 17 309 WLALIVLAV 17 326 MLIFLEgRIR 17 364 LLJIOIAYWA 17 38B VLWASNISSP 17 392 SiN!SSPGCEK 17 486 PL!SAFIRTIL 17 506 ALILTLVQIA 17 551 FIKFLNRNAY 17 580 LLMRNVRWV 17 598 LLFFGKLLWV 17 612 VLSFFFFSGR 17 624 GLGKDFKSPH 17 649 YVIASGFFSV 17 657 SVFGMCVDTL 17 667 FLCFLEDLER 17 68 PVYMSKSLLK 17 689 KSLLKILGKK 17 9 DDEAYGKPVK 16 44 FILGY-IVVGI 16 126 TVGKNEFSQT 16 165 ELCPSFLLPS 16 WO 2004/050828 WO 204100828PCTIUS2002/038264 TableXXXVI.V-HL-A3I Omers.

24PC1 2 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptidle is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 243 SILFILLLRL 16 246 FILLLRLVAG 16 259 LVLILGVLGV 16 272 GIYYCWEEYR 16 304 SVQETWLAAL 16 318 VEAILLML 16 339 ALLKEASKAV 16 363 V1LL1CIAYW 16 453 TLNWVLALGQ 16 457 VLaLGO-VLA 16 459 ALOQCVLAGA 16 487 LISAFIRTLR 16 508 ILTLVQIARV 16 518 ILEYIDHKLR 16 559 AYIMIAIYGK 16 566 YGKNFCVSAK 16 571 CVSAKNAFML 16 579 MLLMRNIVRV 16 596 DLLLFFGKLL 16 640 PIMISILGAY 16 690 SLLKILGKKN 16 693 KILGKKNEAP 16 VICCVLFLLF 15 41 FLLFILGYIV 15 42 LLFILGYIW 15 107 GLOCfPPVC 15 120 CPEDPWTFVGK 15 180 RCFPWTNVTP 15 323 LLLMLIFLRQ 15 329 FLRQRIRIAI 15 367 IClAY WAMTA 15 369 lAY WAMTALY 15 423 MCGYSSK 15 446 GVLGLFWTLN 15 491 FIRTLRYHTG 15 507 LILTLVQIAR 15 510 TC161ARVIL 15 585 IVRVVVLOKV 15 597 LLLFFGKLLV 15 604 LLW7vGVGVL 15 688 SKSLLKILGK 15 694 ILdKKIAEAPP 15 697 KKNEAPPDNK 15 698 KNEAPPDNKK 15 TableXXXVII-V3-HLA-A3-l0mers- 24P14C1 2 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptidle is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 3 RCFPWVTNITP 11 9 NITPPALPGI 11 B TNITPPALPG 9 10 ITPPALPGIT 7 7 WTNITPPALP 5 TableXXXVII-V5-HLA-A3-1 Oniers* 24P4C1 Each peptide is a portion of SEQ ID NO: 11; each start position is specified, the length ot peptidle is 10 amino acids, and the end position for each peptidle is the start position plus nine.

Pos 1234567890 score 1 AVLEAILLLV 19 2 VLEAILLLVL 19 6 ILLLVLIFLR1 19 a LLVLIFLRQR 1s 10 VL!FLRQRIR 17 7 LLE\VLIFLRQ 15 5 AILLLVLIFL 14 Q LVLIFLRQRI 14 4 EALLLVILIF 11 TableXX)CVIl-VS-HLA-A3-10mors- 24P4C12 Each peptide is a portion of SEQ I D NO: 13; each start position is specified, the length of peptide is 10 amrino acids, and the end position for each peptide is the start position plus nine.

PCs 1234567890 score 7 GL!PRSVFNL 16 5 SKGLIPRSVF 14 1 QGYSSKGLIP 12 6 LI'RSMENLQ 11 9 IPRSVFNLQI 11 6 KGLIPRSVFN 10 4 SSKGUPRSV 7 TableXX(XVll-V7-HLA-A3-l0mers- 24P4C1 2 Each peptidle is a portion of SEQ ID NSO: 15; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 9 AVGQMMSTMF 19 6 ILVAVQMMS 16 5 WILVAVGQMM 14 7 LVAVGQMMST 14 2 SWYiWILVAVG 12 8 VAVGQMMSTM 9 TableX(XVII-V8-HLA-A3-1 Omners- 2424C1 2 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptidle is amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 12 PITPTGHVFQ 11 NPITPTGHVF 14 18 HVFQT-SLGA 13 7 PIMRNTP 12 5 WLPIMRNPIT 11 1 LNYYWLPIMR 8 IMRNPITPTG 21 QTSILGAYVI 9 MRNPITPTGH 9 6 LPIMRNPITP 8 19 VFQTSILGAY 8 24P4C12 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptidle is amino acids, and the end posiflon for each peptide is the start position plus nine.

Pos 1234567890 score 7 ALYPLPTQPA 17 TLGYVLWASN 10 PLPTQPATLG 1l4 9 YFTPTPATL 13 1 AYWVAMTALYP 11 18 LGYVLWYASNI 4 AMTALYPLPT 9 11 LPIQPATLGY 9 13 TQPATLGWYL 9 TableX)O(VIII-VI-HLA-A26-l0mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score WO 2004/050828 WO 204/00828PCT/US2002/038264 rableXXXVlI-Vi .HLA-A26-10mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of pcptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 34 DVICCVLFLL 34 138 EVFYTKNRNF 32 307 ETWLAALIVL 31 657 SVFGMCVDTL 28 199 TTIQQGISGL 26 304 SVQETWLAAL 26 588 VV\/LDKVTDL 26 592 DKVTDLLLFF 25 49 IWVGIVAWVLY 24 606 WVGGVGVLSF 24 157 TVITSLQQEL 23 252 LVAGPLVLVL 23 257 LVLVLILGVL 23 320 EAILLLMLIF 23 628 DFKSPHLNYY 23 79 ENKEJKPYLLY 22 353 STMFYPLVTF 22 362 FVLLLICIAY 22 662 CVDTLFLCFL 22 672 EDLERNNGSL 22 48 YWVGIVAWL 20 198 DTTIQQGISG 20 216 DISVKIFEDF 20 240 LVLSLLFILL 20 293 LGFTTNLSAY 20 640 PIMTSILGAY 20 DEAYGKPVKY 19 39 VLFLLFILGY 19 131 EFSQTVGEVF 19 233 LVALGVALVIL 19 237 GVALVLSLLF 19 347 AVGQMMSTMF 19 438 SVFNLQIYGV 19 463 CVLAGAFASF 19 498 HTGSLAFGAL 19 512 VOIARVILEY 19 520 EYIDHKLRGV 19 571 CVSAKNAFML 19 589 WVLDKVTDLL 19 33 TDVICCVLFL 18 203 QGISGLIDSL 18 314 IVLAVLEAIL 18 456 WVLALGQCVL 18 481 DIPTFPLISA 18 486 PLISAFIRTIL 18 493 RTLRYHTGSL 18 502 LAIGALILTL 18 516 RVILEYIDHK 18 TableXXXVIII-VI-HLA*A2610mers- 24P4C12 Each peptide is a portion of SEQ ID) NO: 3; each start position is specified, the length of peptidle is 10 amino acids, and the end position for each pepfide is tihe start position plus nine.

Pos 1234567890 score 532 PVARCIMCCF 18 549 EKFIKFLNRN 18 609 GVGVLSFFFF 18 99 NIISVAENGL 17 102 SVAENGLQCP 17 156 MTVITSLQQE 17 236 LGVAL\/LSLL 17 260 VLILGVLGVL 17 316 LAVLEAILLL 17 317 AVLEAIL[LM 17 321 AILLLMLIFL 17 360 \/TFVLLLICI 17 442 LQIYGVLGLF 17 596 OLLLFFGKLL 17 604 LLWVGGVGVL 17 616 FFFSGRIPGL 17 664 DTLFLCFLED 17 665 TLFLCFLEDL 17 682 DRPYYMSKSL 17 32 CTDVICCVLF 16 37 CCVLFILiFIL 16 123 IJPWTVGKNEF 16 165 LCPSFLLPS 16 186 NVTPPALPGI 16 224 DFAQSWYWfIL 16 239 ALVLSLLFIL 16 262 ILGVLGVLAY 16 266 LGVLAYGIYY' 16 332 QRIRIAIALL 16 359 LVTFVLLLIC 16 380 ATSGQPQYVL 16 400 EKVPINTSCN 15 405 NTSCNPIAHL 16 424 CVFQGYSSKG 16 433 GLIQRSVFNL 16 539 CCFKCCLWCL 16 503 I VTDLLLFFG 16 TableXXXVIII-V3-HLA.A26-1 Diners- Each 24P4C12 Eahpeptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide Is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 6 PWTNITPPAL 10 9 NITPPALPGI 10 10 ITPPALPGIT 7 WTNITPPA[P 3 RCFPWTNIIP 8 TNITPPALPG 4 CFPVVTNITPP TableXXXVIII-V5-HLA-A26i0mers-24P4CI2 Each peptide is a portion of SEQ D NO: 11; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start Position plus nine.

Poe 1234567890 score 4 EAILLLVLIF 27 1 AVLEAILLLV 17 5 AILLLVLIFL 17 2 VLEAILLLVL 13 TableXXXVIII-V6-HLA-A26l0mers-24P4C1 2 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 7 GLIPRSVFNL 17 10 PRSVFNLQIY 14 5 SKGLIPRSVF TableXXXVIII-V7-HLA-A26l0mers-2434C12 Each peptidle is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is 10 amino adds, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 9 AVGQMMSTMF 19 7 LVAVGQMMST 11 4 YWILVAVGQM TableXXXViIII.VO.-LA-A26- 10mefs-24P4C12 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptde is amino acids, and the end position for each peplide is the start position plus nine.

Poe 1234567890 score WO 2004/050828 WO 204/00828PCTIUS2002/038264

HVFQTSILGA

VFQTSILGAY

NPITPTGHVF

ITPTGHVFQT

TGHVFQTSIL

PTGHVFQTSI

TableXXKVIII-V-HLA-A26-10mers- 24P4C1 2 Each peptide is a portion of SEQ ID NO: 19; each start positon is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 12 PTQFATLGY\/ 14 MTALYPLPTQ 13 16 ATLGYVLWAS 13 2 YWVAMTALYPL 12 11 LPTQPATLGY 12 9 YPLPTQPATL 10 13 TQPATLGYVL 10 PATLGYVLWA 6 TableXXXIX-I-HLA-130702- I Omers-24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptidle is amino aods, and the end position for each peptide is the start position plus nine.

Pas 1234567890 score 357 YPLVTFVLLL 23 478 KPQDIPTFPL 23 683 RPYYMSKSLL 21 182 FPVVTNVIPPA 19 83 KPYLLYFNIF 18 192 LPGITNDTTI 18 4B32 IPTFPLISAF 18 639 LPIMTSILGA 18 149 LPGVPWNMTV 17 252 LVAGPLVLVL 17 380 ATSGQPQYVL 17 402 VPINTSCNPT 17 485 FPLISAFIRT 17 123 DPWTVGKNEF 16 235 ALGVALVLSL 16 254 AGFLVLVLIL 15 370 AYWAMTAILYL 15 659 FGMCVDTLFL 15 33 TDVICCVLFL 14 56 WLYGDPRQVL 14 175 APALGRCFPW 14 233 IA'ALGVALVIL 14 241 VLSLLFILLL 14 TableXXXIX-VI-HLA-B0702l0mers-24P4C112 Each peptidle is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptidle is the start position plus nine.

Pos 1234567890 score 331 RORIRIAIAL 14 405 NTSCNIPTAHL 14 451 FWTLNWVLAL 14 502 LAFGALILTL 14 582 MRNIVRVWVL 14 590 VLDKVTDLLL 14 15 KPVKYDPSFR 13 60 DPRQVLYPRN 13 66 YPRNSTGAYC 13 110 CPTPQVCVSS 13 120 CPEDPWTVGK 13 167 CPSIFLLPSAP 13 172 LPSAPALGIRC 13 226 AOSWYWILVA 1l3 227 QSWYWILVAL 13 231 WILVALGVAL 13 250 LRLVAGPLVL 13 284 RDKGASISQL 13 290 ISQLGFTTNL 13 301 AYQSVQETWL 13 31) LAALIVILAVL 13 314 IVLAVLEAIL 13 315 VLEAILLLML 13 321 AILLLIMLIFL 13 35D QMMSTMFYPL 13 355 MFYPLVTFVL 13 358 FYPLVTFVLL 13 368 CIAYWVAMTAL 13 396 SPGCEKVPIN 13 441 NLQIYGVLGL 13 498 HTGSLAFGAL 13 500 GSLAFGALIL 13 510 TLVQIARVIL 13 525 KLRGVQNPVA 13 571 GVSAKNAFML 13 572 VSAKNAFMLL 13 657 SVFGMCVDTL 13 666 YMSKSLLKIL 13 20 DPSFRGPIKN 12 48 YIVVGIVAWL 12 169 SFILLIPSAPAL 12 183 PWTNVTPPAL 12 189 PPALPGITND 12 239 ALVLSLLFIL 12 243 SLLFILLLRL 12 304 SVQETNLAAL 12 307 ETWVLAALIVL 12 309 WLAALIVILAV 12 TableXXXIJX-Vl-HLA-B0702- I Omers.24P34C12 Each peptidie is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 316 ILAVLEAILLL 12 409 NPTAHLVNSS 12 419 CPGLMCVFQG 12 425 VFQGYSSKGL 12 456 WVLALGQCVL 12 493 RTLRYHTGSL 12 581 LMRNIVRVWV 12 588 WVVLDKVTDL 12 604 LLWGGVGVL 12 606 WVGGVGVLSF 12 622 IPGLGKDFKS 12 637 YWLPIMTSIL 12 662 CVDTLFLCFL 12 701 APPDNKKRKK 12 18 KYDPSFRGPI l1 26 GPIKNRSCTD 11 31 SCTDVICCVL 11 44 FILGYIVVGI 11 77 MGENKDKPYL 11 78 GENKDKPYLL 11 140 FYTKNRNFCL 11 152 VPWNMTVITS 11 153 PWNMTVITSL 11 162 LQQELCPSFL 11 188 TPPALPGITN 11 224 DFAQSWYWIL 11 236 LGVALVLSLL 11 240 LVILSILLFILL 11 248 LLLRLVAGPL 11 257 LVLVLILGVL 11 260 VLILGVLGVL 11 274 YYCWEEYRVL 11 312 ALl VLAVLEA 11 315 VLAVLEAILL 11 332 QRIRIAIALL 11 384 QPQYVLWASN 11 395 SSPGCEKVPI 11 413 HLVNSSCPGL 11 433 GLIQRSVFNL 11 435 lQRSVFNLQI 11 439 VFNLQIYGVL 11 445 YGVLGLFWTL 11 449 GLFWrLNWVL, 11 503 AFGALILTLV 11 531 NPV'ARCIMCC 11 536 CIMCCFKCCL 11 539 CCFKCCLWCL 11 546 WCLEKFIKFL I11 WO 2004/050828 WO 204/00828PCT/US2002/038264 Tab~eXXXIX.VI.HLA-B0702- 10mers-24P4C12 Each peptide is a portion of SEQ NO: 3; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 565 IYGKNFCVSA 11 589 WLDKVTDLL 11 595 TDLLLFFGKL 11 616 FFFSGRIPGL 11 625 LGKDFKSP-L 11 630 KSPHLNYYWL 11 672 EDLERNNGSL 11 TableXXXIX.V3-HLA-B0702.

10mers-24P4C12 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start Position plus nine.

Pos 1234567890 score FPWTNITPPA 19 6 PWTNITPPAL 12 1 LGRCFPWTNI 9 TableXXXlX.V5-HLA-BO7O2- 10mers-2434C12 Each peptide is a portion of SEQ ID NO: 11; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 2 VLEAILLLVL 14 AILLLVLIFL 13 1 AVLEAILLLV 10 4 EAILLLVLIF 10 3 LEAILLLVLI 9 9 LVLIFLRQRI 7 TableXXXIX.V6-HLA-B0702- 10rmers-24P4C12 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 9 IPRSVFNLQI 21 7 GLIPRSVFNL 12 TableXXXIX.7.HL11A-B30702lt0mers-24P4Cl2 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1 234567690 score 9 AVGQMMSTMF 10 1 QSWYWILVAV 9 8 VAVGQMMSTM 8 4 YWILVAVGQM 7 7 LVAVGQMMST 7 WVILVAVGQMM~ 6 TableXXXIX*V8-HLA-80702- 10mers;.24P4C12 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptide is 10 amino acids, and the end position for each pepide is the start position plus nine.

Pos 1234567890 score 11 NPITPTGHVF 17 14 TPTGHVFQTS 13 16 TGHVFQTSIL 11 6 LPIMRNPITP 10 4 YWVLPIMRNPI 9 7 PIMRt'PITPT 9 21 QISILGAYVI 9 10 RNJPITPTGHV 8 13 ITPTGHVFQT 8 15 PTGHVFQTSI 8 18 HVFQTSILGA 8 TableXXXIX.V9-HLA.B0702i Omers-24P34C1 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptidle is the start position plus nine.

Pos 1234567890 score 9 YPLPTQPATL 22 11 LPTQPATLGY 13 14 QPA.TLGYVLW 13 2 YWAMTALYPL 12 4 AMTALYPLPU 12 13 TQPATLGYVL 12 7 ALYPLPTQPA 11 TableXLMV-HLA-B308-1 Omers- 24P4CI2 Pos 1234567890 score NoResultsFound.

TableXL-V3-I'LA-B08-1 Diners- 24P4C12 Pos 1234567890 score NoResultsFound.

TableXL-V5-A-B308-l0mers- 24P4C12 Pos 1234567890 score NoResultsFound.

TableXIL-V6HILA-B308-l10mers- 24P34C12 Pos 1234567890 score NoResultsFound.

TableXL-V7-HLA-B08-I0mers- S24P4C12 Pos 1234567890 score NoResultsFound.

TableXIL-VS-HL-A-BOB-l0mers- 24P4C12 Pos 1234567890 score NoResultsFound.

TableXL-Vg-HLA-B08- I Omers-24P4C1 2 Pos 1234567890 score NoResultsFound.

TableXILI-VI-HL-A-B1510-10mors- 24P4C12 Pos 12345676390 score NoResultsFound.

TableXLI-V3-HLA-BI 151 0-1 Omars- 24P4C12 Pos 1234567890 score NoResultsFound.

TableXLI.V5-HLA-BI 510-1 Oiners- 24P4C12 Pos 1234567890 score NoResultsFound.

TableXI-V6-HL11A.BI3 510- 10mers-24P4C12 Pos 1234567890 score NoResultsFound.

TableXI.47.HILA-B51510-110mers- 24P4C12 WO 2004/050828 WO 204/00828PCT/US2002/038264 Pos 1234567890 score TableXLIIl-V3-HLA-B2709-1 Omers- NoResultsFoufld. 24P4C12 Pos 1234567890 score TableXlI.Y8-HL-A-BI 510-11Omers. NoResultsFound.

24P4C12 Pos 1234567890 score TableX~lI-V5-HLIA-132709-11Omers- NoResultsFound. 24P4CI2 Pos 1234567890 score TableXlI.V49-1H11A.BI15IO-11Omers- NoResultsFound.

24P4C1 2 Pos 1234567890 score TableXLIIIN86-HLA-82709-1 Omers- NoResultsFound. 24P4C12 Pos 1234567890 score NoResultsFound.

rableXLlI-VI-HLA-B2705-10mers- 24P4C12 TableXLIIl-V-HLA-B2709-10mers-24P4C1 2 Pos 1234567890 score Pos 1234567890 score NoResultsFound. NoResultsFound.

TableXLII-V3-HLA-B2705-1 Omers-24P4C12 TableXLIII.V8-HLA-B2709-l0mers- Pos 1234567890 score 24P4C12 NoResultsFound. Pos 1234567890 score NoResultsFound.

TableXLII-V5-HLA-B2705-1 Omers- 24P4C12 Pos 1234567890 score NoResultsFound.

TableXLII-V6-I-LA-B2705-l Omers- 24P4C12 Pos 1234567890 score NoResultsFound.

TableXLII-V7-HLA-B27Cl5-1 Omers- 24P4C12 Pos 1234567890 score NoResultsFound.

TableXLII.V8-HLA-82705-1 Diners.

24P4C12 Pos 1234567890 score NoResultsFound.

