[go: up one dir, main page]

WO1999024596A1 - Plasmides d'expression utilises dans des vaccins a base d'acide nucleique a plusieurs epitopes - Google Patents

Plasmides d'expression utilises dans des vaccins a base d'acide nucleique a plusieurs epitopes Download PDF

Info

Publication number
WO1999024596A1
WO1999024596A1 PCT/US1998/023745 US9823745W WO9924596A1 WO 1999024596 A1 WO1999024596 A1 WO 1999024596A1 US 9823745 W US9823745 W US 9823745W WO 9924596 A1 WO9924596 A1 WO 9924596A1
Authority
WO
WIPO (PCT)
Prior art keywords
epitopes
plasmid
nucleic acid
expression
sequence
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/US1998/023745
Other languages
English (en)
Inventor
Harry C. Ledebur, Jr.
Jeffrey L. Nordstrom
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Genemedicine Inc
Urigen Pharmaceuticals Inc
Original Assignee
Genemedicine Inc
Urigen Pharmaceuticals Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Genemedicine Inc, Urigen Pharmaceuticals Inc filed Critical Genemedicine Inc
Priority to CA002309344A priority Critical patent/CA2309344A1/fr
Priority to EP98957671A priority patent/EP1030927A1/fr
Priority to AU13872/99A priority patent/AU1387299A/en
Publication of WO1999024596A1 publication Critical patent/WO1999024596A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

Links

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
    • C12N15/85—Vectors or expression systems specially adapted for eukaryotic hosts for animal cells
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/51—Medicinal preparations containing antigens or antibodies comprising whole cells, viruses or DNA/RNA
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/51—Medicinal preparations containing antigens or antibodies comprising whole cells, viruses or DNA/RNA
    • A61K2039/53—DNA (RNA) vaccination
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2830/00—Vector systems having a special element relevant for transcription
    • C12N2830/42—Vector systems having a special element relevant for transcription being an intron or intervening sequence for splicing and/or stability of RNA
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2840/00—Vectors comprising a special translation-regulating system
    • C12N2840/20—Vectors comprising a special translation-regulating system translation of more than one cistron
    • C12N2840/203—Vectors comprising a special translation-regulating system translation of more than one cistron having an IRES
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2840/00—Vectors comprising a special translation-regulating system
    • C12N2840/44—Vectors comprising a special translation-regulating system being a specific part of the splice mechanism, e.g. donor, acceptor