TableXILlI-V9-LA-B2705-l10mers- 24P4C12 Pos 1234567890 score NoResultsFound.

TableXLIII-VI-HLA-B270910mers- 24P4C12 Pos 1234567890 score NoResultsFound.

TableXILlII-3-HLA-B2709-10mers- 24P4CI2 Pos 1234567890 score TableXIU*V19-HILA-132709-1 Oniers- 24P4C12 Pos 1234567890 score NoResultsFound.

TableXLIV-Vi-HLA-B4402l0mers-24P4C12 Each pepide is a portion of SEQ ID NO: 3 each start position is specified, tihe length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 10 DEAYGKPVKY 23 78 GENKDKPYLL 22 222 FEDFAQSWYW 21 319 LEAILLILMILI 20 47 GYIVVGIVAW 19 332 QRIRIAIALL 18 486 PLISAFIRIL 18 502 LAFGAILILTIL 18 620 GRIPGLGKDF 18 39 VILFILLFILGY 17 241 VLSLLFILLL 17 254 AGPLVLVLIL 17 320 EAILLILMLIF 17 321 AILLILMLIFL 17 476 FHKPQDIPTF 17 512 VQIARVILEY 17 699 NEAPPDNKKR 17 121 PEDPWTVGKN 16 169 SFLLPSAPAL 16 TableXLIV-Vi-HLA-B4402.

l0mers-24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, tie length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 199 TTIQQGISGL 16 203 QGISGLIDSL 16 260 VLILGVLGVL 16 293 LGFTTNLSAY 16 307 ETWL.AALIVL 16 316 LAVLEAILLL 16 380 ATSGQPQYVL 16 546 WVCLEKFIKFL 16 657 SVFGMCVDTL 16 34 C)VICCVLFLL 65 ILYPIRNSTGAY 79 ENKDKPYLLY 99 NIISVAENGL 104 AENGLQCPTP 138 EVFYTKNRNF 213 NARDISVIKIF 235 ALGVALVLSL 239 ALVLSLLFIL 278 EEYRVLRDKG 284 RDKGASISQL 353 STMFYPLVTF 355 MFYPLVTFVL 356 FYPLVTFVLL 362 FVLLLICIAY 363 VLLLIGIAYW 370 AYWAMTALYL 417 SSCPGLMCVF 442 LQIYGVLGLF 451 FWTLNVLAL 482 IPTEPLISAF 561 IMIAIYGKNF 596 DLLLFFGKLL 616 FFFSGRIPGL 637 YWILPIMTSIL 640 PIMTSILGAY 4 KQRDEDDEAY 14.

18 KYDPSFRGPI 14 80 NKDKPYLLYF 14 83 KPYLLYFNIF 14 130 NEFSQTVGEV 14 131 EFSQTVGEVF 14 157 TVITSLQQEL 14 164 QELOPSFLLP 14 173 PSAPALGRCIF 14 175 APALGRCFPW 14 183 FWTNVTPPAL 14 220 KIFEDFAQSW 14 227 QSWYWILVAL 14 WO 2004/050828 WO 204/00828PCT/US2002/038264 TableXLIV-V1.HLA-B4402- I0mers-24P4C112 Each peptide is a port on of SEQ 1I) NO: 3; each start position is specified, the length of peptidle is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1 234567890 score 231 WILVALGVAIL 14 233 LVALGVALVL 14 240 LVLSILLFILL 14 243 SLLFILLLRL 14 250 LRLVAGPLVL 14 252 LVAGPLVLVL 14 253 VAGPILVLVILI 14 262 ILGVLGVLAY 14 304 SVQETWLAAL 14 331 RQRIRIAIAL 14 357 YPLVTFVLLL 14 431 SKGLIQRSVF 14 433 GLIQRSVFNL 14 467 GAFASFYWfAF 14 542 KCCLWCLEKF 14 545 LWCLEKFIKF 14 551 FIKFLNRNAY 14 569 NFCVSAKNAF 14 589 VVLDKVTDLL 14 595 TDLLLFFGKL 14 627 KDFKSPI-LNY 14 629 FKSPI-LNYYW 14 665 TLFLCFLEDL 14 686 YMSKSLLKIL 14 7 DEDDEAYGKIP 13 31 SCTDVICCVL 13 32 CTDVICCVLF 13 VICCVLFLLF 13 49 IWGIVAWLY 13 56 WLYGDPRQVL 13 57 LYGDPRCVLY 13 87 LYFNIFSCIL 13 145 RNFCLPGVPW 13 153 PWNMTVITSL 13 186 NVTPPALPGI 13 237 GVALVLSLLF 13 248 LLLRLVAGPL 13 257 LVLVLILGVL 13 271 YGIYYCWEEY 13 301 AYQSVQETWL 13 310 LAALIVLAVL 13 315 VLAVLEAILL 13 327 LIFLRQRIRI 13 342 KEASKAVGQM 13 347 AVGQMMSTMF 13 405 NTSCNPTAHL 13 425 VFQGYSSKGL 13 441 NLQIYGV1..GL 13 TabeXLIV.Vl-HLA-B4402i omers-2441 2 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of pepfide is 10 amino acids, and the end position for each peptide is the start position pius nine.

Pus 1234567890 score 445 YGVLGLFWTL 13 447 VLGLFWTLNW 13 449 GLFWTLNWVL 13 460 LGQCVLAGAF 13 478 KPQDIPTFPL 13 483 PTFPLISAFI 13 493 RTLRYHTGSL 13 495 LRYHTGSLAF 13 498 HTGSLAFGAL 13 500 GSLAFGAILIL 13 517 VILEYIDHKL 13 539 CCFKCCLWCL 13 557 RNAYIMIAIY 13 582 MRNIVRWVL 13 590 VLDKVTDLLL 13 591 LDKVTDLLLF 13 592 DKVTDLLLFF- 13 606 WVGGVGVLSF 13 659 FGMCVDTLFL 13 61 MCVDTLFLCF 13 662 CVDTLFLCFL 13 671 LEDLERNNGS 13 672 EOILERNNGSL 13 682 DRPYYMSKSL 13 33 TDVICCVLFL 12 37 CCVLFLLFIL 12 44 FILGYIVVGI 12 76 GMGENKDKPY 12 123 DPWTVGKNEF 12 132 FSQTVGEVFY 12 150 PG VP WNMTVI 12 163 QQELCPSFLL 12 216 [)ISVKIFEDF 12 223 EDFAQSWYWI 12 236 LGVALVLSLL 12 266 LGVLAYGIYY 12 274 YYCWVEEYRVL 12 277 WEEYRVLRDK 12 285 KGASlSQLGF 12 290 ISQLGFTTNL 12 300 SAYQSVQETW 12 306 GET WLAALIV 12 313 LIVLAVILEAI 12 318 VLEAILLLML 12 329 FLRQRIRIAI 12 350 QMMSTMFYPL 12 358 PLVTFVLLLI 12 360 VTFVLLLICI 12 TableXLIV-VI-HLA-B4402l0mers-24P4C12 Each peptide is a portion of SEQ I D NO: 3; each start position Is specified, the length of peptidle is amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 368 ClAY WAMVTAL 12 369 lAY WAMTALY 12 378 YLATSGQPQY 12 381 TSGQPQYVLW 12 395 SSPGCEKVPI 12 420 PGLMC\IFQGY 12 436 QRSVFNLQIY 12 439 VFNLQIYGVL 12 443 QIYGVLGLFW 12 456 WVVLALGQCVL 12 463 CVLAGAIFASIF 12 464 VILAGAIFASFY 12 488 ISAFIRTILRY 12 505 GALILTLVQI 12 509 LTLVQIARVI 12 510 TLVQIARVIL 12 548 LEKFIKFLNR 12 556 NRNAYIMIAI 12 571 CVSAKNAFML 12 572 VSAKNAFMLL 12 576 NAFMLLMRMI 12 588 VVVLDKVTDL 12 604 LLWVGGVGVL 12 628 DFKSPHLNYY 12 630 KSPHLNYYWL 12 650 \IIASGFFSVF 12 674 LERNNGSILIDR 12 676 IRNNGSLIDRPY 12 677 NNGSLDRPYY 12 685 YYMSKSLLKI 12 14 GKPVKYDPSF 11 27 lKNRSCTDVI 11 40 LFLLFILGYI 11 48 YIVVGIVAWVL 11 77 MGENKDKPYL 11 116 CVSSCPEDPW 11 137 GEVFYTKNRN 11 161 SLQQELCPSF 11 162 LQQELCPSFL 11 208 LIDSLNARDI 11 212 LNARDISVKI 11 2211 IFEDFAQStNY 11 238 VALVLSLLFI 11 264 GVLGVLAYGI 11 305 VQETWLAALI 11 314 IVILAVILEAIL 11 348 VGQMMSTMFY 11 413 HLVNSSCPGL 11 WO 2004/050828 WO 2004050828PCTIUS2002/038264f TableXLlV.VI-HLA-B4402l0mers-24P4Cl 2 Each pelptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1 234567890 score 479 PODIPTEPLI 11 499 TGSLAFGALI 11 W1 LEYIDHKLRG 11 528 GVQNPVARCI 11 532 PVARCIMCCF 11 536 CIMCCFKCCL 11 537 IMCCFKCCLW 11 543 CCLWCLEKFI 11 552 IKFLNRNAYI 11 607 VGGVGVLSFF 11 608 GGVGVLSFFF 11 609 GVGVLSFFFF 11 625 LGKDFKSPHL 11 632 PHLNYYWLPI 11 642 MTSILGAYVI 11 646 LGAYVIASGF 11 658 VFGMCVDTLF 11 683 RPYYMSKSLL 11 TableXLIV-V3-HLA-B4402-1 Omers- 24P4C12 Each peptide is a portion of SEQ ID NO: 7: each start positlion is specified, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 6 PWTNITPPAL 14 9 NITPPALPGI 13 1 LGRCFPWTNI 8 3 RCFPKTNITP 7 8 TNIIPPALPG 6 TableXI-V5.HLA.B4402-i0mers.

24P4C1 2 Each peptide is a portion of SEQ ID NO: 11;- each start position is specfied, the length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 3 LEAIL.LLVLI 21 4 EAILLLVLIF 18 AILLLVLIFL 17 2 .VLEAILLLVL 13 9 LVLIFLRQRI 1C TableXLIV*V6-HLA-B4402ionmers-24PKC12 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, thne length of peptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 7 GLIPRSVFNL 17 5 SKGLIPRSVF 14 10 PRSVFNLQIY 12 9 IPRSVFNLQI 10 TableXLIV.V7-HLA*B4402* lonmers-24P4C12 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus nine.

Pos 1234567890 score 9 AVGQMMSTMF 13 4 YWILVAVG0QI 6 TableXILIV.-HL-A.B34402- 1 Dmers-24P4C1 2 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of pelptide is 10 amino acids, and the end position for each peptide is the start position plus nine.

Pas 1234567890 score 11 NPITPIGHVF 17 4 YWLPIMRNPI 14 19 VFQTSILGAY 14 16 TGHVFQTSIL 11 21 QTSILGAYVI 11 15 PTGHVFQTSI 8 TableX(LIV.V9-HLA-B4402.

l0mers-24P34C12 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is 10 amino acids, and the end positi on for each peptide is the start position plus nine.

Pos 1234567890 score 9 YPLPTQPATL 16 14 QPATLGYVLW 13 11 LPTQPATLGY 12 13 TQPATLGYVL 12 2 YWAMTALYPIL 11 18 LGYVLWASNI 9 16 ATLGYVLWAS 8 7 ALYPLPTQPA 7 TableXLV-VI-HLA-B5101 -1 niers- 24P4IC1 2 Pos 1234567890 score NoResultsFound.

TabIeXLV-V3-HLA-B51 01 .1 Omners- 24P4C12 Pos 1234567890 score NoResultsFound.

TableXLV-V5-H-LA-B5101-10mers- 24P4C12 Pos 1234567890 score NoResultisFound.

TableXLV-V6-HLA-B5101-1 Omers- 24P4C12 Pos 1234567890 score NoResultsFound.

TableXLV.V7.HLA-B5101-10mers- 24P4C12 Pos 1234567890 score NoResuftsFound TableXLV-V3.HLAB5I01*l0mers- 24P4C12 Pos 1234567890 score NoRosultsFound TableXLV-V9-HLA-B51 01 -l0mers- 24P4C12 Pos 1234567890 score NoResuftsFound.

WO 2004/050828 WO 204/00828PCT/US2002/03826f TableXLVI*V1 *HLA-DRB1 .01 01- 115mers-24341C12 Each pepdde is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start position Plus fourteen.

Pos 123456789012345 scor e 227 QSWYWVILVALGVALV 39 206 SGLIDSLNARDISVK 33 247 ILLLRLVAGPLVLVL 33 313 LIVLAVLEAILLLML 33 601 FGKLLWVGGVGVLSF 33 246 FILLLRLVAGPLVLV 32 262 ILGVLGVLAYGIYYC 32 353 STMFYPLVTFVLLLI 32 368 ClAY WAMTALYLATS 32 652 ASGFFSVFGMCVDTL 32 39 VLFLLFILGYIVVGI 31 181 CFPWTNVTPPALPGI 31 277 WEEYRVLRDKGASIS 31 559 AYIMIAIYGKNFCVS 31 639 LPIMTSILGAYVIAS 31 YLLYFNIFSCILSSN 30 89 FNIFSCILSSNIISV 30 257 LVLVLILGVLGV'LAY 30 259 LVLILGVLGVLAYGI 30 30 646 LGAYVIASGFFSVFG 30 235 ALGVALVLSLLFILL 29 345 SKAVGQMMSTMFYPL 29 LFLLFILGYIWPGIV 28 242 LSLLFILLLRLVAGP 28 359 LVTFVLLLICIAYWA 28 453 TLNWVLALGQCVLAG 28 612 VLSFFFFSGRIPGLG 2a 640 PIMTSILGAYVIASG 28 167 CPSFLLPSAPALGRC 27 243 SLLFILLLRLVAGPL 27 280 YRVLRDKGASISQLG 27 362 FVLLLICIAYWAMvTA 27 423 MCVFQGYSSKGLIQR 27 501 SLAFGALILTLVQIA 27 575 KNAFMLLMRNIVRVV 27 129 KNEFSQTVGEVFYTK 26 230 YWVILVALGVALVLSL 26 254 AGPLVLVLILGVLGV 28 384 QPQYVLWASNISSPG 26 436 ORSVFNLQIYGVLGL 26 437 RSVFNLQIYGVLGLF 26 448 LGLFWTLNWVLALGQ 26 492 IRTLRYHTGSLAFGA 26 551 FIKFLNRNAYIMIAI 26 594 VTDLLLFFGKLLWVG 26 633 HLNYYWVLPIMTSILG 26 688 SKSLLKILGKKNEAP 26 44 FILGYIVVGIVAWLY 25 53 IVA WLYGDPRO VLYP 25 62 RQVLYPRNSTGAYCG 25 NIFSCILSSNIISVA 25 228 SWYWILVALGVALVL 25 TableXLVI-VI -HLA-DRBI -0101l5mers-24P4CI2 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 123456789012345 scor e 231 WILVALGVALVLSLL 239 ALVLSLLFILLLRLV 293 LGFTNLSAYQSVQE 299 LSAYQSVQETWLAA[ 304 SVQETWLAALIVLAV 319 LEAILLLMLIFLRQR 326 MLIFLRQRIRIAIAL 337 AIALLKEASKVGQM 354 TMFYPLVTFVLLLIC 371 YWAMTALYLATSGQP 399 CEKVPINTSCNPTAH 451 FWTfLNW\'LALGQCVL 454 LNWVLALGQCVLAGA 471 SFYWAFHKPQDIPTF 482 IPTFPLISAFIRTLR 526 LRGVQNPVARCIMCC 583 RNIVRVWVLDKVTDL 603 KLLVVGGVGVLSFFF 51 VGIVAWLYGDPRQVL 24 97 SSNIISVAENGLQCP 24 229 WYWILVALGVALVLS 24 238 VALVLSLLFILLLRL 24 255 GPLVLVLILGVLGVL 24 256 PLVLVLILGVLGVLA 24 279 EYRVLRDKGASISQL 24 307 ETWLAALIVLAVLEA 24 310 LAALIVLAVLEAILL 24 383 GQPQYVLWASNISSP 24 420 PGLMCVFQGYSSKGL 24 459 ALGQCVLAGAFASFY 24 506 ALILTLVQIARVILE 24 523 DHKLRf3VQNPVARCI 24 569 NFCVSAKNAFMLLMR 24 579 MLLMRNIVRVWLDK 24 588 WVLDKVTDLLLFFG 24 607 VGGVGVLSFFFFSGR 24 644 SILGAYVIASGFFSV 24 660 GMCVDTLFLCFLEDL 24 47 GYIWGIVAWLYGDP 23 59 GDPRQVLYPRNSTGA 23 165 ELCPSFLLPSAPALG 23 166 LCPSFLLPSAPALGR 23 241 VLSLLFILLLRLVAG 23 374 MTALYLATSGQPQYV 23 412 AHLVNSSCPGLMCVF 23 507 LILTLVQIAR\/ILEY 23 508 ILTLVQIARVILEYI 23 566 YGKNFCVSAKNAFML 23 604 LLWGGVGVLSFFFF 23 636 YYWLPIMTSILGAYV 23 33 TDVICCVLFLLFILG 22 43 LFILGYIVVGIVAWL 22 88 LLYFNIFSCILSSNI 22 WO 2004/050828 WO 204100828PCT/US20021038264 TableXLVI-V1.HLA-DRB1 -01 01 1 5mers*24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is spefied, the length of peptide is 15 amino acids, and the end position for each peptide is the start position plus fourteen, Pos 123456789012345 scar e 160 TSLQOELCPSFLLPS 22 198 DTTIQQG(SGLIDSL 22 312 ALl VLA VLEAILLLM 22 316 LAVLEAILLLMLIFL 22 349 GQMMSTMFYPLVTFV 22 363 VLLLICIAYWAMTAL 22 419 CPGLMCVFQGYSSKG 22 439 VFNLQIYGVLGLFWT 22 441 NLQIYGVLGLFWrLN 22 458 LALGQCVLAGAFASF 22 481 DIPTFPLISAFIRTL 22 511 LVOIARVILEYIDH 22 587 RWVLDKVTDLLLFF 22 598 LLFFGKLLWGGVGV 22 655 FFSVFGMCVDTLFLC 22 689 KSLLKILGKKNEAPP 22 138 EVFYTKNRNFCLPGV 21 151 GVPWYNMTVITSLQQE 21 153 PWNMTV1TSLQQELC 21 203 QGISGLIDSLNARDI 21 300 SAYQSVQETWLAALI 21 329 FLRQRIRIAIALLKE 21 331 RQRIRIAIALLKEAS 21 409 NPTAHLVNSSCPGLMV 21 518 ILEYIDHKLRGVQNP 21 546 LEKFIKFLNRNAYIM 21 606 WGGVGVLSFFFFSG 21 DEAYGKPVKYDPSFR 20 DPSFRGPIKNRSCTD 20 272 GIYYCWEEYRVLRDK 20 333 RIRIAIALLKEASKA 20 449 GLFWVTLNWVLALGQC 20 476 FHKPQCDIPTFPLISA 20 543 CCLWCLEKFIKFLNR 20 563 IAIYGKNFCVSAKNA 20 599 LFFGKLLVVGGVGVL 20 614 SFFFFSGRIPGLGKD 20 634 LNYYWLPIMTSILGA 20 645 ILGAYVIASGFFSVF 20 656 FSVFGMCVDTLFLCF 20 657 SVFGMCVDTLFLCFL 20 37 CCVLFLLFILGYIVV 19 38 CVLFLLFILGYIWVG 19 82 DKPYLLYFNIFSCIL 19 122 EDPWTVGKNEFSQTV 19 179 GRCFPWTNVTPPALP 19 184 WTNVTPPALPGITND 19 245 LFILLLRLVAGPLVL 19 271 YGIYYCWEEYRVLRD 19 317 AVLEAILLLMLIFLR 19 323 LLLMLIFLRQRIRIA 19 336 IAIALLKEASKAVGQ 19 369 IAYWAMTALYLATSG 19 TableXLVI-V1 -HLA-DRBI-01 01- 15rrmers-24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of pepide is amino acids, and the end position for each peptide is the start position plus fourteen, Pos 123456789012345 scar e 411 TAHLVNSSCPGLMCV 19 442 LQIYGVLGLFWTfLNW 19 460 LGQCVLAGAFASFYW 19 495 LRYHTGSLAFGALIL 19 M03 AFGALILTLVQIARV 19 657 RNAYIMIAIYGKNFC 19 586 VRVVVLDKVTDLLLF 19 683 RPYYMSKSLLKILGK 19 684 PYYMSKSLLKILGKK 19 TablaXLVl-V3-HLA-DRSI-01 01- I 5mers-24P4C1 2 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 15 amnino acids, aid the end position for each peptide is the start Position plus fourteen.