Definitions

  • the invention relates generally to gene therapy, in particular, the invention relates in part to improved plasmids and methods for nucleic acid based vaccines.
  • Plasmids are an essential element in genetic engineering and gene therapy. Plasmids are circular DNA molecules that can be introduced into bacterial cells by transformation which replicate autonomously in the cell. Plasmids allow for the amplification of cloned DNA. Some plasmids are present in 20 to 50 copies during cell growth, and after the arrest of protein synthesis, as many as 1000 copies per cell of a plasmid can be generated. (Suzuki et al., Genetic Analysis, p. 404, (1989).)
  • Plasmid design and construction utilizes several key factors.
  • plasmid replication origins determine plasmid copy number, which affects production yields. Plasmids that replicate to higher copy number can increase plasmid yield from a given volume of culture, but excessive copy number can be deleterious to the bacteria and lead to undesirable effects (Fitzwater, et al . , EMBO J. 7:3289-3297
  • genes that code for antibiotic resistance phenotype are included on the plasmid and antibiotics are added to kill or inhibit plasmid-free cells.
  • Most general purpose cloning vectors contain ampicillin resistance ( ⁇ -lactamase, or bla) genes. Use of ampicillin can be problematic because of the potential for residual antibiotic in the purified DNA, which can cause an allergic reaction in a treated patient.
  • ⁇ -lactam antibiotics are clinically important for disease treatment. When plasmids containing antibiotic resistance genes are used, the potential exists for the transfer of antibiotic resistance genes to a potential pathogen.
  • neo gene which is derived from the bacterial transposon Tn5.
  • the neo gene encodes resistance to kanamycin and neomycin (Smith, Vaccine 12:1515-1519 (1994)).
  • This gene has been used in a number of gene therapy studies, including several human clinical trials (Recombinant DNA Advisory Committee Data Management Report, December, 1994, Human Gene Therapy 6:535-548). Due to the mechanism by which resistance is imparted, residual antibiotic and transmission of the gene to potential pathogens may be less of a problem than for ⁇ -lactams.
  • plasmid vectors have also been shown to affect transfection and expression in eukaryotic cells. Certain plasmid sequences have been shown to reduce expression of eukaryotic genes in eukaryotic cells when carried in cis (Peterson, et al . , Mol . Cell . Biol . 7:1563-1567 (1987); Yoder and Ganesan, Mol . Cell . Biol . 3:956-959 (1983); Lusky and Botchan, Nature 293:79-81 (1981); and Leite, et al . , Gene 82:351-356 (1989) ) .
  • Plasmid sequences also have been shown to fortuitously contain binding sites for transcriptional control proteins (Ghersa, et al . , Gene 151:331-332 (1994); Tully and Cidlowski, Biochem. Biophys . Res . Comm . 144:1-10 (1987); and Kushner, et al . , Mol . Endocrinol . 8:405-407 (1994)). This can cause inappropriate levels of gene expression in treated patients.
  • nucleic acid vaccines or the use of plasmid encoding antigens, has become an area of intensive research and development in the last half decade. Comprehensive reviews on nucleic acid vaccines have recently been published [M.A.
  • the use of epitopes small immunologically relevant protein sequences that are capable of inducing both cellular and humoral responses that result in a protective or therapeutic immune response against large and complex pathogens, is an attractive and amenable strategy provided by the present invention for incorporation into nucleic acid-based vaccines. If multiple epitopes are expressed in the context of a synthetic gene construct, immunity against many antigenic targets, multiple strain variants or multiple pathogens is possible. This disclosure describes the structures and characteristics of gene expression systems that are capable of expressing multiple epitopes.
  • the invention provides a method of genetic immunization comprising the step of presenting multiple epitopes to an organism in need of said immunization.
  • the multiple epitopes are presented with one or more augmenting cytokines and/or are presented with a delivery vehicle selected from the group consisting of cationic lipids, delivery peptides, and polymer based deliver systems .
  • the invention features a plasmid for expression of multiple epitopes comprising a nucleic acid sequence encoding multiple epitopes, wherein each of said epitopes is capable of creating an immune response.
  • the plasmid includes a promoter, a 5' UTR sequence, and a 3' UTR sequence, a nucleic acid sequence encoding polyubiquitin, there are spacers between the nucleic acid regions encoding each of said epitopes, there are proteolytic cleavage sites between each of said epitopes, there are ER targeting signals between each of said epitopes, there are lysosomal and/or endosomal targeting sequences between each of said epitopes.
  • the invention provides a multivalent expression system as shown in Figure 8 and selected from the group consisting of two plasmids, two genes, IRES, and alternative splicing and a method of making a plasmid producing the appropriate nucleic acid sequence.
  • Figure 1 shows a plasmid for multiple epitopes using a beads on a string approach.
  • Figure 2 shows a plasmid for multiple epitopes using beads on a string fused to polyubiquitin.
  • Figure 3 shows a plasmid for multiple epitopes using beads on a string with spacers between epitopes .
  • Figure 4 shows a plasmid for multiple epitopes using beads on a string with proteolytic cleavage sites between epitopes.
  • Figure 5 shows a plasmid for beads on a string epitopes with ER targeting sequences.
  • Figure 6 shows a plasmid for multiple epitopes with ER targeting sequences.
  • Figure 7 shows a plasmid for multiple epitopes with lysosomal/endosomal targeting sequences.
  • Figure 8 shows types of multivalent expression systems.
  • Figure 9 shows a DNA vaccine expression plasmid with two genes .
  • Figure 10 shows a design of a drug-controlled DNA vaccine expression plasmid.