Pos 123456789012345 scare 9 CFPWTNIrPPALPGI 31 7 GRCFPWTNITPPALP 19 12 WTNITPPALPGITND 19 10 FPWTNITPPALPGIT 18 14 NITPPALPGITNDTT 16 TableXLVI.V5-HLA-DRBI -01 01.

l5mers-24P4CI2 Each peptide is a portion of SEQ ID NO: 11; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Pus 123456789012345 score 2 LIVLAVLEAILLLVL 33 8 LEALLLVLIFLRQR 15 VLIFLRQRIRIAIAL 1 ALl VLAVLEAILLLV 22 5 LAVLEAILLLVLIFL 22 6 AVLEAILLLVLIFLR 19 12 LLLVLIFLRQRIRIA 19 13 LLVLIFLRQRIRIAI 18 7 VLEAILLLVLIFLRQ 17 11 ILLLVLIFLRQRIRI 17 14 LVLIFLRQRIRIAIA 17 4 VLAVLEAILLLVLIF 16 10 AILLLVLIFLRQRIR 16 TableXLVI-V6-HLA-DRBI-0101l5mersi-24PC1 2 WO 2004/050828 WO 204100828PCTIUS2002/038264 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 123456789012345 score 2 MCVFQGYSSKGLIPR 27 PRSVFNLQIYGVLGL 26 7 GYSSKGLIPRSVFNL 24 4 VFQGYSSKGLIPRSV 16 SKGLIPRSVFNLQIY 16 12 GLIPRSVFNLQIYGV 16 1 LMCVFQGYSSKe3LIP 15 8 YSSKG3LIPRSVFNLQ 15 TableXLVI-VT-HLA-DRBi -01 01- I 5mers-24P34CI2 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 123456789012345 score 6 QSWYWILVAVGQMMS 31 12 LVAVGQMMSTMFYPL 29 7 SWYWILVAVGQMNIST 25 8 WVYWILVAVGQMMSTM 24 9 YINILVAVGQMMSTMF 24 1 FEDFAQSWYWVILVAV 18 AQSWYWILVAVGQMM 16 11 ILVAVGQMMSTMFYP 15 TableXILVII-V8I-11-LA-DRB-01101- I 5mers-24P4C12 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptice is the start position plus fourteen.

Pos 123456789012345 score 24 VFQTSILGAYVIASG 28 7 NYYWLPIMRNPITPT 24 23 H-VFQTSILGAYVIAS 23 6 LNYYWLPIMRNPITP 20 HLNYYWLPIMRNPIT 18 21 TGHVFQTSILGAYVI 1iB 3 SPHLNYYWLPIMRNP 17 8 YYWLPIMRNPITPTG 17 13 IMRNPITPTGI-VFQT 17 11 LP(MRNPITPTGHVF 16 12 PIMRNPITPTGHVFQ 16 14 MRNPITPTGHVFQTS 16 26 QTSILGAYVIASGFF 16 9 YWLPIMRNPITPTGH 15 18 ITPTGHVFQTSILGA 15 19 TPTGHVFQTSILGAY 14 PTGHVFQTSILGAYV 14 TableXLVII*VS-HL-11A-DRIB 10101- 15mers-24P4C12 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 123456789012345 score 4 ClAY WAMTALYPLPT 32 10 MTALYPLPTQPATLG 22 TLGYVLWASNISSPG 26 21 ATLGYVLWASNISSP 24 7 YWAMTALYPLPTQPA 23 13 LYPLPTQPATLGYVL 23 5 IAY WAMTALYPLPTQ 19 2 LICIAYWAMTALYPL 17 1 LLICIAYWAMTALYP 16 16 LPTQPATLGYVLWAS 16 23 LGYVLWASNISSPGC 16 24 GYVLWASNISSPGCE 16 9 AMTALYPLPTQPATL TabIeXLVI*VI -HLA-DRB1 .0301- I 5rers-24P4C1 2 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 12345678901 2345 score 54 VAWLYGDPRQVLYPR 36 586 VRVIJ\JLDKVTDLLLF 31 667 FLCFLEDLERNNGSL 29 312 ALIVLAVLEAILIIM 28 97 SSNIISVAENGLQCP 27 155 NMTVITSLQQELCPS 27 454 LNWVLALGQCVLAGA 27 549 EKFIKFLNRNAYIMI 27 136 VGEVFYTKNRNFCLP 26 506 ILTLVQIARVILEYI 26 622 IPGLGKDFKSPHLNY 26 376 ALYLATSGQPQYVLW 447 VLGLFWTLNWVLALG 279 EYR\/LRDKGASISQL 24 534 AROIMCCFKCCLWCL 24 567 GKNFCVSAKNAFMLL 24 229 WYWILVALGVALVLS 23 238 VALV[SLLFILLLRL 23 14 GKPVKYDPSFRGPIK 22 218 SVKIFEDFAQSWYWI 22 219 VKIFEDFAQSWYWIL 22 235 ALGVALVLSLLFILL 22 241 VLSLLFILLLRLVAG 22 360 VTFVLLLICIAYWAM 22 515 ARVILEYIDHKLRGV 22 594 VTDLLLFFGKLLWVG 22 33 TDVICCVLFLLFILG 21 167 CPSFLLPSAPALGRC 21 192 LPGITNDTTIQQGIS 21 237 GVALVLSLLFILLLP 21 239 ALVLSLLFILLLRLV 21 260 VLILGVLGVLAYGIY 21 302 YQSVQETWLAALIVL 21 319 LEAILLLMLIFLRQR 21 431 SKGLIQRSVFNLQIY 21 WO 2004/050828 WO 204100828PCTIUS2002/038264 TableXLVII-VI-HLADRB1.0301l5mers-24P4C1 2 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start positionl plus fourteen.

Pos 123456789012345 score 461 GQCVLAGAFASFYWA 21 587 RWVLDKVTDLLLFF 21 590 VLDKVTODLLLFFGKL 21 595' TDLLLFFGKLLVVGG 21 658 VFGMCVDTLFLCFLE 21 32 CTDVICCVLFLLFIL 20 37 CCVLFLLFILGYIW 20 46 LGYIVVGIVAWLYGD 20 47 GYIWVGIVAWLYGDP 20 74 YCGMGENKOKPYLLY 20 76 GMGENKDKPYLLYFN 20 231 WILVALGVALVLSLL 20 233 LVALGVALVLSLLFI 20 246 FILLLRLVAGPLVLV 20 250 LRLVAGPLVLVLILG 20 255 GPLVLVLILG3VLGVL 20 258 VLVLILGVLGVLAYG 20 313 LIVLAVLEAILLLML 20 316 LAVLEAILLLMLIFL 20 323 LLLMLIFLRQRIRIA 20 338 IALLKEASKAVGQMM 20 411 TAHLVNSSCPGLMCV 20 439 VFNLQIYGVLGLFWT 20 484 TFPLISAFIRTLRYH 20 559 AYIMIAIYGKNFCVS 20 588 VVLDKVTDLLLFFG 20 602 GKLLVVGGVGVLSFF 20 604 LLWVGGVGVLSFFFF 20 691 LLKILGKKNEAPPDN 156 MTVITSLQQELCPSF 19 159 ITSLQQELCPSFLLP 19 205 ISGLIDSLNARDISV 1s 335 RIAIALLKEASKAVG 1s 348 VGQMMSTMFYPLVTF 19 366 LICIAYWAMTALYLA 19 385 PQYVLWASNISSPGC 19 505 GALILTLVQIARVIL 19 576 NAFMLLMRNIVRWV 1S 607 VGGVGVLSFFFFSGR 19 626 GKDFKSPH-LNYYWLP 19 638 WLPIMTSILGAYVIA 19 648 AYVIASGFFSVFGMC 19 663 VDTLFLCFLEDLERN 19 668 LCFLEDLERNNGSLD 19 684 PYYMSKSLLKILGKK 19 689 KSLLKILGKKNEAPP 19 3 GKQRDEDDEAYGKPV 18 61 PRQVLYPRNSTGAYC 18 98 SNIISVAENGLOCPT 18 114 QVCVSSCPEDPWTVG 18 214 ARDISVKIFEDFAQS 18 243 SLLFILLLRLVAGPL 18 263 LGVLGVLAYGIYYCW 18 327 LIFLRQRIRIAIALL 18 345 SKAVGQMMSTMFYPL 18 TabIOXLVII-V1 -HLA-DRBI-0301 1 5mers-24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start positon is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 123456789012345 score 462 QCVLAGAFASFYWAF 18 530 QNPVARCIMCCFKCC 18 560 YIMIAIYGKNFCVSA 18 569 NFCVSAKNAFMLLMR 18 579 MLLMRNIVRVWLDK 18 585 IVRVVVLDKVTDLLL 18 555 FFSVFGMCVDTLFLC 18 656 FSVFGMCVDTLFLCF 18 560 GMCVDTLFLCFLEDL 18 664 DTLFLCFLEDLERNN 18 284 RDKGASISQLGFTTN 17 290 ISQLGFTTNLSAYQS 17 324 LLMLIFLRQRIRIAI 17 325 LMLIFLRQRIRIAIA 17 353 STMFYPLVTFVLLLI 17 423 MCVFQGYSSKGLIQR 17 437 RSVFNLQIYGVLGLF 17 485 FPLISAFIRTLRYHT 17 517 VILEYIDHKLRGVOI\ 17 519 LEYIDHKLRGVQNPV 17 523 DHKLRGVQNPVARCI 17 542 KCCLWVCLEKFIKFLN 17 545 LWCLEKFIKFLNRNA 17 546 LEKFIKFLNRNAYIM 17 614 SFFFFSGRIPGLGKD 17 619 SGRIPGLGKDFKSPH 17 670 FLEDLERNNGSLDRP 17 692 LKILGKKNEAPPDNK 17 TableXLVII-V3-HLA-DRBI-0301 24P4C1 2 Each peptide is a porton of SEQ 10 NO: 7; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 123456789012345 score 12 WTNITPPALPGITND 12 3 APALGRCFPWVTNIrP 9 CFPWTNITPPALPGl 7 GRCFPWTNITFPALP 8 6 LGRCFPVVTNITPPAL 7 TableXLVlI-V5-HLA-DRBi *0301l5mners-24P4C12 Each peptide is a portion of SEQ ID NO: 11; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 123456789012345 score 1 ALIVLAVLEAILLLV 28 8 LEAILLLVLIFLRQR 21 2 LIVLAVLEAILLLVL WO 2004/050828 WO 204100828PCT/US2002103826-t -0301.

I 5mers-24P4C12 Each peptidle is a portion of SEQ ID NO: 11; each start posi Von is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 123456789012345 score LAVLEAILLLVLIFL 20 12 L.LLVLIFLRQRIRIA 20 13 LLVLIFLRQRIRIAI 17 14 LVLIFLRQRIRIAIA 17 4 VLAVLEAILLLVLIF 15 9 EAILLLVLIFLRQRI 15 AILLLVUIFLRQRIR 13 TableXLVII-V6-HLA-DRBI-0301- 15mers-24P4C12 Each peptidle is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide Is the start position plus fourteen.

Pos 123456789012345 score SKGLIPRSVFNLQIY 22 2 'MCVFQGYSSKGLIPR 17 8 YSSKGLIPRSVFNLQ 16 11 KGLIPRSVFNLQIYG 12 1 LMCVFQGYSSKGLIP 11 PRSVFN[QIYGVLGL 10 TableXLVII-V7-HLA*DRBi-0301-1 5mers- 24P4C12 Each peptidle is a portion of SEQ ID NO: each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptidle is the start position plus fourteen.

Pos 123456789012345 score 9 YWILVAVGQMMSTMF- 18 12 LVAVGQM~MSTMFYPL 18 1 FEDFAQSWYWILVAV 16 8 WVYWILVAVGQMMSTM 13 WILVAVGQMMSTMFY 10 13 VAVGQMMSTMFYPLV 10 TableXLVIl-V8.HLA.DRBI-0301-1 5rers- 24P4C12 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peplidle is the start position plus fourteen.

Pos 123456789012345 score 22 GHVFQTSILGAYVIA 17 8 YYWLPIMRNPITPTG 16 RNPITPTGHVFQTSI 14 26 QTSILGAYVIASGFF 13 21 TGHVFQTSILGAYVI 12 WLPIMRNPITPTGHV 11 11 LPIMRNPITPTGHVF 11 3 SPHLNYYWLPIMRNP 10 7 NYYWLPIMRNPITPT 10 14 MRNPITPTGHVFQTS 19 TPTGHVFQTSILGAY TableXL\lI-V9-HLA-DRB1-0301 -1 24P4C1 2 Each peptidle is a portion of SEQ ID NO: 19; each start position is specified, the length of pepide is 15 amino acids, and the end position for each peptidle is the start position plus fourteen.

Pos 123456789012345 score 2 LICIAYWAMTALYPL 19 23 LGYVLWASNISSFGC 19 10 MTALYPLPTQPATLG 13 7 YWAMTALYPLPTQPA 12 12 ALYPLPTQPATLGYV 12 13 LYPLPTQPATLGYVL 12 20 PATLGYVLWASNISS 12 3 ICIAYWVAMTALYPLP 14 YPLPTQPATLGYVLW 24 GYVLWASNISSPGCE 5 IAYWVAMTALYPLPTQ 9 16 LPTQPATLGYVLWAS 9 T ab~eXLVIII-VI-DR1 -0401-1 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptidle is the start position plus fourteen.

Pos 123456789012345 score 85 YLLYFNIFSCILSSN 28 89 FNIFSCILSSNIISV 28 243 SLLFILLLRLVAGPL 28 353 STMFYPLVTFVLLLI 28 469 FASFYWVAFHKPQDIP 28 548 LEKFIKFLNRNAYIM 28 575 KNAFMLLMRNIVRVJV 28 635 NYYWLPIMTSILGAY 28 54 VAWLYGDPRQVLYPR 26 98 SNIISVAENGLQCPT 26 153 PWNMTVITSLQQELC 26 189 PPALPGITNDTTIQQ 26 192 LPGITNOTTIQQGIS 26 323 LLLMLIFLRQRIRIA 26 337 AIALLKEASKAVGQM 26 385 PQYVLWASNISSPGC 26 419 CPGLMCVFQGYSSKG 26 454 LN'WLALGQCVLAGA 26 508 ILTIVOIARVILEYI 26 523 DI-KLRGVQNPVARCI 26 579 MLLNRNIVRVVVLDK 26 16 PVKYDPSFRGPIKNR 22 38 O\/LFLLFILGYIWVG 22 82 DKPYLLYFNIFSCIL 22 86 LLYFNIFSCILSSNI 22 122 EDPVV[FVGKNEFSQTV 22 138 EVFYTKNRNFCLPGV 22 181 CFPWTNVTPPALPGI 22 219 VKIFEDFAQSVVIL 22 227 QSWYWILVALGVALV 22 228 SWYWILVALGVALVL 22 WO 2004/050828 TableXLVIII-V-OR-040)1-5mers- 24P4C1 2 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start position pls fourteen.

Pos 123456789012345 score 272 GIYYCWEEYRVLRDK 22 277 WEEYRVLRDKGASIS 22 292 QLGFTTNLSAYQSVQ 22 299 LSAYOSVQETWLAAL 22 306 QETVLAUVLAVLE 22 354 TMFYPLVTFVLLLIC 22 359 LVTFVLLLICIAYVVA 22 384 QPQYVLWASNISSPG 22 423 MCVFQGYSSKGLIQR 22 442 LQIYGVLGLFWTLNW 22 448 LGLFWTLNVVVLALGQ 22 453 TLNWVLALGQCVLAG 22 488 ISAFIRTLRYHTGSL 22 5011 SLAFGALILTLVQIA 22 557 RNAYIMIAIYGKNFC 22 633 HLNYYWLPIMTSILG 22 646 LGAYVIASG3FFSVFG 22 652 ASGFFSVFGMCVDTL 22 667 FLCFLEDLERNNGSL 22 682 DRPYYMSKSLLKILG 22 14 Gr(PVKYDPSFRGPIK 20 39 VLFLLFILGYIVVGI 20 LFLLFILGYIWGIV 20 43 LFILGYIVVGIVAWL 20 97 SSNIISVAENGLQCP 20 133 SQTVGEVFYTKNRNF 20 146 NFCLPGVPWNMTVIT 20 149 LPGVPWNMTVITSLQ 20 155 NMTVITSLQQELCPS 20 156 MTVITSLQQELCPSF 20 198 DTTIOQGISGLIDSL 20 202 QQGISGLIDSLNARD 20 206 SGLIDSLNARDISVK 20 216 DISVKIFEDFAQSWY 20 229 WIYWILVALGVALVLS 20 230 YWILVALGVALVLSL 20 233 LVALGVALVLSLLFI 20 235 ALGVALVLSLLFILL 20 238 VALVLSLLFILLLRL 20 239 ALVLSLLFILLLRLV 20 241 VLSLLFILLLRLVAG 20 242 LSLLFILLLRLVAGP 20 246 FILLLRLVAGPLVLV 20 247 ILLLRLVAGPLVLVL 20 254 AGPLVLVLILGVLGV 20 255 GPLVLVLILGVLGVL 20 257 LV.LVLILGVLGVLAY 20 259 LVLILGVLGVLAYGI 20 262 FLGVLGVLAYGIYYC 20 279 EYRVLRDKGASISQL 20 287 GASISQLGFTTNLSA 20 290 ISQLGFTTNLSAYQS 20 307 ETWLAALIVLAVLEA 20 310 LAALIVLAVLEAILL 20 311 MALIVLAVLEAILLL 20 PCTIUS2002/038264 TableXLVlIl-VI -DRI-0401-1 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Pas 123456789012345 score 312 ALIVLAVLEAILLLM 313 LIVLAVLEAILLLML 315 VLAVLEAILLLMLIF 316 LAVLEAILLLMLIFL 319 LEAILLLMLIFLRQR 321 AILLLMLIFLRQ)RIR 324 LLMLIFLRQRIRIAI 331 RQRIRIAIALLKEAS 333 RIRIAIAIIKEASKA 335 RIAIALLKEASKVG 356 FYPLVTFVLLLICIA 353 VLLLICIAYWAMTAL 364 LLLICIAYWAMTALY 371 YWAMTALYLATSGQP 374 MrALYLATsGQPQYV 401 KVPINTSCNPTAHLV 420 PGLMCVFQGYSSKGL 436 QRSVFNLQIYGVLGL 444 IYGVLGLFWTLNW'/[ 445 YGVLGLFWTLNWVLA 447 VLGLFWTLNWVVLALG 451 FWVTLNWVLALGQCVL 475 PQDIPTFF[ISAFIR 484 TFPLISAFIRTLRYH 485 FPLISAFIRTLRYHT 505 GALILTLVQIARVIL 506 ALILTLVQIARVILE 511 LVQIARVILEYIDHK 514 IARVILEYIDHKLRG 516 RVILEYIDHKLRGVQ 542 KCCLWCLEKFIKFLN 545 LWCLEKFIKFLNRNA 549 EKFIKFLNRNAYIMI 558 NAYIMIAIYGKNFCV 582 MRNIVRVWVLDKVTO 583 RNIVRVVVLDKVTD[ 586 VRVVVLDKVTDLLLF 588 VVVLDKVTDLLLFFG 594 VTOLLLFFGKLLWVG 595 TDLLLFFGKLLWVGG 601 FGKLLVVGGVGVLSF 619 SGRIPGLGKDFKSPH 639 LPIMTSILGAYVIAS 642 MTSILGAYVIASGFF 660 GMCVDTLELCF[EDL 668 LOFLEDLERNNGSLD 688 SKSLLKILGKKNEAP G0 NIFSCILSSNIISVA 18 125 WTVGKNEFSQTVGEV 18 152 VPWNMTVITSLQQEL 18 166 LCPSFLLPSAPALGR 18 195 ITNDTTiQQGISGLI 18 203 QGISGLIDSLNARDI 18 210 DSLNARDISVKIFED 18 289 SISQLGFTTNLSAYQ 18 WO 2004/050828 WO 2O4IOO82~PCT/US20021038264 TableXLVIIIVI-DRI -0401 -i5mers.

24P4C12 Each pepide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 123456789012345 score 295 FTTNLSAYQSVQETW 18 342 KEASKAVGQMMSTMF 18 373 AMTALYLATSGQPQY 18 398 GCEKVPINTSCN4PTA 18 428 GYSSKGLIQRSVFNL 18 433 GLIQRSVFNLQIYGV 18 476 FHKPQDIPTFPLISA 18 481 DIPTFPLISAFIRTL 18 502 LAFGALILTLVQIAR 18 527 RGVQNPVARCiMCCF 18 568 KNFCVSAKNAFMLLM 18 611 GVLSFFFFSGRIPGL 18 623 PGLGKDFKSPHLNYY 18 657 SVFGMCVDTLFLCFL 18 669 CFLEDLERNNGSLOR 18 DPSFRGPIKNRSCTD 16 ILGYIVVGIVAWVLYG 16 53 IVAWLYGDPRQVLYP 16 AWLYGDPRQVLYPRN 16 63 QVLYPRNSTGAYCGM 16 144 NRNFCLPGVFWNMTV 16 151 GVPWWMTVITSLQQE 16 167 CPSFLLPSAPALGRC 16 222 FEDFAQSWYWILVAL 16 226 AQSWYWILVALGVAL 16 271 YGIYYCWEEYRVLRD 16 326 MLIFLRQRIRIAIAL 16 368 CIAYWVAMTALYLATS 16 369 IAYWAMTALYLATSG 16 375 TALYLATSGQPQYVL 16 387 YVLWASNISSPGCEK 16 437 RSVFNLQIYGVLGLF 16 449 GLFWTfLNWVVLALGQC 16 466 AGAFASFYVVAFHKPQ 16 470 ASFYWAFHKPQDIPT 16 471 SFYWAFHKPQDIPTF 16 473 YWAFHKPODIPTFPL 16 482 IPTFPLISAFIRTLR 16 518 ILEYIDHKLRGVQNP 16 543 CCLWCLEKFIKFLNR 16 563 IAIYGr(NFCVSAKNA 16 598 LLFFGKLLVWGGVGV 16 612 VLSFFFFSGRIPGLG 16 613 LSFFFFSGRIPGLGK ,16 614 SFFFFSGRIPGLGKD 16 634 LNYYWLPIMTSILGA 16 653 SGFFSVFGMCVDTLF 16 664 DTLFLCFLEDLERNN 18 62 RQVLYPRNSTGAYCG 15 325 LMLIFLRQRIRIAIA 15 327 LIFLRQRIRIAIALL 15 519 LEYIDHKLRGVQNPV 15 587 RVVVLO)KVTDLLLFF 15 32 CTDVICCVLFLLFIL 14 33 TDVICCVLFLLFILG 14 TableXLVIl-VI-DR1-0401-l5mers- 24P4C1 2 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length ot peptidle is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 123456789012345 score 36' ICCVLF[LFILGYIV 14 37 CCVLFLLFILGYIVV 14 42 LLFILGYIWVGIVAW 14 46LGYIV VGIVAWLYGD 14 47 GYIWVGI.VAWLYGDP 14 48 YIWVGIVAWLYGDPR 14 51 VGIVAWLYGDPRQVL 14 61 PRQVLYPRNSTGAYC 14 83 KPYLLYFNIFSCILS 14 84 PYLLYFNIFSCILSS 14 88 YFNIFSCILSSNIIS 14 92 FSCILSSNIISVAEN 14 93 SCILSSNIISVAENG 14 124 PWTVGKNEFSQTVGE 14 136 VGEVFYTKNRNFCLP 14 159 ITSLQQELCPSFLLP 14 163 QQELCPSFLLPSAPA 14 169 SFLLPSAPALGRCFP 14 175 APALGRCFPWTMVTP 14 184 WTNVTPPALPGITND 14 205 ISGLIDSLNARDISV 14 218 SVKIFEDFAQSWYWI 14 231 WVILVALGVALVLSLL 14 237 GVALVLSLLFILLLR 14 244 LLFILLLRLVAGPLV 14 249 LLRLVAGPLVLVLIL 14 250 LRLVAGPLVLVLILG 14 256 PLVLVLILGVLGVLA 14 258 VLVLILGVLGVLAYG 14 260 VILGVLGVLAYGIY 14 263 LGVLGVLAYGIYYCV4 14 296 TTNLSAYQSVQETWVL 14 302 YQSVIQETWLAALIVL 14 322 ILLLMLIFLRQRIRI 14 338 IALLKEASKAVGQMM 14 345 SKAVGQMMSTMFYPL 14 348 VGQMMSTMFYPLVTF 14 349 GQMMSTMFYPLVTFV 14 352 MSTMFYPL'IrFVLLL 14 357 YPLVIFVLLLIC[AY 14 360 VTFVLLLICIAYWAM 14 361 TFVLLLICIAYWAMT 14 362 FVLLLICIAYWAMvTA 14 366 LICIAYWAMTALYLA 14 376 ALYLATSGQPQYVLW 14 391 ASNISSPGCEKVPIN 14 399 CEKVPINTSCNPTAH 14 411 TAHLVNSSCPGLMICV 14 412 AHLVNSSCPGLMCVF 14 422 LMCVIFQGYSSKGLIQ 14 432 KGLIQRSVFNLQIYG 14 439 VFNLQIYGVLGLFWT 14 441 NLQIYGVLGLFWTLN 14 455 NWVLALGQCVLAGAP 14 457 VLALGQCVLAGAFAS 14 WO 2004/050828 WO 2004050828PCT/US2002/038264t TableXLVIII-VI .DRI-0401-1 5mers- 24P4C1 2 Each peptide is a portion of SEQ ID NO: 3: each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 123456789012345 score 462 QCVLAGAFASFYWAF 14 489 SAFIRTLRYIHTGSLA 14 492 IRTLRYHTGSLAFGA 14 499 TGSLAFGALILTLVQ 14 504 FGALILTLVQIARVI 14 LTLVQIARVILEYID 14 515 ARVILEYIDHKLRGV 14 526 LRGVQNPVARCIMCC 14 534 ARCIMCCFKCCLWCL 14 535 RCIMCCFKCCLWCLE 14 552 IKFLNRNAYIMIAIY 14 559 AYIMIAIYGKNFCVS 14 576 NAFMLLMRNIVRVVV 14 578 FMLLMRNIVRWVLD 14 585 IVRVVVLDKVTOLLL 14 591 LDKVTDLLLFFGKLL 14 596 DLLLFFGKLLVVGGV 14 602 GKLLVVGGVGVLSFF 14 603 KLLWVGGVGVLSFFF 14 604 LLVVGGVGVLSFFFF 14 607 VGGVGVLSFFFFSGR 14 609 GVGVLSFFFFSGRIP 14 610 VGVLSFFFFSGRIPG 14 622 IPGLGKDFKSPHLNY 14 631 SPHLNVYWLPIMTSI 14 636 YYVVLPIMTSILGAYV 14 647 GAYVIASGFFSVFGM 14 655 FFSVFGMCVDTLFLC 14 658 VPGMCVDTLFLCFLE 14 663 VDTLFLCFLEDLERN 14 665 TLFLCFLEDLERNNG 14 678 NGSLDRPYYMSI SLL 14 684 PYYMSKSLLKILGKK 14 689 KSLLKILGKKNEAPP 14 TableXLVIII-V3-HLADRI-0401.

l15mers-24P34C12 Each peptide is a portion of SEQ ID NO: 7; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptidle is the start position plus fourteen.

Pos 123456789012345 score 9 CFPWTNITPPALPGI 22 3 APALGRCFPWNITP 14 12 WTNITPPALPGITND 14 4 PALGRCFPWTNITPP 12 ALGRCFPWTNITPPA 12 8 RCFPWNITPPALPG 12 13 TNITPPALPGITNDT 12 14 NITPPALPGITNDTT 12 7 GRCFPWVTNITPPALP 10 TableXLVIII-V-R-0401-5mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 11; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 123456789012345 score 12 LLLVLIFLRQRIRIA 26 1 ALl VLAVLEAILLLV 2 LIVLAVLEAILLLVL 4 VLAVLEAILLLVLIF 5 LAVLEAILLLVLIFL 8 LEAILLLVLIFLRQR 10 AILLLVLIFLRQRIR 13 LLVLIFLRQRRIAI 15 VLIFLRQRIRIAIAL 16 14 LVUIFLRQRIRIAIA 9 EAILLLVLIFLRQRI 14 11 ILLLVLIFLRQRIRI 14 3 IVLAVLEAILLLVLI 12 6 AVLEAILLLVLIFLR 12 TableXLVIIi-V6-HLA-DRIl .0401-1 24P4C12 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 123456789012345 score 2 MCVFQGYSSKGLIPR 22 15 PRSVFNLQIYGVLGL 12 GLIPRSVFNLQIYGV 18 1 LMCVFQGYSSKGLIP 14 11 KGLiPRSVFNLQIYG 14 7 GYSSKGIPRSVFNL 12 8 YSSKGLIPRSVFNLQ 12 9 SSKGLIPRSVFNLQI 12 TableXLVIII-V7-HLADRI-0401-l5mers- 24P4C12 Each peptide is a portion of SEQ ID NO: 15; each start position is specified, the length of peptidle is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Poe 123456789012345 score 9 YWILVAVGQMMSTMF 26 6 QSWYWILVAVGQMMS 22 7 SWYWILVAVGQMMST 22 8 WYWILVAVGQMMSTM 1 FEDFAQSWVYWILVAV 16 5 AQSVVYWILVAVGQMM 16 10 WILVAVGQMMSTMFY 14 12 LVAVGQMMSTMFYPL 14 TableXLVIII-VB.HLA.DR1 -0401M 24P34C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is '17 amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 123456789012345 score WO 2004/050828 TableXLVIII.V8HLADRI-0401l5mers- 24P4C12 Each pepide is a portion of SEQ ID NO:, 3; each start position is specified, the length of peptide is 17 amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 123456789012345 score 7 NYYVVLPIMRNPITPT 28 HLNYYWLPIMRNPIT 22 8 YYWLPIMRNPITPTG 20 RNPITPTGH-VFQTSI 20 26 QTSILGAYVIASGFF 20 18 ITPTGHVFQTSILGA 18 19 TPTGHVFQTSILGAY 18 3 SPH-LNYYWLPIMRNP 14 WLPIMRNPITPTGHV 14 11 LPIMRNPITPTGHVF 14 21 TGHVFQTSILGAYVI 14 TabIBXLVIII-V9HLA-DR11.04011- I 5mers-24P4C12 Each peptide is a portion cf SEQ ID NO: 19; each start position is specified, the length of peptide is 15 amnino acids, and the end position for each peptide is the start position plus fourteen.

Pos 123456789012345 score MTALYPLPTQPATLG 26 23 LGYVLWASNISSPGC 26 11 TALYPLPTQPATLGY 22 22 TLGYVLWASNISSPG 22 7 YWAMTALYPLPTQPA 20 PATLGYVLWASNISS 20 lAY WAMTALYPLPTQ 16 2 LICIAYWAMTALYPL 14 3 ICIAYWAMTALYPLP 12 PLPTQPATLGYVLWA 12 21 ATLGYVLWASNISSP 12 TableXLIX-VI -ORSi -11101-l5mers- 24P4C12 Each pepide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 123456789012345 score 243 SLLFILLLRLVAGPL 31 DEAYGKPVKYDPSFR 26 DPSFRGPIKNRSCTO 26 668 LCFLEDLERNNGSLD 26 575 KNAFMLLMRNIVRWV 25 613 LSFFFFSGRIPGLGK 25 226 AQSWYWILVALGVAL 23 228 SWYWILVALGVALVL 23 277 WEEYRVLRDKGASIS 23 359 LVTFVLLLICIAYWA 23 448 LGLFWT[NWVLALGQ 23 579 MLLMRNIVRWVVLE)K 23 598 LLFFGKLLWGGVGV 22 633 HLNYYWVLPIMTSILG 22 PCTIUS2002/038264 TableXLlX-VI-DRBi -11101 24P4C12 Each peptide is a portion of SEQ ID NO: 3; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start posiion plus fourteen.

Pos 123456789012345 score 276 CWEEYRVLRDKGASI 21 338 IALLKEASKAVGQM4M 21 508 ILTLVOIARVILEYI 21 516 RVILEYIDH-KLRGVQ 21 542 KCCLWCLEKFIKFLN 21 585 IVRVVVLDKVTDLLL 21 685 YYMSKSLLKILGKKN 21 172 LFSAPALGRCFPWTN 334 IRIAIALLKEASKAV 371 YWAMTALYLATSGQP 549 EKFIKFLNRNAYIMI 591 LDKVTDLLLFFGKLL 619 SGRIPGLGKDFKSPH- 689 KSLLKILGKKNEAPP 36 ICCVLFLLFILGYIV 19 122 EDPWTVGKNEFSQTV 19 256 PLVLVLILGVLGVLA 19 259 LVLILGVLGVLAYGI 19 310 LAALIVLAVLEAILL 19 353 S7MFYPLVTFVLLLI 19 523 DHKLRGVQNPVARCI 19 587 GKNFCVSAKNAFMLL 19 612 V[SFFFFSGRIPGLG 19 638 YYWLPIMTSILGAYV 19 16 PVKYDPSFRGPiKN'R 18 48 YIVVGIVAWLYGDPR 18 8b YLLYFNIFSCILSSN 18 137 GEVIFYTKNIRNFCLPRG 18 181 CFPWTNVTPPALPGI 18 227 QSWYWILVALGVALV 18 244 LLFILLLRLVAGPLV 18 326 MLIFLRQRIRIAIAL 18 419 OPGLMCVFQGYSSKG 18 469 FASFYWAFHKPQDIP 18 470 ASFYWAFHKPODIPT 18 488 ISAFIRTLRYHTGSL 18 489 SAFIRTLRYH-TGSLA 18 597 LLLFFGKLLWVGGVG 18 41 FLLFILGYIWGIVA 17 45 ILGYIWVGIVAWLYG 17 -71 TGAYCGMGENKDKPY 17 86 LLYFNIFSCILSSNI 17 306 QETWLAALIVLAVLE 17 325 LMLIFLRQRIRIAIA 17 354 TMFYPLVrFVLLLIC 17 369 lAY WAMTALYLATSG 17 384 QPQYVLWASNISSPG 17 442 LQIYGVLGLFWTLNW 17 482 IPTFPLISAFIRTLR 17 501 SLAFGALILTLVQIA 17 548 LEKFIKFLNRNAYIM 17 615 FFFFSGRIPGLGKDF 17 635 NYYVA.PIMTSILGAY 17 652 ASGFFSVFGMCVCTL 17 82 DI<PYLLYFNIFSCIL 16 WO 2004/050828 WO 204/00828PCT/US2002/038264 TableXLIX-V1 -DR1111 01-15mers.

Pos 123456789012345 score 89 FNIFSCILSSNIISV 16 179 GRCFPWTNVTPPALP 16 253 VAGPLVLVLILGVLG 16 299 LSAYQSVQETWLMAL 16 323 LLLMLIFLRQRIRIA 16 368 ClAY WAMTALYLATS 16 387 YVLWASNISSPGCEK 16 490 AFIRTLRYHTGSLAF 16 494 TLRYHTGSLAFGALI 16 506 ALILTLVQIARVILE 16 517 VILEYIDFIKLRGVQN 16 557 RNAYIMIAIYGKNFC 16 563 IAIYGKNFCVSAKNA 16 583 RNIVRWVVLDKVTDL 16 646 LGAYVIASGFFSVFG 16 43 LFILGYIWVGIVAWL 15 44 FILGYIVVGIVAWLY 15 47 GYIWVGIVAWLYGDP 15 54 VAWLYGDPRQVLYPR 15 73 AYCGMGENKDKPYLL 15 153 PWNMTVITSLQQELC 15 156 MTVITSLQQELCPSF 15 195 ITNDITIQQGISGLI 15 207 GLIDSLNARDISVKI 242 LSLLFILLLRLVAGP 15 357 YP[VTFVLLLICIAY 15 429 YSSKC3LIQRSVFNLQ 15 485 FPLISAFIRTLRYHT 15 519 LEYIDHKLRGVQNPV 15 527 RGVQNPVARCIMCCF 15 545 LWCLEKFIKFLNRNA 15 595 TDLLLFFGKLLWGG 15 600 FFGKLLVVGGVGVLS 15 603 KLLWVGGVGVLSFFF 15 681 LDRPYYMSKSLLKIL 15 TableXLIX-V3-HLA-DRBI-1 101-1 5mers- 24P4C12 Each peptidle is a portion of SEQ IC NO: 7; each start position is specified, the length of peptide is 15 amino acids, and the end position for each paptidle is the start position plus fourteen.

Pos 123456789012345 score 9 CFPWTNITPPALPGI 18 7 GRCFPWTNITPPALP 16 12 WTNITPPALPGITND 8 TabIeXLIX-V5.HLA.DRBI-1 I 01-l5mers- 24P4C12 Each peptidle is a portion of SEQ ID NO: 11; each start position-is specified, the length of pepfide is 15 amino acids, and the end position for each peptidle is the start position plus fourteen.

Pos 123456789012345 15 VLIFLRQRIRIAIAL 14 LVLIFLRORIRIAIA 12 LLLVLIFLRQRIRIA 10 AILLLVLIFLRQRIR 2 LIVLAVLEAILLLVL 8 LEAILLLVLIFLRQR 13 LLVLIFLRQRIRIAI 1 ALl VLAVLEAILLLV 5 LAVLEAILLLVLIFL 9 EAILLLVLIFLRQRI 11 ILLLVLIFLRQRIRI score 18 17 16 14 14 14 13 13 13 13 TabIeXLIX*VIS.HLA.DRBI-l1 101 l5mers-24P4C12 Each peptide is a portion of SEQ ID NO: 13; each start position is specified, the length of peptidle is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 12345678901 2345 score 8 YSSKGLIPRSVFNLQ 1 LMCVFQGYSSKGLIP 14 15 PRSVFNLQIYGVLGL 13 2 MCVFQGYSSKGLIPR 5 FQGYSSKGLIPRSVF 3 CVFQGYSSKGLIPRS 9 11 KGLIPRSVFNLQIYG 9 6 QGYSSKGLIPRSVFN 8 4 VFQGYSSKGLIPRSV 7 7 GYSSKGLIPRSVFNL 7 TableXLIX-W*.HLA-DRBI-1 101 24P4C12 Each peptido is a portion of SEQ I D NO: 15; each start position is specified, the length of peptidle is 15 amino acids, and the end position for each peptide is thre start position plus fourteen.

Pos 123456789012345 score 5 AQSWYWILVAVGQMM 23 6 QSWYWVILVAVGQMMS 18 9 YWILVAVGQMMSTMF 18 7 SWYWILVAVGQMMST 16 12 LVAVGQMMSTMFYPL 12 I FEDFAQSWYWILVAV 11 TableXLIX*V8-HLA-DRBl1*101 24P4C12 Each peptide is a portion of SEQ ID NO: 17; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptidle is the start position plus fourteen.

Pos 123456789012345 score 7 NYYWLPIMRNPITPT 24 5 HLNYYWLPIMRNPIT 18 6 LNYYWLPIMRNPITP 17 15 RNPITPTGHVFQISI 16 8 YYWLPIMRNPITPTG 13 21 TGHVFQTSILGAYVI 13 Tab~eXLiX-V9-HLA-DRBI1 101 -1 WO 2004/050828 PCTIUS2002/038264I 24P4C12 Each peptide is a portion of SEQ ID NO: 19; each start position is specified, the length of peptide is 15 amino acids, and the end position for each peptide is the start position plus fourteen.