  • the multiple epitopes are directly linked to each other. No spacer sequences between the epitopes are included.
  • the epitope sequences themselves are sufficient for the formation of a functional "pseudo" protein that can be processed into individual peptide epitopes via proteosome cleavage. This concept, i.e. beads-on-a-string, is supported by data that shows full CTL responses to numerous epitopes when they are placed into novel locations within different proteins
  • the present invention provides an exemplary expression system for beads-on-a-string as shown below (See Figure 1) :
  • Promoter CMV tissue-specific (e.g. APC-specific) , or synthetic promoter
  • Intron Synthetic intron that has optimized 5' ss, 3' ss and branch point sequences. Current optimal sequence is IVS 8.
  • Initiation codon AUG is placed in the context of the Kozak sequence to ensure optimal initiation of translation.
  • epitope String of epitopes, each having a length of 9-10 amino acid residues in length for class I presentation, or >10 amino acid residues for class II presentation. It appears that at least 15 epitopes may be strung together.
  • Stop codon For termination of translation. To ensure efficient termination, it is desirable to string two stop codons in tandem. 3' To ensure efficient processing of the
  • UTR/poly(A) mRNA an efficient poly (A) signal, such signal as from human growth hormone, is required .
  • Ub chains which target the substrates to the 26S proteasome, an abundant cellular protease.
  • Ub chain that is fused to the N-terminus of the multiple epitopes that are arranged as beads-on-a-string.
  • Expression plasmids with Ub fused to the antigen have been used to achieve class I presentation (Gueguen and Long, Proc . Natl . Acad. Sci . USA 93:14692-97 (1996) ) .
  • flanking sequence can profoundly influence the generation of epitopes (Yellenshaw et al . , J. Immunol . , 158:1727-33 (1997); Shastri et al., J. Immunol . , 155:4339-46 (1995); Del-Val et al . , Cell , 66:1145-93 (1991); Eggers et al . , J " . Exp. Med. 182:1865-70 (1995); Niedermann et al .
  • Spacers may facilitate the formation of epitopes that induce immunity.
  • the spacer sequence should be one that does not conform at all to the rules for class I or II epitope.
  • it may be desirable to include a hydrophobic, basic or acidic residue at the C-terminus of the spacer to facilitate cleavage between the spacer and the adjacent epitope.
  • the length of the spacer that would be optimal is not known and would have to be determined empirically.
  • viruses e.g. retroviruses, flaviviruses
  • Cleavage which occurs at specific sites, is catalyzed by host proteinases or by virally encoded proteinases.
  • the polyprotein from hepatitis C virus is structured as follows: H2N-C-El-E2-p7-NS2-NS3-NS4A-NS4B-NS5A-NS5B-COOH.
  • Host cell signal peptidases cleave the junctions in the region between C and NS2.
  • the viral proteinase (NS2-3 proteinase) cleaves the junction between NS2 and NS3.
  • Another viral proteinase cleaves the junctions between NS3 and NS5B .
  • One approach is to insert host cell cleavage sites between the epitope sequences. This may be achieved by insertion of the sequences that are located at the junctions between the C-El-E2-p7-NS2 proteins of the hepatitis polyprotein. Other possibilities are to utilize the recognition site for specific cellular proteases.
  • a second approach is to insert cleavage sites for the viral proteinase between the epitope sequences. Thus, the site for the sequence recognized by the NS3 proteinase, Asp/Glu-X-X-X-X-Cys/ThrlSer/Ala (Koch and Bartenschlager, Virology, 237:78-88 (1997)), may be inserted.
  • the NS3 proteinase must be also encoded by the expression plasmid.
  • two transcription units are required, one for the multiepitopes, one for the viral proteinase. See Section V. Multivalent expression plasmids for nucleic acid-based vaccines.
  • the classical pathway for antigen presentation in the context of class I involves the partial degradation of antigenic proteins into peptides by the proteasome.
  • the peptides are then transported into the endoplasmic reticulum by peptide transporters (TAP-1 and TAP-2) . It is within the lumen of the ER or cis-Golgi that the peptides are loaded into the binding pocket of the MHC class I molecules.
  • TAP-1 and TAP-2 peptide transporters
  • Ciernik et al . , J. Immunol . 2369-75 (1996) have demonstrated that the immunogenicity of an epitope may be enhanced if the epitope sequence is fused in frame with the adenovirus E3 leader sequence and expressed from a plasmid delivered by particle bombardment.
  • ER targeting signals have also been used by Overwijk et al . , Identification of a Kb-restricted Ctl Epitope of Beta- galactosidase : Potential Use in Development of Immunization Protocols for "Self” Antigens, Methods 12: 117-23 (1997) .
  • the signal sequence allows the epitope to be targeted to the ER by the standard protein translocation process.
  • the N- terminal leader sequence targets the peptide to the ER by first binding, as a nascent sequence, to the 54 kDa subunit of the SRP particle.
  • the leader sequence subsequently binds to the b-subunit of the membrane-bound transporter protein, Sec61p. Following entry into the lumen of the ER through a putative channel, a peptidase cleaves the peptide to remove the leader sequence.
  • This mechanism is independent of the TAP transporter system. This alternative mechanism may be advantageous if epitope formation by proteasome cleavage, or epitope transport by the TAP system, are limiting steps in antigen presentation.
  • a leader sequence preferably needs to be attached to the N-terminus of each epitope. Adding a leader sequence to the multiple epitopes that are arranged as beads-on-a string concept is unlikely to work, since the leader will be attached only to the first epitope sequence. Placing an individual targeting sequence on each of the epitopes that is arranged in a bead-on-a-string assembly is a possibility. However, this strategy will depend on accurate proteolytic cleavage at the N-terminus of the leader sequence and at the C-terminus of the adjoined epitope sequence. A gene expression system that utilizes alternative splicing will yield individual epitopes with their own leader sequences. The peptide epitopes produced by this strategy will not depend on random degradation of a protein precursor.