Pos 123456789012345 score 4 CIAYWAMTALYPLPT 22 MTALYPLPTQPATLG 18 22 TLGYVLWASNISSPG 17 7 YWAMIALYPLPTQPA 14 13 LYPLPTQPATLOYVL 13 PATILGYVILWASNISS 12 23 LGYVLVYASNISSPGC 12 24 GYVLWASNISSPGCE 12 IAYWAMTALYPLPTQ I0 11 TALYPLPTQPATLGY WO 2004/050828 WO 204/00828PCTIUS2002/038264 Table L: Properties of 24P4C1 2 Bioinformatic URL Outcome Program ORF U1{I finder Protein length Transmernbrane region TM Prod H-MMfrop Sosui http:/ www.ch.embnet.org/ bttp://www.enzim.hu/hnuntop/ http:/www.genore.ad.jp/SOSuI 6 to 2138 710aa I I TM, 39-59, 86-104, 23 1-250, 252-273, 309- 330, 360-380, 457-474, 497-515, 559-581, 604- 626, 64t-663 IITM, 25-59 84-104231- 250 257-277 308-330 355- 377 456-475 500-5 19 550- 572 597-618 649-671 13TM, 34-65, 86-108, 145- 167, 225-247, 307-329, 357-379, 414-436, 447-469, 501-523,564-586, 600-622, 644-666 Signal Peptide p1 Molecular weight Localization TMH1M1M Signal P p1/MW tool p1/MW tool

PSORT

P SORT If httpj/www.cbs.dtudk/srvices/TMHMM http;//www.cbs.dtu.dk/services/SignalP/ http://www.expasy.chitools/ http://www.expasy.ch/tools/ http://psort.nibb.ac.jp/ http://psort.nibb.ac.jp/ http://www.sangerac.ukfPfarm/ http://www biochemn.uci.ac.uk/ http://www.blocks fhcrecrg/ IOTM, 36-5b,228-250, 252- 274, 308-330, 356-378, 454-476, 497-519, 559-581, 597-619 no 8.9 p1 79.3 kD 80% Plasma Membrane, Golgi 65% Plasma Memnbrane, 38% endoplasmic reticulum DUF580, uknown fuinction Anion exchanger fumily 313-359 CYS-RICH 536 547 Motifs Pfam Prints Blocks Pro-ite http://www.prosite.org/ Table LI. Exon compositions of 24P4C12 v.1 Exon number Start End Length 1 1 45 2 46 94 49 395 168 74 4 169 247 '79 248 347 100 6 348 473 126 7 474 534 61 8 535 622 88 9 623 706 64 707 942 236 11 943 1042 100 12 1043 113E 93 13 1136 1230 103 14 1239 1492 254 1493 1587 16 1588 1691 104 17 1692 1765 74 18 1766 1836 71 19 1837 1931 1932 2016 185 21 2072573 1557 WO 2004/050828 WO 2O4IOO82~PCT/US2002103826-t Table LII.

gaqccatggg acgacccctc tcctcttcct gagacccccg agaacaaaga acatcatctc cctqcccgga tcttctatac tcacaagcct ggegctgctt oaccataca ttaagatctt tgatgtctac actgggccat catccaacat cggcccacct ccaaaggcct tctggaccct ccttctactg tcatccgcac ttgtgcagat accctgtagc tt~argt t gtgtctcagc tggacaaagt gggtcctgtC gcccccacct tcgccagcgg tggaagacct ttctaaagat gacagctccg acttcgcctt tcacgcctgt tcgagaccag ccgagagtgg gcttgaaccc gggtgacaga tttqttaact Nuclaotide gggaaagcag ctttcgaggc gctcttcatt gcaagtcctc taagccgtat agttgctgag ggacccatgg aaaaaacagg gcaacaggaa tecatggacc gcaggggatc tgaagatttt catgttctac gactgctctg cagctccccc tgtgaactcc aatccaacgt taactggta ggccttccac actccgttac agcccgggtc ccgctgcatc crtaaaccgc caaaaatgcg cacagacctg cttctttcttt caactattac cttcttcagc gqagcggddC tctgggcadg gccctgatcc acagqtctcc aatccaacac cctggccaac tgqcatgcac ggqaggcaga ctctgtctcc caqtaaaaaa sequence of transcript variant 24P4Cl2 v.7 (SEQ ID NO: 94) cgggacqagg cccatcaaga ctaggttaca taccccagga ctcctgtact aacggcctac actgtgggaa aacttttgtc ctctgcccca aacgttactc agcggtctta gcccagtcct ccactggtca tacctggcta qgctgtqaga tcgtgercag tctgtcttca ctggccctgg aagccccagg cacactgggt atcttggagt atgtgctgLt aatgcataca ttcatgctac Ctgctqttct ttctccggtc tggctgccca gttttcggca aeCggCtCCC aagaacgagg aggactgcac attttgtggt tttgagaggc atggtgaaac Ctgtcatccc qgttgcagtq aaaacaaaac aaaaaaaaaa atgacqaggc acagaagctg tcgtggtggg actctactqg tcaacatctt agtgccccac aaaacgagtt tgccaggggt gtttcctcct caccggcgct ttgacagcct ggtattggat cctttgtcct cat cggggca aagtgcc-aat ggctgatgtg atctgcaaat gccaatgcgt acatccctac cattggcatt atattgacca tcaagtgctg tcatgatcgc tcatgcgaaa ttgggaagct gcatcccggg tcatgacctc cgtgtgtgqa tggaccggcc cgcccccgga cccaccccca aaaaaaaggt tgaggcgggc ctccgtctct agctactcgg agccgegatc aaacaaacaa ctacgggaag cacagatgtc gattgtggcc ggcctactgt cagctgcatc accccaqtg ct cacaga ct accctggaat cccctctgct cccagggatc caatgcccga tcttgtggct cctcctcatc accccagtat datacatca cgtcttccag c tatggggtc cctcgctgga cttcccetta tggagccctc caagctcaga cctctggtgt catctacggg cattgtcagg gctggtggtc gctgggtaaa catcctgggg cacgctcttc ctactacatg caacaagaag ccgtccagcc tttagqccag ggatcacctg attaaaaata gaggctgagg gcgccactgc aaagatttta ccagtcaaat atctgctgcg tggttgtatg ggcatggggg ctgtccagca tgtgtgtcct gttggggaag atgacggtga ccagctctgg accaatgaca gacatcagtg gtgggacaga tgcattgcct gtgctctggg tgcaacccca ggctactcat ctggggctct gcctttgcct atctctgcct atcctgaccc ggagtgcaga ctggaaaaat aagaatttct gtggtcgtcc ggaggcgtgg gactttaaga gcctatgtca ctctgcttcc tccaagagcc aggaagaagt atccaacctc gcgccgtggc agtcaggagt caaaaattag caggagaatc actccaacct ttaaagatat 120 180 240 300 360 420 480 540 600 650 720 780 840 900 960 1020 1080 1140 1200 1260 1320 1380 1440 1500 1560 1620 1680 1740 1800 1860 1920 1980 2040 2100 2160 2220 2251 Table LIII. Nuclaotida sequence alignment of 24P4C12v.1 v.1 (SEQ ID NO: 95) and 24P4C12 v.7 (SEQ ID NO: 96).

Score 135E bits (706; Expect 0.Oldantities 706/706 (100%) Strand Plus/ Plus 24P4C12v. 1: 1 gagccatggggggaaagCagcgggdcgdggatgacgaggcctacgggaagccagtcaaat 24P4C12v. 7: 1 gagccatggggggaaagcagcgggacgaggatgacgaggcctacgggaagccagtcaaat 24P4C12v. 1: 61 acgacccctcctttcgaggccccatcaagaacagaagctgcacagatgtcatctgctgcg 120 24P4C12v. 7: 61 acgacccctcctttcgaggccccatcaagaacagaagctgcacagatgtcatctgctgcq 120 24P4C12v. 1: 121 tcctcttcctgctcttcattctaggttacatcgtggtggggattgtggcctggttgtatg 180 24P4C12v. 7: 121 tcctcttcctgctcttcattctaggttacatgtggtggggattgtggcctggttgtatg 180 24P4C12v. 1: 181 gagacccccggcaagtcctctaccccaggaactctactggggcctactgtggcatggggg 240 24P4Cl2v. 7: 181 gagacccccggcaagtcctctaccccaggaacrctactggggcctactgtggcatggggg 240 24P4C12v. 1: 241 agaacaaaqataagccgtatctcctgtacttcaacatcttcagctgcatcctgtccagca 300 WO 2004/050828 PCT/US2002/038264 24P4C12v.7: 241 agaacaaagataagccgtatctcctgtacttcaacatcttcagctgcatcctgtccagca 24P4C12v.I1: 301 acatcatctcagttgctgagaacggcctacagtgccccacaccccaggtgtgtgtgtcct 360 24P4C12v.7: 301 acatcatctCagttgctgagaacggcctacagtgccccacaccccaggtgtgtgtgtcct 360 24P4C12v.1: 361 cctgcccggaggacccatggactgtgggaaaaaacgaqttctcacagac-tqttgqggaag 420 24P4C12v.7 361 cctgcccggaggacccatggactgtgggaaaaaacgagttctcacagactgttggggaag 420 24P4C12v. 1 421 tcttctatacaaaaaacaggaacttttgtctgccaggggtaccctggaatatgacggtga 480 24P4C12v.7: 421 tcttctatacaaaaaacaggaacttttgtctgccaggggtaccctggaatatgacggtga 490 24P4C12v.1: 481 tcacaagcctgcaacaggaactctgccccagtttcctcctcccctctgctccagctc-tcg 540 24P4C12v. 7: 481 tcacaaqcctgcaacaggaactctgccccagtttcctcctcccctctgctccagctctgg 540 24P4C12v. 1: 541 ggcqctgctttccatggaccaacgttactccaccggcgctcccagggatcaccaatgaca 600 24P4C12v.7: 541 ggcgctgctttccatggaccaacgttactccaccggcgctcccagggatcaccaatgaca 600 24 P4C12v.l1 601 ccaccatacagcaggqatcagcggtcttattgacagectcaatgcccgagacatcagtg 660 24P4C.12v.?: 601 ccaccatacagcaggqatcagcgqtcttattgacagcctcaatgcccgagacatcagtg 660 24P4C12v. 1: 661 ttaagatctttgaagattttgcccagtcctggtattggattcttgt 706 24P4C1[2v-7: 661 ttaagatctttgaagattttgcccagtcctggtattggattcttgt 706 Score 2971 bits (1545), Expect 0.Oldentizies 1545/1545 (100%) Strand PlusI Plus 24P4C12v. 1: 1043 egctgtgggacgatattctaccatttctacccactggtcacctttgtcctcctcct 1102 24P4C12v. 7: 707 ggctgtgggacagatgatgtctaccatgttctacccactggtcacctttgtcctcctcct 766 24P4C12v. 1: 1103 catctgcattgcctactgggccatgactgctctgtacctggetacatcggggcaacccca 1162 24P4C12v.7; 767 catCtgCattgCCtactgggCCatgaCtgCtctgtacctggCtacatcggggcaacccca 826 24P4C12v. 1: 1163 gtatgtgctctgggcatccaacatcagctcccccggctgtgagaaagtgccaataaatac 1222 24P4CI2v. 7: 827 gtatgtgctctgggcatccaacatcagctccccggctgtgagaaagtgccaataaatac 886 24P4C12v. 1: 1223 atcatgcaaccccacggcccaccttgtgaactcctcgtgcccagggctgatgtgcgtctt 1282 24P4CI2v. 7: 887 atcatgcaaccccacggcccaccttgtgaactcctcgtgcccagggctgatgtgcgtctt 946 24P4C127. 1: 1283 ccagggctactcatccaaaggcctaatccaacgttctgtcttcaatctgcaaatctatgg 1342 24P4C12v. 7: 947 ccagggctactcatccaaaggcctaatcaacgttctgtcttcaatctgcaaatctatgg 1006 24P4C12v. 1: 1343 ggtcctggggctcttctggacccttaactgggtactggcccgggccaatgcgtcctcgc 1402 24P4Cl2v. 7: 1007 ggtcctggggctcttctggacccttaactgggtactggccctgggccaatgcgtcctcgc 1066 WO 2004/050828 WO 2O4IOO82~PCT/US2002103826S 24 P4Cl2v. 1: 24PIC12v 7t 24 P4Cl2v. 1: 24 P4Cl2v.17: 24 P4Cl2v. 1: 24P4C12v.7: 24 P4C12v. 1: 24P4Cl2v.7: 24 P4 C2v. 1: 24P4Cl2v.7: 24P4C12v.1! 24P4C12v.7: 24 P4 12v.1: 24 P4Cl2v. 7: 24 P4Cl2v. 1: 24P4C12v.7: 24P4C12v.1: 24P4C12v.7: 24P'iCl2v. 1: 24P4Cl2v.7: 24 P4C12v. 1: 24P4Cl2v.7: 24P4C12v.1: 24P4Cl2v.7: 24 P4Cl2v. 1: 24P4C12v.7: 24P4Cl2v.l: 24 P4Cl2v. 7: 1403 tgacttctcttcggctcaagccgectct~~tc 1462 1067 tggagcctttguctccttctactgggccttccacaagccccaggacatccctaccttcc 1126 1463 cttaatctctgccttcatccgcacactccgttaccacactgggtcattggcatttggagc 1522 1127) cttaatctctgcctteatccgcacactccgttaccacactgggtcattggcctttggagc 1186 1523 cctcatcctgacccttgtgcagatagcccgggtcatcttggagtatattgaccacaagct 1582 1187 ccLcatcctgaccctttcagatagccqggtcatcttggagtatattgaccacaagct 1246 1583 cagaggagtgcagaaocctgtagcccgctgcatcatgtgctgtttcaagtgctgcctctg 1642 1247 cagaggagtgcagaaccctgtagcccgctgcatcatgtgctgtttcaagtgctgcctctg 1306 1643 gttctggaaaaatttatcaagttcctaaaccgcaatgcatacatcatgatcgccatcta 1702 1307 gtgtctggaaaaatttatcaagttcctaaaccgcaatgcatacatcatgatcgccatcta 1366 1733 cgggaagaatttctgtgtctcagccaaaaatgcgttcatgctactcatgcgaaacattgt 1762 1367 cgggaaqaatttctgtgtctcagccaaaaatgCqttcatgCt~tatqCtgaadCattgt 1426 1763 cagggtggtcgtcctggacaaagtcacagacctgctgctgctctttgggaagctgctggt 1822 1427 cagggtggtcgtcctggacaaagtcacagacctgctgctgrtctttgggaagctgctggt 148b 1823 ggtcggagcgtgggggtcctgtccttcttttttttctccggtcgcatcccgqggctqgg 1882 1497 ggtcggaggcgtgggggtcctgtccttcttttttttctcccgqtcgcatcccggggctggg 1546 1883 taaagactttaagagcccccacctcaactattactggctgcccatcatgacctccatcct 1942 1547 taaagactttaagagccccczacctcaactattactggctgcccatcatgacctccatcct 1606 1943 gggggcctatgtcatcgccagcggcttcttcagcgttttcggcatgtgtgtggacacgct 2002 1607 gggggcctatgtcatcgccagcggcttcttcagcgttttcggcatgtgtgtggacacgct 1666 2003 cttcctctgcttcctggaagacctgjgagcggaacaacggctccctggaccggccctacta 2062 1667 cttcctctgcttcctggaagacctggagcggaacaacggctccctggaccggccctacta 1726 2063 cattccaaaccttctaaagattctgggcaagaagaacgaggcgcccccggacaacaa 2122 1727 catgtccaagagccttctaaagattctgggcaagaagaacgaggcgcccccggacaac-aa 2786 2123 gaagaggaagaagtgacagctccggccctgatccaggactgcaccccacccccaccgtcc 2182 1797 gaagaggaagaagtgaca~gctccggccctgatccaggactgcaccccacccccaccgtcc 1846 2183 agccatccaacctcacttcgccttacaggtctccattttgtggtaaaaaaaggttttagg 2242 1847 agccatccaacctcacttcgccttacaggtctccattttgtggtaaaaaaaggttttagg 1906 WO 2004/050828 WO 204100828PCTIUS2002/038264L 24E'4C12v. 1: 2243 ccaggcgccgtggctcacqcctgtaatccaacactttgagaggotgngqcgq~qgatca 2302 24P4012v. 7: 1907 ccagqcgccggctcecgccrgtsatccsacactstgagaggcsgagqcgggcggatca 196 24P4C12v. 1: 2303 cctgagtcaggagttcgagaccagcctggccaacetggtgaaaccrccgsctctattaaa 2362 24P4C12v. 7: 1967 cctgagtcaggagtztogagaccagcctggccaacstggtgaaaccccqtctotattnaaa 2026 24P4C12v.1: 2363 eatacaaaeattegccgagagcggtggjcatgoacctgtcatcccagctactcgggaggcs 2422 24P4012v.7: 2027 EaLuciddEattegccgagagtggtqgcatgoacctgtcatcccagctactcgggaggct 2086 24r'4C12v. 1: 2423 geggcaqqagascgcttgaecccqqqaggcagaggttgcagtgagccgagatcqcgcca 2482 2424C12v.7: 20687 gaggcaqgageatcgctcgeacccgggaggcagaggttgcagtgagccgagatogcgcca 2146 24P4C12v. 1: 2483 coqcactccaacccgggtgacagactcgsctccaaaacaaaacaaacaaacaaaaagat 2542 24P4C12v. 7: 2147 ctgcactccaacctggqtqacagactcsgtcccaaaacaaaacaaacaaacaaaaagat 2206 2424012v. 1! 2543 tttattaaagatattttgttaactcagtaaaaaaaaaaaaaaaaa 2587 24P4C12v. 7 2207 tcattaaagatattttgscaactcagtaaaaaaaaaaaaaaaaa 2251 Table LIV. Peptide sequences of protein coded by 24P4c12 v.7 (SEQ ID NO: 97) *IGGKQRDEDD EAYGI PVKYD PSFRGPIKNR SOTOVICCVL FLLFILGYIV VGIVAWLYOD PRQVLYPRNiS TGAYCGNGE4 KDKFYLLYWN ZFSCILSSN] ISVASNGLQC PTPQVCVSSC 120 PEDPWTVGKN EFSQTVCEVF YTKNRNFCLP GVPWIIMTVIT SLQQELCPSF LLPSAPALGR 180 CFPWTNVTPP ALPCITNDTF IQQGISGLID SLNAADISV< IFEDE'AQSWY WILVAVGQMM 240 STMFYPLVTF VLLLICIAYW AD4TALYLATS GQPQYVLWAS NISSPGCEKV PINTSCUPTA 300 HLVNSSCPGL MCVFQGYSSK GLIQRSVFNL QIYCVLGLFW TLNWVLALGQ CVLAGAFASF 360 YWAFHKFQDI PTFPLISAFI RTLRY-TGSL AFGALILTLV QIARVILEYI DHKLRGVQNP 420 VARCIMCCFK CCLWCLEKFI 1 FLMRNAYIM IAIYCKNECV SD&NAFMLLM RNIVRVVVLD 400 KVTOLLLFFG KLLWVGGVGV LSFFFFSGRI PGLGKDFXS? HLNYYWLPI4 TSILGAYVIA 540 SGFFSVF3MC VOTLWLCELE DLERNNGSLD RPYYMSKSLL KITJGKKNEAP PDNKKRKX 598 Table LV. Amino acid sequence alignment of 24P4C12v. 1 v.1I (SEQ ED) NO: 98) and 24P4C12 v. 7 (SEQ ]ID NO: 99).

Score 1195 bits (3091), Expect =0.Ol~dentlties 598/-710 Positives 598/710 Gaps 112/710 24P4C12v 1 MGGKQROED0EAYGKPVKYDPSFRGPIKbNRSCTDV1CCVLFLLFTLGY1VVG1VANLYGD

MGGKQRDEDDEAYGKPVKYDPSFPPKRSCTOVICCVLFLLFILGYIVVGIVAWLYGD

24E4C12v. 7: 1 MGGRQRDEDDEAYGKVYDPSFRGPIKNRSCTDVICCVLFLLFILGYIVVGIVAWLYGD 24P4C12v. 1: 61 PRQVLYVRNSTGAYCGGENDKPYILYFNIFSCILSSNIISVAENGLQCPTPQVCVSSC 120 PRQVLYPRI4STGAYCGMGENKDKPYLLYFNFSCILSSNIISVAEHGLQCPTPQVCVSSC 24P4C12v. 7: 61 PRQVLYPRNSTGAYCG%GEKDKPYLLYFNISCILSSNIISVAENGLQCPTPQVCVSSC 120 24p4C12v. 1: 121 PEDPWTVGKNEFSQTVCEVFYTKNRNFCLPGVPWNMTVITSLQQELCPSFLLPSAPALCR 180 PEDPWrVGKNEFSQVVGEVrYTIOPNFCLPGvPNNMTVITSLQQELCPS £LLPSAPALGR 24P4C12v. 7: 121 PEDFWTVGENEFSQTVGEVFYTKRRNFCLPGVWNITVITSLCQELcFSFLLPSAPALGR 180 2494012v. 1: 181 CFPWTNVTPPALPGITNDTTIQQGISGLIDSL4ARDISVKFEDFAQSWYWILVALGVAL 240 CFPWTNVTPPALPGITNDTTIQQGISGLIDSLNAHDI SVKIFEDFAQSWYWILVA 24P4CI2v. 7: 181 CFPWTNVTPPALFGITNu)TT1QQcGISGL1DSLWARflISVKIFEDFAQSWYWILVA--235 24p4C12v. 1: 241 VLSLLFILLLRLVAGPLVLVLILGVLGVLAYGIYYCWEEYRVLRDKGASISQLGFTTNLS 300 WO 2004/050828 24P4C12v.7: PCT/US2002/038264 235 24P4CI2v.1: 301 24P4C12v.7: 236 24P4C12v. 1: 24P4C12v. 7 24P4C12v.1: 421 24P4Cl2v.7: 309 24P4CI2v. 1: 24F4C12v. 7: AYQSVQETWIALIVIAVLEAILLLMLI FLRQRIRIAIALLKEASAVGQM!MSTMFYPLV 360 VGQMI4STMy2YPLV VGQMFSTb4FYPLV 248 TVLLLICIAYWATALYLATSGQPQYVLWASNISSPGCEKVPINTSCPTA.1LVNSSCP 420 TFLLCAWMTLLTGPY WSISG2VINSMTHVSCP TFVLLLbCIAYWAMTALYLATSGQPQYWASNISSPGCTPINTSCPTAJ1LVNS3qCP 308 GL!ACVFQGYSSKGLIQRSVFNL0IYGLGLFWTNWVLALGQCVLAGAFASFYWAFIKFQ 480 GLMCVFQGYSSKGLIQRSVFNLQIYGVLGLFWTLNJWVLALGQCVLAGAF'AS FYWAFHKPQ GLMCVFQGYS SKGLIQRS VFNLOI YGVLGLFWTLNWVLALGQCVLAGA3SFYWAFHKPQ 368 DIPTFPLTSAFIRTLRYHTGSLAFGALILTLVQIARVLEYIDHK1RGVQNPVARCIMCC 540 DIPTFPLISAFIRTLRYHTGSLAWFGALILTLVQIARVILEYIDHKLRGVQN9VARCIMCC DIPTFPLISAFIRTLRYHTGSLAFGALILTLVQIARVILEYIDHLRGVQNPVAflCIMCC 428 FKCCLWCLEKFIKFLNRNAYIMIAIYGKNFCVSAKNAFMLLMR4IVRVVVLDKVTDLLLF 600 FKCCLWCLEKFIKFLNtU4AYIMIAilYGNFCVSAKNAFMLLMRIVRVVVLDKVTDLLLF FKCCLWiCLEKFIKFLNRNAYIMIAIYGKNFCVSAKNAFMLLMRI4IVRVVVLDKVTDLLLF 488 FGKLLVVGGVGVLSFFFFSGP.IPGLGKDFKSPHLNYYWLPIMTSILGAYVIASGFFSVFG 660

FGKLLVGGVGVLSFFFFSGRIPCLGKDFKSPHLNYYWLPIMTSILGAYVIASGFFSVFG

FGKLLVGGVGVLSFFFSGRE'GLGKDFKSPHLNYYWLPIMTSILGAYVIASGFFSVF0 548 MCVDTLFLCFED~LERNNGSLDRYYMSKSLLKILGKKNEAPN(E(RKK 710

MCVDTLFLCFLEDLERNNGSLDRPYYSKSLLKILGK(NEAPPDNKKJCF(

MCVDTLFLCFLEDLERNNGSLDPYYMSSLLKILGKIC3IEAPPDIIKKy 598 24P4C12v.1: 541 24P4C12v.7: 429 2424C12v.1: 601 24P4C12v.7: 489 24P4C12v.1: 661 2424C12v.7t 549 Table LVI. Nucl~otide sequence of transcript variant 24P4C12 v.8 (SEQ ID NO: 100) gagccatggg gggaaagcag cgggacgagg atgacgagqc ctacgaag ccagtcaaat acgacccctc ctttcgaggc cccatcaaga acagaagctg cacagatgtc atctgctgcg 120 tcctcttCct gctcttcatt ctaggttaca tcgtggtggg gattgtggcc tggttgtatg 180 gagacccccg gcaagtcctc taccccagga actctactgg ggcctactgt qgcatggggq 240 aqaacaaaga taagccgtat ctcctgtact tcaacatctt cagctgcatc ctgtccagca 300 acatcatctc agttgctgag aacggcctac agtgccccac accccaggtg tgtgtgtcct 360 cctqcccgga ggacccatgg actgtgggaa aaaacgagtt ctoacagact qttggggaag 420 tcttctatac aaaaaacagg aacttttgtc tgccaggggt accctqgaat atgacqgtga 480 tcacaa'gcct gcaacaggaa ctctgcccca gtttcctcct cccctctgct ccagctctgg 540 ggcgctgctt tccatggacc aacgttactc caccggcgct cccagggatc accaatgaca 600 ccaccataca gcaggggatc agcggtctta ttgacagcct caatgcccga gacatcagtg 660 ttaagatctt tgaagatttt gcccagtcct ggtattggat tcttgttgcc ctgggggtgg 720 ctctggtctt gaqcctactg tttatcttgc ttctgcgcct ggtggctggg cccctggtgc 780 tggtgctgat cctgggagtg ctgggcgtgc tgqcatacgg catctactac tgctgggagg 840 agtaccgagt gctgcgggac aagggcgcct ccatctccca gctgggtttc accaccaacc 900 tcagtgccta ccagagcqtg caggagacct ggctggccgc cctgatcgtg ttggcggtgc 960 ttgaagccat cctgctgctg atgctcatct tcctgcggca gcggattcgt attgccatcg 1020 ccctcctgaa ggaggccagc aaggctgtgg gauagatgat gtctaccatg ttctacccac 1080 tggtcacctt tgtcctcctc ctcatctgca ttgcctactg ggccatgact gctctgtacc 1140 tggctacatc ggggcaaccc cagtatgtgc tctgggcatc caacatcagc tcccccggct 1200 gtgaqaaagt gccaataaat acatcatgca accccacggc ccaccttgtg aactcctcgt 1260 gcccagggct gatgtgcgtc ttccagggct actcatccaa aggcctaatc caacgttctg 1320 tcttcaatCt gcaaatctat ggggtcctgg ggctcttctg gacccttaac tgggtactgg 1380 ccctgggcca atgcgtcctc gctgqagcct ttgcctcctt ctactgggcc ttccacaagc 1440 cccaggacat ccctaccttc cccttaatct ctgccttcat ccgcacactc cgttaccaca 1500 ctgggtcatt ggcatttgga gccctcatcc tgacccttgt gcagatagcc cgggtcatct 1S60 tggagtatat tgaccacaag ctcagaggag tgcagaaccc tgtagcccgc tgcatcatgt 1620 gctgtttcaa gtgctgcctc tggtgtctgg aaaaatttat caagttccta aaccgcaatg 1680 catacatcat gatcgccat2 tacgggaaga atttctgtgt ctcagccaaa aatgcgttca 1740 tgctactcat gcgaaacatt gtcagggtgg tcgtcctgga caaagtcaca gacctgctgc 1800 tgttctttcg gaagctgctg gtggtcggag gcgtgggggt cctgtccttc ttttttttct 1860 ccggtcgcat cccggggctg ggtaaagact ttaagagCcc ccacctcaac tattactggc 1920 tgcccatcat gaggaaccca ataaceuuaa cgggtcatgt ctliccagacc tccatcctgg 1980 gggcctatqt catcgccagc ggcttcttca gcgttttcgg catgtgtgtg gacacgctct 2040 tccI-ctgctt cctggaagac ctggagcgga acaacggctc ccr-ggaccgg ccctactaca 2100 tgtccaagag ccttctaaag attctgggca agaagaacga ggcgcccccg gacaacaaga 2160 agaggaagaa gtgacagctc cggccctgat ccaggactgc accccacccc caccgtccag 2220 WO 20041050828 ccatccaacc tcacttcgcc ttacaggtct czattttgtg qtaaaaaeqg gttttaggcc aqqcgccgtg gctcacgcct gtaatccaac actttgagag gctgaggcgg gcggatcacc tgagtcagg gttcgagacc agcctggcca acatggtgaa acctccgtct ctattaaaaa tacaaaaatt aqzcgagaqt gqtgqcatgc acctgtcatc ccagctactc gggaggctga ggcaggagaa tcgcttgaac ccgggaggca gaggttgcag tgagccgaga tcgcgccact gcactccaac ctgggtgaca gactctgtct ccaaaacaaa acaaacaaac aaaaagattt tattaaagat attttgttaa ctcagtaaaa aaaaaaaaaa aaa PCT/US20021038264 2280 2340 2400 2460 2520 2580 2623 Table LVII. Nucleotide sequence alignment of 24P4C12v.. v.1. (SEQ ID NO: 101) and 24P4C12 v.8 (SEQ IM NO: 102) Score 3715 bits (1932), Expect 0.01dentities 193211932 (100%) Strand Plus/ Plus 24P4C12v.l: I 24P4Cl2v.8: 1 24P4C12v.1: 61 24P4Cl2v.8: 61 24P4C12v.l: 121 24P4012v.8: 121 24P4Cl2v.2.: 181 24P4C12v,8: 181 24P4C12v.1: 241 24P4C12v.8: 241 2424C12v.l: 301 24P4C12v.8: 301 24P4C12v.1l: 361 24P4C12v.8: 361 24P4C12v.l: 421 24P4C12v.8: 421 24P4C12v.1: 481 24P4C12v.6: 481 24P4C12v.l: 541 24P4C12v.8: 541 24P4C2.2v.1: 601 24P4Cl2v.8: 601 gagccatggggggaaagcagcggecgaggatgacgaggcctacgggaagccagtcaaatI gagccatqgggggaaagcagcgggacgaggatgacgaggcctacgggaagccaqtcaaat acgacccctccttcaggcccatcaagaacagadqctgcacagatgtcatctgct9gg acgacccctcctttcgaggcccr atcaagaa-cagaagctgcacagatgtcatct-gctgcg tcctcttcctgctcttcattctaggttacatcgtggtggggattgtggcctggttgtatg tcctcttcctgctcttcattctaggttacatcgtgqtgggqattgtggcctggttgtatg gagacccccgaagtcctctacccaggaactctactggggcctactgtggcatggggg gagacccccggcaagtcctctaccccaggaactctactggggcrtactgtggcatggggg agaacaaagataagccgtatctcctgtacttcaacatCttcagctgcatCCtgtccagca agaacaaagataagccgtatCtcctgtdcttCaaeatcttcagctgcatcctgtccagca acatcatctcagttgctgagaacggcctacagtgccccacaccCaggtgtgtgtgtcct acatcatctcagttgctgagacggcctacagtgccccacaccccaggtgtgtgtgtcct cctgcccggaggacccatggactgtgggaaaaaacqaqttctcacagactgttggggaag cctgeccggaggacccatggactgtgggaaaaaacgagttctcacagactgttggggaag tcttctatdcaaaaaacaggeacttttgtctgcagggtaccctggaatatgacggtga tcacaagcctgcaacaggaactctgccccagtttcctcctcccctctgctc~agctCtgg tcacaagcctgcaacaggaactctgccccagtttcctcctcccctctgctccagctCtgg ggcgctgctttccatqpacca8.CqttactCCaccgcqtcccaggatCaccaatg&a ggcgctgctttccatggaccaacgttactccaccggcgctcccagggatcaccaatgaca ccaccatacagcaggggatcagcggtcttattgacagcctraatgcOccagacatcagtg ccaccatacagcaggggatcagcggtct tattqacagcctceatgcccgagacatcagtg WO 2004/050828 WO 204/00828PCTIUS2002/038264 24P4Cl2v.1:* 24P4C12v.8; 24P4C12v. 1: 24P4Cl2v.8: 24P4C12v.1: 24P4Cl2v .8: 24P4 Cl2v 1: 24P4C12v. 8: 24P4C12v. 1: 24P4C12v. 8: 24 P4Cl2v. 1: 24P4C12v. 8: 24P4C12v.l1: 24P4C12v. 8: 24P4C12v. 1; 24P4C12v. 8: 24P4C12v. 1: 24P4C12v. 8: 24P4C12v. 1: 24P4C12v. 8: 24P4C12v.1: 24P4Cl2v. 8: 24P4C12v. 1: 24P4Cl2v.O8: 24P4Cl2v. 1: 24P4Cl2v.B: 661 661 721 721 781 781 841 841 901 901 961.

961 1021 1021 1001 1081 1141 1141 1.201 1201 1261 1261 1321 1321 1381 1381 ttaagatctttgaagattttgcccagtcctggtattggLLcttgttgccctgggggtgg ttaagatctttgaagattttgcccagtcctgta~ttggattcttgttgccctggqggtgg ctctggccttgagcctactgtttatcttgcttctgcgcctggtggctgggccctggtgc ctctggtcttgagcctactgtttatcttgcttc2tgcgcctggtggctgggcccctggtgc tggtgctgatcctgggagtgctgggcgtgctggcatacggcatctactactgctgggagg tggtgctgatcctgggagtgctgggcgqctggcatacggcatctactactgctgggagg agtaccgagtgctgcgggacaagggcgcctccatctcccagctgggtttcaccaccaacc agtaccgagtgctgcgggacaagggcgcctccatctcccagctgggtttcaccaccaacc tcagtgcctaccagagcgtgcaggagacctqctggccgGcctgatCgtgttggCggtgc tcagtgcctaccagagcgtgcaggagacctggctggccgccctgatcgtgttggcggtgc ttgaagccatcctgctgctgatgctcatcttcctgCggcagcggattcgtattgccatcg ttgaagccatcctgctgctgatgctCatcttcctqcgqtzagcggattcgtattgccatcg ccctcctgaaggaggccagcaaggctgtgggacagatgatgtctaccatgttctGcccac ccctcctgaaggaggccagcaaggctgtgggacagatgatgtcta~catgttctacccac tggtcacctttgtcctcctcctcotctgcattgcctactgggccatgactgctctgtacc tggtcacctttgtcctcctcctcatctgcatitgcctactgggccatgactgctctgtacc tggctacatcggggcaacccagtatgrtgctctgggcatccaacatcagctCCCCCggc tggctacatcggggcaaccccagtatgtgctctgggcatccaacatcagcCCccggct gtgagaaagtgccaatdaatacatcdtgcaaccccacqgcccaccttgtgaactcctcgt gtgagaaagtgccaataaatacatcatgcaaccccacggcccaccttgtgaaCtcCtCgt gcccagggctgatgtgcgtcttccaggctactCatccaaaggcctaatcc!aacgttctg gcccagggctgatgtgugtcttccaggctactcatccaagcctaatcC8acgttctg tcttcaatctgcaaattatggggtcctggggctcttctggacccttadctggtactgg tcttcaatctgcaaatctatggggtcctggggctCttctggacccttaactgggtactgg ccctgggccaaigcgtcctcgctgagcctttcctccttctactgggccttccacaagc ccctgggccaatgcgtcctcgctggagcctttgCtccttctactgggccttccacaagc 720 720 780 780 840 840 900 900 960 960 1020 1020 1080 1080 1140 1140 1200 1200 1260 1260 1320 1320 1380 1380 1440 1440 24P4C12v.1: 1441 24P4C12v.8: 1441 cccaggacatccctaccttc~ccttaatctctgccttcatccgcacactccgttaccaca 1500 cccaggacatccctaczttccccttaatctctgccttcatccgcacactccgttaccaca 1500 WO 2004/050828 WO 204/00828PCT/US2002JO38264 24P4Cl2v. 1: 1501 ctgggtcattgcatttggaqccctcatcctgaeccttgtgcagatagcccgggtcatct 1560 24P4C12v.8: 1501 ctgggtcattggcatttggaqccctcatcctgacccttgtgcagataqcccgqgtcatct 1560 24P4Cl2v. 1: 1561 tggagtatattgaccacaagctcagaggagtgcagaaccctgtagcccgctgcatcatgt 1620 24P4C12v. 8: 1551 tggagtatattgaccaceaqctcagaqagtqcagaaccctgtagcccgctgcatcatgt 1620 24P4Cl2v.1: 1621 gctgtttcaagtgctgcctctggtgtctggaaaaatttatcaagttccteaaccgcaatg 1680 24P4Cl2v.8: 1621 gctctttcaagtgctgcctctggtgtctggaaaaatttatcaagttcctaaaccgcaatg 1680 24P4C12v.1: 1631 catacatcatgatcgccatctacgggaagaatttctgtgtctcagccaaaaatgcgttca 1740 24P4C12v.8. 1681 catacatcatgatcgccatctacgggaagaatttctgtgtctcagccaaaaatgcgttca 1740 24P4Cl2v.1: 1741 tqctactcatgcgaaacattgtcagggtggtcgtcctggacaaagtcacagacctgctgc 1800 24P4C12v.8: 1741 tqctactcatgcgaaacattgtcagggtggtcgtcctggacaaagtcacagacctgctqc 1800 24P4C12v. 1: 1801 tgttctttgggaagctgctggtggtcggaggcgtgggggtcctgtcrntcttttttttct 1860 24P4C12v.8: 1801 tgttctttgggaagctgctggtggtcggagqcgtgqgggtcctgtccttcttttttttct 1860 24P4C12v. 1: 1061 ccggtcgcatcccggggctggaaagacctttaagagcccccacctcaactattactggc 1920 24P4CI2v.8; 1861 ccggtcgcatd ccggggctgggtaaqactttaagagcccccacctcaactattactggc 1920 24P4CI2v.I: L921 tgcccatcatga 1932 111FF 111111] 24P4C12v.8: 1921 tgcccatcatga 1932 Score 1263 b~its (657), Expect 0.Otdentities 6571657 (100%) Strand Plus/ Plus 24P4C12v. 1 1931 gacc catrctgggccctatgtcatcgccagcggcttcttcagcgttttcggcatgtg 1990 24P4C12v.8: 1967 gacctccatcctgggggcctatgtcatcgccagcggcttcttcagcgttttcggcatgtg 2026 24P4C12v. 1: 1991 tgtggacacgctcttcctctgcttcctqaagacctgagcggaacaacggctccctgga 2050 24P4C12v. 8: 2027 tgtggacacgctcttcctctgcttcctggaagacctggagcggaacaacggctccctgga 2086 24P4C12v. 1: 2051 ccggccctactacatgtccaagagccttctaaagattctgggcaagaagaacgaggcgcc 2110 24P4C12v. 8: 2087 ccggccctactacatgtccaagagccttctaaagattctgggcaagaagaacgaggcgcc 2146 24P4C12v. 1: 2111 cccggacaacaagaagaggaagaagtgacagctccggccctgatccaggactgcacccca 2170 24P4C12v. 8: 2147 cccggacaacaagaagaggaagaagtgacagctccggccctgatccaggactgcacccca 2206 24P4C12v. 1: 2171 cccccaccgtccagccatccaacctzcacttcgccttacaggtctccattttgtggtaaaa 2230 24P4C12v. 8: 2207 cccccaccgtccagccatccaacctcacttcgccttacaggtctccattttgtggtaaaa 2266 24P4C12v. 1: 2231 aaaggttttaggccaggcgccgtggctc acgcctgtaatccaacactttgagaqgctgag 2290 WO 2004/05 0828 PCT/US2002/038264 111111111M1 IHIM 1111 (1111111111111 11 III 1111 IIIM 24P4C12v.8: 2267 aagttagcqccggcccgcgatcaattaag~a 2226 24P4C12v,.1: 2291 qcggcggatcacctgagtcaggagttcgagaccagcctggccaacatggtgaaacctcc 2350 24P4C12v.8: 2227 gcgggcgqatcacctgagtcaggagttcgagaccagccrggccaacarggtgaaaccccc 2.386 24P4C12v.1: 2251 gtctctatcaaaaatacaaaaattagcc~agagtggtggcatgcacctgtcatcccagct 2410 24P4C12v. 8: 2387 gtctctattaaaaatacaaaaattagccgagaqtggtggcatqcacctgtcatcccadgct 2446 24P4C12v.1: 2411 actcgggaggctgagocaggagaatcgcttgaacccgggaggcagaggtsgcagtgagcc 2470 24P4012v. 8: 2447 actcgggaqgctgaggcaggagaatcgcttgaacccgggaqgcagaggttgcagtgagcc 2506 24P4C12v.1: 2471 gagatcgcgccactgcactccaacctgggtgacagactctgtccccaaaacaaaacaaac 2530 24P4C12v.8: 2507 gagatcgcgccactgcactccaacctggqtgacaqactctgtctccaaaacaaaacaaac 2566 24P4C12v.1: 2531 aaacaaaaagattttattaaagatattttqtoaacrcagraaaaaaaaaaaaaaaaa 2587 24P4C12v. 8: 2567 aaacaaaaaqattttattaaqatattttgttaactcagtaaaaaaaaaaaaaaaaa 2623 Table LVIII. Peptide, sequences of protein coded by 24P4C12 v.0 (SEQ MGGKQRDBDE EAYGKPV Y FSFRGPIKUR SCTIDVTCCVL FLLEILGYIV VGIVAWLYGD PRQVLYPRNS TGAYCGNGEN KDKPYLLYFN IFSCILSSNI ISVAENGLQC PTPQVCVSSC PEDPWTVGKN EFSQTVGEVF YTKNRNFCLP GVPWNMTVIT SLQQELCPSF LLPSAPALGR CFPWTNVT'P ALPGITNDTT IQQGISGLID SLNAROISVK IFEDFA2SWY WILVALGVAL 'JLSILFILLL RLVAGPLVLV LILGVLGVLA YGIYYCWEEY RVLROKGASI SQLSFTTLS AYQSVQETWL AALIVLAVLE AILLLMLIFL RQRIEIAIAL LKEASKAVGQ MM1STMFYPLV TFVLLLICIA YWAIITALYLA TSGQPQYVLW ASNISSPGCE KVPINTSCNP TAHLVNSSCP GLMCVFQGYS SKGLIQRSVF NLQIYGVLGL FWTLNWVLAL GQCVLAGAFA SFYWAFHEPQ DIFTFFLISA FIR PLRYHTG SIAFSALILT LVQIZ\RVILE YIDHICLRGVQ NPVARCINCC FKCCLWCLE( FIKFLWRNAY IMIATYCKNE CVSAKNAFML LMRNIVRVVV LDKVTDLLLF BGKLLVVGGV GVLSFFFFSG RIPGLG'CFK SPH-LNYflWLP IMRNPITPTG HiVFOTSILGA YVTASGFFSV FGMCVDTLFL CFLEDLERNN GSLDRPYYMS KSLLKILGKK NSAPPDNRKR