  • the only processing that is required is N-terminal processing that is associated with protein translocation.
  • the C-terminal ends of the epitopes are defined by the stop codons that are designed into the system.
  • the preprotein products may be incompletely synthesized until protein translocation through the pore into the ER has occurred.
  • the prepeptides may be synthesized in their entirety prior to ER translocation. This may expose the prepeptide to the proteasome and transport of proteins that transport epitopes to the ER by the standard pathway.
  • Table II describes the genetic elements used in the alternative splicing strategy.
  • Element Description ER Signal The N-terminal leader sequence from sequence 5' ss adenovirus E3 or preprolactin Strong 5' splice site, one that exactly matches the consensus sequence . Such a sequence is found in the synthetic intron, IVS8.
  • This sequence is derived from the intron synthetic intron, IVS8. It extends from sequence the 3' end of the 5' splice site to the 5' end of the polypyrimidine tract of the 3 'splice site.
  • 3' splice sites sites will be designed to be used equally. Thus, their relative strengths need to be mathed. This will be accomplished by introducing purines within the polypyrimidine regions of the splice site sequences.
  • Balanced splicing will be achieved by controlling the purine content of the pyrimidine-rich sequences of the 3' splice sites. In general, the greater the purine content, the weaker the splice site.
  • one way to design an appropriate alternatively spliced system for epitopes is to model the 3' splice sites of adenoviral late transcripts.
  • leader sequence after splicing, the leader sequence must be fused in frame with the peptide sequence of each epitope. Also, by altering the strengths of the 3' splice sites, the relative amounts of the epitopes may be varied. This may important if certain epitopes are more dominant than others . Table III below shows an example of a balanced set at 3' splice sites derived from the adenoviral late transcript.
  • TTGTATTCCCCTTAG ⁇ T Ad2 (14149) GTTGTATGTATCCAG ⁇ C Ad2 (16515) GTAACTATTTTGTAG ⁇ A Ad2 (17999) CCATGTCGCCGCCAG ⁇ A Ad2 (18801) ATGTTTTGTTTGAAG ⁇ T Ad2 (21649) TTCCTTCTCCTATAG ⁇ G Ad2 (24094)
  • ADRCG ADRCG . Numbers in parentheses indicate the nucleotide position of each 3' splice site. Note the locations of the purines (A or G) that interrupt the polypyrimidine (C or T) region.
  • FIG. 7 Promoter / 5' UTR / AUG / ER signal sequence / 5'ss / internal intron sequence / 3'ss-l / epitope-1 / LAMP-1 transmembrane-cytoplasmic tail / stop codon-1 / 3'ss-2 / epitope-2 / LAMP-1 transmbrane-cytoplasmic tail / stop codon-2 / 3' ss-3 / epitope-3 / LAMP-1 transmbrane- cytoplasmic tail / stop codon-3 / 3' UTR/poly(A) signal
  • each epitope is preceded by an N-terminal leader sequence (e.g.
  • adenovirus E3 adenovirus E3 and followed by the C-terminal endosomal/lysosomal targeting sequence (e.g. the transmembrane and cytoplasmic tail region of LAMP-1) .
  • endosomal targeting sequence e.g. the transmembrane and cytoplasmic tail region of LAMP-1 .
  • Another sequence that may be employed for endosomal targeting is the cytoplasmic tail of membrane immunoglobulin (Weiser et al . , Science 276:407-9 (1997); Achatz et al., Science 276:409-11 (1997)).
  • nucleic acid-based vaccines it may be important to have the capability of expressing multiple gene products.
  • expression of multiple intact antigens or multiepitope gene product may enhance the potency of the these vaccines.
  • Co-expression of costimulatory proteins, such as IL-2, IL-6, IL-12, GM-CSF, B7.1 or B7.2 have been demonstrated to enhance the immune response to an encoded antigen (Geissler et al . , J. Immunol . 158:1231-1237 (1997), Irvine et al . , (1996); Kim et al., Vaccine 15:879-83 (1997); Okada et al., J. Immunol .
  • Co-expression of proteins that facilitate peptide epitope formation such as proteolytic enzymes (e.g. the NS3 protease from hepatitis C (Koch and Bartenshlager, 1997)) or chaperone proteins (e.g. heat shock protein Hsp65 (Wells et al . , Scand. J. Immunol . , 45:605-12 1997)), may also enhance the response.
  • proteolytic enzymes e.g. the NS3 protease from hepatitis C (Koch and Bartenshlager, 1997)
  • chaperone proteins e.g. heat shock protein Hsp65 (Wells et al . , Scand. J. Immunol . , 45:605-12 1997)
  • Hsp65 heat shock protein
  • nucleic acid-based vaccine expression plasmid that has two genes is shown in Figure 9.
  • the GeneSwitch is a chimeric protein that consists of human progesterone receptor with a modified ligand binding domain, a DNA binding domain from yeast GAL4, and an activator domain from Herpes Simplex VP16.
  • a synthetic steroid mifepristone
  • the GeneSwitch protein becomes activated (binds the synthetic steroid, presumably dimerizes, and translocates to the nucleus) .
  • the activated GeneSwitch then binds to a target sequence (multiple GAL4 binding sites linked to a minimal promoter) and thereby stimulates the transcription of the desired transgene (Wang et al., Proc . Na tl . Acad . Sci . USA 91:8180- 84 (1994); Wang et al . , Nature Biotechnology 15:239-243 (1997a); Wang et al . , Gene Therapy 4:432-41 (1997b)).
  • the GeneSwitch may be used to regulate the expression of a plasmid for nucleic acid-based vaccines. It is possible that the timing of expression may influence the immune response. Thus, with a GeneSwitch regulated system, the genes that encode the multiepitopes may be turned on at a defined time after DNA delivery by the administration of the ligand (mifepristone) to the animal. If the expression plasmid persists in vivo for a long enough time, the GeneSwitch system also can be used to provide pulsatile expression of the multiepitope gene products. An example of the design of a system that is regulated by the GeneSwitch is shown in Figure 10.