KK

ID HO: 103) 120 180 240 300 360 420 480 540 600 660 720 '722 Table LIX. Amino acid sequence alignment of 24P4012v. 1 v.1I (SEQ 11) NO: 104) and 24P4C12 v.8 (SEQIDNO: 105) Score 1438 bits (3722), Expect 0.OcIdentities 710/722 Positives 710/722 Gaps 12/722 24P4C12v. 1: 1 IGGKQRUEDDEAYGKPVKYDPSFRGPIKNRSCTDVICCVLFLLFILGYIVVGIVAWLYGD MGGKQRIDEDDEAYGKPVKYDFSFRGPIKNRSCTDVICCVLFLLFILGYIVVGIVAWT YGD 24P4C12tu8: 1 MGGI(QRDEDDEAYGKPVRYDPSFRGPIKNRSCTDVICCVLFLLFILGYIVVGIVA41Y0D 24P4C12v. 1: 61 PRQVLYPRNSTGAYCGMGENKDKPYLLYFNIFSCILSSNIISVAENGLOCPTPQVCVSSC PRQVLYPRNSTGAYCGMGENKDKPYLLYFNI FSCILSSIISVAEWGLQCPTPQVCVSSC 24P4C12v. 8: 61 PRQVLYPRNSTGAYCGMGENKDRPYLLYFUXFSCILSSMIISVAENGLQCPTPQVcVSSc 24P4C12v. 1: 121 PEDPWPVGRNEFSQTVGEVFYTKNRNFOLPGVPWNMTVITSLQQELCPSFLLPSAPALGR PEDPWPVGKNEFS QPVGEVFYTKNPNFCLPGVPWNMTVITSLQQELG PS FLLPSAPALGR 24P4C12v. 8: 121 PEDPWPVGKNEFSQ'PVGEVFYTINRNFCLPGVPWNMTVITSLQOELOFSFLLPSAFALG 24P4C12v.1: 181 CFPWTNVTPPALPGITNDTTIQQGISGLIDSLNARDISVKIFEDFAQSWYILVALGVAxL CFPWTNVTPPIUJFGITNDTTIQQGISGLIDSLNRDISVKIFEDFAQSWYWgILVALGVAL 2424012v.8: 181 CFPWTNVTPPALPGITNDTTIQQGISGLIDSLNARJTSVKIFEDFAQSWYWtILVALGVA 24P4012v. 1: 241 VLSLLFILLLRLVAGPLVLVLILGVLGVLAYGIYYCWEEYRVLRDKGASIsQLGFTTNLS9 VLSLLFILLLRLVAGPLVINLILGVLGVLAYGIYYCWEEYRVLRDKGAS ISQLGFTTMLS WO 2004/050828 WO 204/00828PCTIUS2002/038264 24P4CL2v.8: 241 VLSLLFILLLRLVAGPLVLVLILGVLGVLAYGIYYCWEEYRVLRDKGASISQLGFTTNLs 300 24P4C12v.1: 24P4Cl2v.8: 24P4Cl2v. 1: 24P4Cl2v.@: 24P4C12v. 1: 24P4CI2v.8: 24P4C12v. 1: 24F4012v.8; 301 AYQSVQETWLAALIVLBVLEAILLLM1LIFLRQRI1RIAIALLKEASKAVGQmNSTMFYPLv 360 AYQSVQErWLAALTVTAkVIEAI LLLMILI FLRQRIRIAIALLKEASKAVGO4MSTMFYP'LV 301 AYQSVQETWLAALIVLAVLEAILLLMIFLRQRIRIAIALLKEASKAVGQMMSTMFYPLV 360 361 TFVLLLICIAYWAMTALYLATSGQPQYVLWASNISSPGCEKVEINTSCNPTAHLVNSSCP 420 TFVLLLICIAYWAMTALYLATSGQPQYVLt4ASM4ISS PGCEKVPINTSCNPTAHLVNSSCP 361 TFVLLLICIAYWAMI'ALYLATSGQPQYVLWASISSPGCEKV9INTSCNPTAHLVNSSCP 420 421 GLMCVFQGYSSKGLIQRSVFNLQIYGVLGLFWTLNWVLALGQCVLAGAFASFYWAFHKPQ 480 GLMCVFQGYS SKGL rQRSVFNLQ]YGVLGLFWTLNWVLALGQCVLAGAFASFYWAFHKPQ 421 GLMCVFQ3YSSKGLIQRSVFNLQIYCVLGLFWTLNWVLALGQCVIAGAFASFYWAFHKPQ 480 481 481 24P4C12v.1: 541 24P4C12v.8: 541 24P4Cl2v, 1: 24P4Cl2v. 8: DIPTFELISAFIRTLRYHTGSLAFGALILTLVQIARVILEYID)HKLRGVQNFVARCIMCC 540

DIPTFPLISAFIRTLRYHTGSLAFGALILTLVQIARVILEYIIDHKLRGVQNPVARCIMCC

DIPTFE'LI5AFIRTLRYHTGSLAFGALILTLVQIARVILEYID)HKLRGVNPVARIM4CC 540 FKCCLWCLEKFIKFLNRNAYIMIAIYGKNFCVSPI(NA'MLLMRN IVRVVVLDKVT LLLF 600 FKCCLWCLEKFIKFLNRNAYIMIAYGKNFCVSP.KNAFMLtMRNIVRVVVLDKVTDLLLF FKCCLWCLEKFIKFLNRNAYIMIAIYGKNFCVSnKNAFMLLMRN IVRVVVLDKVT OLLI F 600 FGKLLVVGGVGVLSFFFFSGRI PGI(DFKSPfHLNYYWLPIM TSILGA 648 FGKLLVVCGVGVLSFFFFSGRIPGLGKDFKSPHLNYYWLPIM

TSTLGA

FGKLLVVGGVGVLSFEFFSGRIPGLGKDFKSPHL4YYWLPIMRI4PITPTGHVFOTSILGA 660 YVIASGFFSVFGMCVETLFLCVLEELERNNGSLDRPYYMSKSLLKILGBKNEAPPDNKKR 708 YVIASGEFSVFGMETLCE'LEDLERNSTRPYYMSSLLKAILGKENEAPPDNK R 720 KK 710

KK

KK 722 24P4C12v.1: 649 24P4Cl2v.g: 661 24P4C12v.l1: 709 24P4C12v.B: 721 Tabl~e LX. Nucleotide sequence of gagccatggg gggaaagcag cgggacgagg acgacccctc ctttcgaggc cccatcaaga tcctcttcct gctcttcatt ctaggttaca gaqacccccg gcaagtcctc taccccagga agaacaaaga taagccgtat ctcctgtact transcript variant 24P4C12 v.9 (SEQ ID NO: 106) acatcatctc cctgcccgga tcttctataC tcacaagcct ggcgctgctt ccaccataca ttaagatctt ctctggtctt tgqtqCtgdt agtaccgagt tcagtgecta ttgaagccat ccctcctgaa tggtcacctt ctctgcccaC ccggctgtqa CCtcgtgCCC gttctgtctt tactggccct acaagc~cca accacactgg tcatcttgga tcatgtgctg gcaatgcata cgttcatgct tgctgctgtt ttttctccgg actggctgc gcgttttcgg agttqctgag ggaccuatgq aaaaaatagg tccatggacc gcaqgggatc tgaagatttt gagcctactg cctgggagtg gCtgCgggac ccagagcgtg cctgctgctg ggaggccagc tgtcctcctc gcagccagcc qaaaqtgcca agggctgatg caatctgcaa gggccaatgc ggacatccct gtcattggca qtatattgac tttcaagtgc catcatgatc actcatgcga Ctttgggaag tcgcatcccg catcatgacc catgtqtgtq aacggcctac a ctgtqggaa aacttttgtc ctctgcccca aacgttactc agcggtctta gcccagtcct tttatcttgc ctggqgtgc aagggCgCCt caggagacct atgctcatct aaggctgtg ctcatctgca actcttgga:atgacgaggc acagaagctg tcgtggtggg actctactgg tcaacatctt agtgccccac aaaacgagtt tgccaggggt gtUtCCtcC ttgacagct ggtattggat ttCtgCgCCt tggcatacgg CCatCtCCCd ggctggccgc tcctgcggca gacagatgat Qtacgggaag cacagatgtc gattgtggcc ggcctactgt cagctgcatc accccaggtg ctcacagact accctggaat CCCCtctgCt cccagggatc caatgcoccga tcttgctgcc ggtggctggg catctactac ccagtcaaat atctgctgcg tggttgtatg ggcatggggg ctgtccagea tgtgtgtcct gttggggaag atgacggtga ccaqctctqq accaatgaca gacatcagtg ctqqgggtgg cccctggtgc tgotgggagg gctgggtttc accaccaac cctgatCgtg ttggcggtgc gcggattcgt attgccatcg 120 180 240 300 360 420 480 540 600 660 720 780 840 900 960 1020 1080 1140 1200 1260 1320 1380 1440 1500 1560 1620 1680 1740 1800 1860 1920 1980 2040 gtctaccatg ttgcctactg ggccaogact atgtgctttg ggcatccaac ataaataca: catgcaaccc tgcgtcttcc agggctactc atctatgggg tcctggggct gtcctcgctg gagcctttgc accttcccct taatctctgc tttggagccc tcatcctgac cacaagctca gaggagtgca tgcctctggt gtctggaaaa gccatctacg ggaaqaattt aacattgtca gggtggtcgt ctgctggtq tcggaggt gggctgggta aagactttaa tccatcctgg gggcctatgt gacacgctct tcctctgctt cacggzccac atccaaaggc cttctgqacc CtCCttCtaC cttcatccgc ccttgtgcag gaaccctgta atttatcaag ctgtgtctca cctggacaaa gggggtCctg gagcccccac catcgccagc cctggaagac ttctacccac gctctgtatc atcagctccc cttgtgaact ctaatccaac cttaactggg tgggccttcc acactccgtt atagcccggg gcccgctgca ttcctaaacc gccaaaaatg gtcacagacc tzcttctttt ctcaactatt ggcttcttca ctggagcgga WO 2004/050828 PCT/US2002/038264 acaacggctc cctgqaccqg ccctactaca tgtccaagag ccttctaaag attctgggca 2100 agaagaacga ggcgcccccg gacaacaaga agaggaagaa gtgacagctc cggccctgat 2160 ccaggactgc accccacccc caccgtccag ccatccoace tcacttcgcc ttacaggtct 2220 ccattttgtg gtaaaaaaag gttttaggcc aggcgccgtg gctcacgcct gtaatccaac 228C actttgagag gctgagjgcgg gcggatcacc tgagtcaqqa gLtcgagacc agcctggcca 2340 acatggtgaa acctccgtct ctattaaaaa tacaaaaatt agccgagagt ggtgqcatgc 2400 acctgtcatc ccagctactc gggaggctga ggcaggagaa tcgcttgaac ccgggaggca 24G0 gaggttgcag tgagccgaga tcgcgccact gcactccaac ctgtgaca gactct9tct 2520 ccaaaacaaa acaaacaaac aaaaagattt tattaaagat attttgttaa ctcagtaaaa 2580 aaaaaaaaaa aaa 2593 Table LXI. Nuclaotide sequence a:lignmient of 24P4Cl2v.1 v.1 (SEQ IDJ NO: 107) and 24P4C12 V-9 (SEQ ID NO: 108) Score 2188 bits (1138), Expect O.Dldentities =1138/1138 (100%) Strand PlusI Plus 241P4C12v.1:- 1 24P4Cl2v.9: 1 24F4C12v.1: 6 24P4C12v.9: 6: 24P4C12v.1: 1L 24P4C12v,9* 1; 24P4C12v.l: 1L 24P4C12v. 9: L~ 24P4C12v1l: 2 24P4C12v.9: 2 24P4C12v.1: 3 24P4C12v.9: 3 24P4Cl2v.1: 3 24P4Cl2v,9: 3 24P4Cl2v.1: 4 24P4Cl2v.9: 4 24P4C12v.l: 4 24P4C12v,.9: 4 24P4Cl2v.1: 5 24P4Cl2v.9; 5 24P4C12v,1: 6 gagccatggggqgaaagcagcgggacgaggatgacgaggcctaCgggadgccagtcaaat gagccatggggggaaagcagcgggacgaggatgacgaggcctacgggaagccagtcaaat acgacccctcctttcgaggccccatcaagaacagaagctgcacagatgtcatctgctgog acgjacccctcctttcgaggccccatcaagaacagaagctgcacagatgtcatctgctgg tcctcttcctgctcttcattctaqgttacatcgtggtCgggattqtggcctggttgtatg tcctcttcctgctcttcattctaqgttacatcgtggtggggattgtggcctggttgtatg gagacccccggcaagtcctctaccccaggaactctactggggcctactgtggcatggggg gagacccccggcaagtcctctaccccaggaactctactggggcctactgtggcatggggg agaacaaagataagccgtatctcctgtauttcaacatcttcagctgcatcctgtccagca agaacaaagataagccgtatctcctgtacttcaacatrttcagctgcatcctgtccagca acatcatctcagttgctgagaacggcctacagtgccccacaccccaggtgtgtgtgtcct acatcatctcagttqctgagdacggcctacagtgccccacaccccaggtgtgtgtgtcct cctgcccggaggacccatggactgtgggaaaaaacgagttctcacagactgttggggaag cctgcccggaggacccatggactgtgggaaaaaacgagttctcacagactqttggggaag tcttctatacaaaaaacaggaacttttgtctccaggggtaccctggaatatgacggtga tcttctatacaaaaaacaggaacttttgtctgccaggggtaccctggaatatgacggtga tcacaagcctgcaacaggaactctgccccatttcctcctcccctctqctccagctctgg tcacaagcctgcaacaggaacttgoccccagtttcctcctcccctctgctccagctctgg ggcgctgctttccatggaccaacgttactccaccggcgctcccagggatcaccaatgaca ggcgctgctttccatggaccaacgttactccaccggcgctccagggatcaccaatgaca ccaccatacagcaggggatcagcggtcttattgacagcctcaatgcccgagacatcagtg WO 2004/050828 PCT/US2002i038264 24P4C12v. 9: 601 ccsccatacagcaggggatcagcggtcttatcgacagcctcaatgcccgagacatcagtg 660 242'LCl2v.1: 661 ttsagacctttgaagattttgcccagtcctggtattggattcttgttgccctgggggtgg 720 24P4C12v. 9: 661 ttaagatctttgaagattttccagtcctggtattggattcttgttgccctgggggtgg 720 24P4C12v.1: 721 cnctggtcttgagcctactgtttatcttgctcctgcgcctggtgqctgggcccctggtgc 780 24P4C12v. 9: 721 rtrtggtrttgagcctactgttcatcttgcttrtgcgcctggtggctgggcccctggtgc 780 24P4C12v.1: 781 tggtgctgatcctgggagtgctgggcgtgctggcatacggcatctaccactqctgggagy 840 24P4C12v. 9: 781 tggtgcrgatcctgggaqtgcogggcgtgctggcatacggcatctactactgctgggagg 840 24P4C12v.1: 841 agtaccgagtgctgcgggacaagggcgcctccatctcccagcggtttcaccaccaacc 900 24P4C12v. 9: 841 agtaccqagtgctgcgggacaagggcqcctcratrtcccagctcggtttcsccaccaarc 900 24P4C12v.1: 901 tcagtgcctaccagagcgtgcaggagacctggctggccgccctgatcgtgttggcggtgc 960 24P4C12v-9: 901 tcagtgcctaccagagcgtgcaggagacctggctggccgccctgatcgtgttqgcggtgc 960 24P4012v. 1: 961 ttgaagccatcctgctgctgatgctcatcctcctgcggcagcggattcgtattgcratCg 1020 24P4O12v. 9: 961 ttgaagccatcctgctgctgatgctcatcttcctgcqgragcggattcgtattgCcatcg 1020 24F4C12v.1: 1021 ccctcctgaaggaggccagcaaggctgtgggacagatgatgtctaccatgttctacccac 1080 24P4C12v-9: 1021 crctcctgaaggaggcragcaaggctgtgggacagatgatgtctaccatgttctacccac 1080 24F4012v.1: 1081 tgqtcacctttgrcctcctcctcatctgcattgcctactgggccatgactgcrctgta 1138 24P4C12v. 9: 1081 tgqtcacctttqucctcctcc-tcatctqcattqcctactgggccatgactgctctgta 1138 Score 2738 bits (1424), Expect 0 Jldentities 1424/1424 (100%) Strand PlusI Plus 24P4C12v. 1: 1164 tatgtgctctgggratccaar~atcagctcccccggctgtgagaaagtgccaatsaataea 1223 24P4012v. 9: 1170 tatgtgctctgggcatccaacatcagcrcrccccggcrgtgaqaaagtgccaataaataCa 1229 24P4C12v. 1: 1224 tcatgcaarcccacggcccaccttgtgaactcctcgtgcccagggctgatgl;cgtCttc 1283 24P4C12v. 9: 1230 tcacgcaaccccacqgcccaccttgtgaactcctcgtgcccagggctgatqtgcgtcttc 1289 24P4C12v. 1: 1284 cagqgctactcatccaaaqqcctaatccaacgttccgtcttcaatctgcaaatctatggg 1343 24P4C12v. 9: 1290 cagggctactcatccaaaggcctaatccaacgttccgtcttcaatctgcaaatctatgqg 1349 .24P4C12v. 1: 1344 qtcctqgggctcttctggacccttaactgggtactggcZctgggrccatgCgtcrtCgct 1403 24P4C12v,. 9: 1350 gtcctggggcocotctggacccttaactgggtaccggccctgggccaatqcgtcctrgct 1409 2424C12v. 1: 1404 ggagccttgcctccttctactgggccttccacaaqccccaggacatccctaccttcccr 1463 24P4C12v. 9: 1410 ggagcctttgcctccttctactgggccttccaCaagccccaggacatccctaccttcccc 1469 WO 2004/050828 WO 204/00828PCTIUS2002/038264 24P4C12v. 1: 24P4Cl2v. 9: 24P4Clv. 1: 24P4C12v. 9: 24P4C12v. 1: 24P4C12v. 9: 24E'4Cl2v. 1: 24P4C12v. 9: 24P4C12v. 1: 24P4Cl2v. 9: 24P4C12v. 1: 24r4Cl2v. 9: 24P4C12v.l1: 24P4ClZv. 9: 24P4C12v. 1: 24P4Cl2v. 9: 24 P4Cl2v. 1: 24P4C12v.9: 24P4Cl2v.l: 24P4Cl2v.9: 24 P4Cl2v 1: 24P4Cl2v. 9: 24P4Cl2v. 1: 24P4Cl2v. 9: 24P4C12v. 1: 24P4C12v, 9: 24P4C12v. 1: 24P4Cl2v. 9: 1464 1470 1524 1530 1584 1590 1644 1650 1704 1710 1764 1770 1824 1630 1884 1890 1944 1950 2004 2010 2064 207C 2124 2130 2194 2190 2244 2250 ttaatctctgccttcatccgcacactccgttaccacactgggtcattggcatttggagcc ttaatctcrgccttcatc.gcncactcgttacacactggtcttggcatttggagtc ctcatcctgacccttgtgcagatagcccgggtcatcttggagtatattgaccacaagctc ctcatcctgacccttgtgcagatagcccgggtcatcttggagtatattgac.cacaagctc agaggagtgcagaaccotgtagcccgotgcatcatgtgctgtttcaagtgctgcctctgg agaggagtgcagaaccctgtagcccgctgcatcatgtgctgtttcaagtgctgcctCtgg tgtctggaaaaatttatcaagttcctaccgcadatgcatdcatcatgatcgCCatctac tgtctggaaaaatttatcaagttcctaaaccgcaatgcatacatcatgatcgccatctac gggaagaatttctgtgtctcagccaaaaatgcgttcatgCtactcatgCgaaacattgtc gggaagaatttctgtgtctcagccaaaaatgcgttcatgctactcatgCgaaacattgtc agggtggtcgtcctggacaaagtcacagacctgctgctgttctttggqaagctgctggtg agggtggtcqtcctggacaaagtcacaqacctgctgctgttctttgggaagctgctggtg StcggaggcgtgggggtcctgtccttcttttttttctccggtcgCatccc9ggtgggt gtcggdggcgtggqqgtcctgtccttcttttttttctccggtcgcatcCcggggctgggt aaagactttaagagcccccacctcaactattactggctgcccatcatgacctccatcctg aaagactttaagagcccccacctcaactattactggctgccatcatgacctccatcctl ggggcctatgtcatcgccagcggcttcttcagcgttttcggcatgtgtgtggacacgctc ggggcctatgtcatcgccagcggcttcttcagcgttttcggCatgtgtgtggacacgctc ttcctctgcttotggaagacctggagcggaacaacggctccctggaccggCCctactac ttcctctgcttcctggaagacctggagcggaacaacggctccctggaccggCcctactac atgtccaagagccttctaaagattctgggcaagaagaacgaggCgcccccggacaacaag atgtccaagagccttctaaagattctgggcaagaagaaCaggcgcccccggacaacaag adgdggaagadgtgacagctccggccctgatccaggactgcaccccaCccccaccgtcca aagaggaagaagtgacagctccggccctgatccaggactgcaccccacccccaccgtcca gccatccaacctcacttcgocttacaggtctccattttgtggtaaaaaaaggttttaggc caggcgccgtggctcacgcctg taatcc-aacactttgagaggCtgaggcgggcggatcac caggcgccgtggctcacgcctqtaatccaacactttgagaggctgaggcgggcggatcac 1523 1529 1583 1589 1643 1649 1703 1709 17 F 3 1769 1823 1829 1883 1989 1943 1949 2003 2009 2063 2069 2123 2129 2183 2189 2243 2249 2303 2309 WO 20041050828 WO 204100828PCT/US20021038264 24E'4Cl2v. I: 2424C12v. 9: 2424012v. 1: 24?4012v. 9: 24P4C12v. 1: 2424212v. 9: 24?4C12v. 1: 2404012v. 9: 24P4Cl2v. 1: 24P4C12v. 9: 2304 cLgagtcaggagtrcgagaccagccrggccaacatggtgeeacctccgtctctattaaaa 2363 2310 ctataggtqgcactgcactggactcttttaa 2369 2364 atacaeaattagccgagagtggtggcatgcaccsgscascccegctactcqggeqqgctg 2423 2370 aracaaaeatzagccgagagtggtggcetgcaccsgtcascccegcsectcgqqeggctq 2429 2424 agqcaggagaetcgcttgaacccgggaggcagaggttgcegtgagccgeqetcgcgccec 2483 2430 eggceggegaatcgcttgaacccgggeqgcaqeqgttgcagtgagccgegatcgcgccac 2489 2484 tgcactccaacccgggsgacagactcsgscsccaaeacaaaacaaacaaacaaaaegatt 2543 2490 tqcectcceecctgggtqacagectctgsctccaaeacaaeaceeacaeaceeeaget 2549 2544 ttattaeagerettttgtteactcagoaaaaaaaaaaaaa 2567 2550O ttattaeagacattttgtteactcegtaaaeaaaaeeaaaaaaa 2593 Table LXII. Peptide, sequences of protein coded by 2494012 v.9 (SEQ ID NO:- 109) MGGKQRDFDD EAYGKPVKYD PSFRGPIKNR SOTDVICCVL FLLFILGYIV VGIVAWLYOD PRQVLYPRNS VGAYCGMGEN KDKFYLLYFN IFSCILSSNI ISVAENGIJOC PTPQVCVSSC 120 PEOPWTVGKN EFSCTVGEV' YTKNRNFCLF GVPWNMTVIT SLQQELCPSF LLPSAPALGR 180 CFPWTNVTPP ALPGITNDTr IQOGISOLID SLNARDISVK IFEDFAQSWY WILVALEVAL 240 VLSLLFILAJ RLVAGPLVLV LILGVLGVLA YGIYYCWEEY RVLRDKGASI SQILGFTTNLS 300 AYQSVQETWL AALTVLAVLE AILLJJMLIFL RORTRLMIAL LKEASKAVGQ MMSTNEYPLV 360 TFVLLLTCIA YWANTALYPL FTQPATLGYV LWASNrSSPG CEKVPINTSC NP EAHLVNSS 420 CPGLMCV'QG YSSKGLIQRS VFNLQTYGJI GLFWTLNWAVL ALGQCVIAGA FASFYWAFUC 480 PODIPTEPTA SAFIRTLRY- TOSLAFOALI LTLVQEARVI LSYIDHRLRG VQNPVARCI4 540 CCFBZCCLWCL EKFIKFLNRN AYIMIATYI{ WFCVSAKNAF MLLMRNIVRV VVLDKVTDLL 600 LFFGKLLVVG GVGVLSFFFF SGRIPGLGED FKSPELNYYW LPIb4TSILGA YVIASGFFSV 660 FGMCVDTLFL CFLEDLER4W GSLDRYYMS KSLLKILGKK NEAPFDNKKR KK1 712 Table LXIII. Amino acid sequence alignment of 24P4C12v. 1 v. 1 (SEQ ED) NO: 110) and 24P4C12 v.9 (SEQIDiNO: 111) Score 1424 bits (3696), Expect 0.Oldentities 704/713 Positives= 705/713 Gaps 4/713 24P4C12v.2: 1 2424C12v.9: 1 24F4C12v.1: 6 24P4C12v.9; 6 2454012v.1: 1 2424C12v.9: 1 24P4C12v.1: 1 2 4?4C12v.9: 1 24P4C12v.1: 2 24P4C12v.9: 2 24P4C12v.1: 3 24P4C12v.9: 3 MGGKQRDEDDEAYGKPVKYOPSFRGPIKNRSCTDVICCVLFLLFILGYIVVGIVAWLYGD bGGKQRDEDDEAYGKPVKYDPSFRGPIKNRSC2EDVICCVLFLLFILGYIVVGIVANLYGD t4GGKQPDEDDEAYG;KPVKyDPSFRGPIKNRSCTDVICCVLFLLFILCYIVVGIVAWLYGD PRQVLYPRNSTGAYCGMGENKDKFYLLYFNIFSCILSSNIIOVAENGLOCPTPOVCVSSC 12( PRQVLYPRNSTGAYCGMGEI4KDKEYLLYFNIFSCILSSIIVAFt4GLOCFT FQVCVSSC FRQVLYPRNSTGAYCGMGENKDKPYLLYFNIFSCILSSWIISVAENGLQCFTPQVCVSSC 12( PEDPNTVGKNEY-SQTVGEVFYTKN4RNFCLPGVPWNMTVITSLQQELCPSFLLPSAPAILGR 18( PEDPWTVGKNEFSQTVGEVFYTKNRNFCLPGVPWNMTVITSLQQELCPS FILPSAPALGR PEDPWTVGKNEFSQTVGEVFYTKNRNFCLPGVFWNMTVITSLQQELCPSFILPSAPALGR 2.8) CFPWTNVTPPALPGITNDTTIQQGISGLIDSLNARDISVKIFEDFAQSWYWAILVALGVAL 24(

CFPWTNVTPPALPGITNDTTIOQGISGLIDSLNARDISVCIFEDFAQSWYWILVALGVAL

CFPWThVTPPALPGITNDTTIQGISGLIDsLNAPRDISvKIFE0FAOSWYWqILVALGVAL 24( VLSLLFILLLRLVAGPLVLVLILGVIGVLAYGIYYCWEEYVLRDKGASISQLGFPTNLS VLSLLFI LLLRLVAGPLVLVLI LGVLGVLAYGIYYCWEEYRVLRDKGAS ISQLGFTTNLS VLSLLFILLLRLVAGPLVLVLILGVLGVLAYGIYYCWEEYRVLRDKGASISQLGFTTNLS AYQSVQETWLAALIVLAVLEAILLLMLIFLRQIRIAIALLKjEASKAVGQMqMSTMFYPLV 36( AYQSVOETWLAALIVLAVLEAI LLLMLIFLRQRIRIAIALLKEASKAVGQMbSTMFYFLV AYQSVOETWLAALIVLAVLEAILLLr4LIFLRQRIRIAIALLKEASKAVGI4DSTMFYPLV 362 WO 2004/050828 PCT/US2002/038264 24P4Cl2v. 1: 24P4CL2v. 9: 24D4C!2v. 1: 24P4C12v. 9: 24P4C12v. 1: 24P4C12v. 9: 24 24Cl2v 1: 2424CI2v.9: 2424 Cl2v. 1: 24P4 Cl2v.9: TFVLLLICIAYWAI4TALYLATSGQPQ YVLWASNISS PGCEKVPINTSCNPTA{LVNS TFVLLLICAZYWAATALY 02 YVLWASNISSPGCSKVPINTSCNPTAHLVNS TFVLLLICIZYWANTALYPLPT-QPATLGYVLWASNISS PGCEKVPINTSCNPTAHLVNqS SCFGLNCVEQGYS SKGLIQRS VFN4LQIYGVLGLFWTLNWVLALGQCVLAGAFAS FYWAPH SCPGLMCVFQGYS SKGLIQRSVFNILQIYGVLGLFWTLNWVLALGQCVLAGAFAS FYWAFE SCPGLMCVFQGYSSKGLIQRSVF4LQIYGVLGLFWTLNWVLAJGQCVLAGAFAS FYWAFH I(PQDT PT TEPdSAFiRTLRYHTGSLAFGALILTLVOIARVI LEYI DHRLRGVQNPVAPZI KPQDIPTB'PLI SAF:RTLRYHTGSLAFGALT LTLVOIARVTLJEYIDHKLRGVQNPVARCI KPQDIPTFPLI SAFIRTLRYHTGSLAFXALT LTLVQIARVTLEYI DHRLRGVQNPVARCI MCGFKCCLWCLEKFIKFLHRNAYIMIAIYGEN FCVSAEKHAFMLL14NIVRVVVLD)KVTDL MCCFKCCLWCLEKFTKFLNRNAY EMTATYCKNFCVSAZNAFb4LLRN TVRVVVLOKVTD)L MCCFKCCLWCLEEFLKELNRNAY EMTAIYGKNFCVSAKNAFMLLMRN TVRVVVLDK\JTEL

LLFFGKLE

4 VVGGVGVELSFFFFSGRIPGLGKDFKSPHLNYYWLPIMTS ThGAYVIASGFFS LLFFGKLLVVGGVGVLSFFFFSGRFGTJGKDFKSPHLNYYWLPIMTS TLCAYVIASGFES

LLFFGKLLVVGGVGVLSFFFFSGRIPGLGKDFKSPHLNYYWLPIMTSTLGAYVIASGFFS

24P4C12v.1: 658 VFGNCVDTLFLCFLEIJLERNNGSLE)RPYYMSKSLLKILGKEZNEAFPDNKFRKK 710 VFGNCVDTLFLCFLEDLERNNGSLDRPYYD4SKS3LLKIIGKKNEAPPNKKRKK 2424 Cl2v. 9: 663 VFGNCVDTLFLCFLEDLERNNGSELRPYYNSKSLLKILGKKNEAPPDNKKRKK 712

Claims

4. A process for producing a 24P4C12 protein comprising culturing the host cells of claim 3 under conditions sufficient for the production of the 24P4C12 protein. The process of claim 4, further comprising recovering and isolating the 24P4C12 protein so produced.
6. An isolated and purified protein produced by the process of claim 5 having an amino acid sequence shown in SEQ ID NO: 7, 11, 13, 15, 17, or 19.
7. An isolated 24P4C12 protein, wherein the 24P4C12 protein has an amino acid sequence comprising SEQ ID NO: 7, 11, 13, 15, 17, or 19.
8. An antibody or fragment thereof that immunospecifically binds to an epitope on a 24P4C12 protein of SEQ ID NO: 7, 11, 13, 15, 17, or 19.
9. An antibody or fragment thereof that immunospecifically binds to a 24P4C12 protein comprising an amino acid sequence set forth in SEQ ID NO: 7, 11, 13, 15, 17, or 19. The antibody or fragment thereof of claim 8 or 9, which is monoclonal.
11. The fragment of any one of claims 8 to 10 which is an Fab, F(ab') 2 Fv or Sfv fragment.
12. The antibody or fragment thereof of claim 8 or 9, which is a human antibody. COMS ID No: ARCS-165644 Received by IP Australia: Time 14:22 Date 2007-10-22 22/10 '07 14:19 FAX 613 8618 4199 F.B. RICE Co. _008 O S225
13. The antibody or fragment thereof of any one of claims 8 to 12, which is bound to O an agent. C"1
14. The antibody or fragment thereof of claim 13, wherein the agent selected from the group consisting of radioactive isotopes, chemotherapeutic agents, labels, and toxins. The antibody or fragment thereof of claim 14, wherein the radioactive isotope is C' selected from the group consisting of 1 At, 131, 12 s, 0 1 Re, '1Re, 53 Sm, 21 2 Bi, 2 P and Cl radioactive isotopes of Lu. <C
16. The antibody or fragment thereof of claim 14, wherein the chemotherapeutic agent is selected from the group consisting of taxol, actinomycin, mitomycin, etoposide, tenoposide, vincristine, vinblastine, colchicine, gelonin, and calicheamicin.
17. The antibody or fragment thereof of claim 14, wherein the toxin is selected from the group consisting of diphtheria toxin, enomycin, phenomycin, Pseudomonas exotoxin (PE) A, abrin, abrin A chain, mitogellin, modeccin A chain, and alpha-sarcin.
18. A hybridoma that produces an antibody of claim 9.
19. A recombinant vector comprising a polynucleotide encoding a monoclonal antibody of claim 9. Host cells comprising the vector of claim 19.
21. A method to produce antibodies specifically immunoreactive with a 24P4C12 protein which comprises culturing the hybridoma of claim 18 or the cells of claim 20 and recovering said antibodies.
22. An in vitro method for detecting the presence of a 24P4C12 protein or polynucleotide in a test sample comprising; contacting the sample with the antibody or fragment of any of claims 8-15 or with polynucleotide that is complementary to the polynucleotide of claim 1 that specifically binds to the 24P4C12 protein or polynucleotide, respectively; and COMS ID No: ARCS-165644 Received by IP Australia: Time 14:22 Date 2007-10-22 22/10 '07 14:19 FAX 613 8618 4199 F.B. RICE Co. .009 O S226 O detecting binding of 24P4C12 protein or polynucleotide, respectively, in the sample. O ci
23. The method of claim 22, wherein the determining step further comprises comparing an amount of binding of the antibody or polynucleotide that specifically binds to the NO 24P4C12 protein or polynucleotide in the test sample to the amount of said binding in a control sample.
24. The method of claim 23, wherein the presence of elevated 24P4C12 C polynucleotide or protein in the test sample relative to the control sample provides an indication 0 O of the presence of cancer. A method of inhibiting growth of cells expressing a 24P4C12 protein comprising an amino acid sequence set forth in SEQ ID NO: 7, 11, 13, 15, 17, or 19, comprising providing an effective amount of an antibody according to any one of claims 5-16 to the cells, whereby the growth of the cells is inhibited.
26. A method of inducing an immune response to a 24P4C12 protein comprising an amino acid sequence set forth in SEQ ID NO: 7, 11, 13, 15, 17, or 19, which compromises administering at least an epitope of said protein to a subject in an amount effective to elicit an immune response.
27. The method of claim 26, wherein the immune response is a B cell or T cell response.
28. Use of an epitope of a 24P4C12 protein comprising a sequence set forth in SEQ ID NO; 7,11, 13, 15,17, or 19 epitope for the preparation of a medicament to induce an immune response in a subject
29. Use of an antibody for the preparation of a medicament which delivers an agent to cells expressing a 24P4C12 protein comprising a sequence set forth in SEQ ID NO: 7, 11, 13, 17, or 19, wherein the antibody comprises an antibody according to any one of claims 13- 17. COMS ID No: ARCS-165644 Received by IP Australia: Time 14:22 Date 2007-10-22 22/10 '07 14:19 FAX 613 8618 4199 F.B. RICE Co. __010 O S227 O
30. Use of an effective amount of an antibody according to any one of claims 8-17 for O the preparation of a medicament which inhibits growth of cells expressing a 24P4C12 protein i comprising an amino acid sequence set forth in SEQ ID NO: 7. 11, 13, 15, 17, or 19.
31. A method of isolating a peptide useful as a vaccine to elicit an Immune response l- to a 24P4C12 protein comprising an amino acid sequence set forth in SEQ ID NO: 7, 11, 13, 17, or 19, which method comprises In C identifying an HLA supertype for which binding of epitopes of said vaccine is desired; 0 l selecting from the peptides listed in any one of Tables VIIl to XLIX any epitopes of SEQ ID NO: 7, 11, 13, 15, 17, or 19 disclosed to bind alleles of said identified supertype: experimentally assessing the ability of said peptides to bind to at least one allele of said HLA supertype and isolating a peptide useful as a vaccine a peptide that binds with an IC50 equal to, or less than, 500 nanomolar to said HLA supertype allele.
32. The method of claim 31, wherein peptides are selected that bind within equal to, or less than, 500 nanomolar to three alleles of said HLA supertype.
33. A peptide when isolated by the method of claim 31 or 32 34 Use of a peptide isolated by the method of claim 31 or 32 or the peptide of claim 33 for the preparation of a vaccine to elicit an immune response to a 24P4C12 protein comprising an amino acid sequence set forth in SEQ ID NO: 7, 11, 13, 15, 17, or 19. The use of claim 34, wherein said immune response is a CTL response.
36. The use of claim 34, wherein the immune response is a humoral response.
37. The polynucleotide of claim 1 or the nucleic acid of claim 2 or the host cell of claim 3 or 20 or the vector of claim 19 or the process of claim 4 or 5 or the protein of claim 6 or 7 or the antibody or fragment thereof of any one of claims 8 to 17 or the hybridoma of claim 18 COMS ID No: ARCS-165644 Received by IP Australia: Time 14:22 Date 2007-10-22 22/10 '07 14:20 FAX 613 8618 4199 F.B. RICE Co. S228 or the method of any one of claims 21 to 27, 31 or 32, or the use of any one of claims 28 to O or 34 to 36 or the peptide of claim 33 substantially as described herein with reference to any i one or more of the examples and/or drawings. SDATED this TWENTY SECOND day of OCTOBER 2007 Ci LC Agensys, Inc. Patent Attoeys for the Applicant: Patent Attorneys for the Applicant: aioii F.B. RICE CO. COMS ID No: ARCS-165644 Received by IP Australia: Time 14:22 Date 2007-10-22