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Chemical & Material Sciences (AREA)
  • General Health & Medical Sciences (AREA)
  • Microbiology (AREA)
  • Engineering & Computer Science (AREA)
  • Medicinal Chemistry (AREA)
  • Immunology (AREA)
  • Animal Behavior & Ethology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Epidemiology (AREA)
  • Mycology (AREA)
  • Biomedical Technology (AREA)
  • Organic Chemistry (AREA)
  • Biotechnology (AREA)
  • General Engineering & Computer Science (AREA)
  • Zoology (AREA)
  • Wood Science & Technology (AREA)
  • Physics & Mathematics (AREA)
  • Plant Pathology (AREA)
  • Molecular Biology (AREA)
  • Biochemistry (AREA)
  • Biophysics (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)

Abstract

L'invention porte sur des plasmides et sur des procédés améliorés utilisés sur des vaccins à base d'acide nucléique; sur l'utilisation d'épitopes, de petites séquences de protéines à application immunologique qui sont capables d'induire des réponses cellulaires et humorales déclenchant une réponse immune protectrice ou thérapeutique contre des agents pathogènes nombreux et complexes, et qui sont destinés à être incorporés dans des vaccins à base d'acide nucléique; sur les structures et les caractéristiques des systèmes d'expression génique pouvant exprimer plusieurs épitopes; sur un procédé d'immunisation génétique consistant à introduire plusieurs épitopes dans un organisme nécessitant cette immunisation; sur un plasmide d'expression de plusieurs épitopes comprenant une séquence d'acide nucléique codant plusieurs épitopes, chacun de ces épitopes pouvant générer une réponse immune; sur un système d'évaluation polyvalent tel qu'indiqué en figure 8 et sélectionné dans le groupe comprenant deux plasmides, deux gènes, un IRES (site d'entrée ribosomique interne) et un épissage alternatif, et sur un procédé de fabrication d'un plasmide produisant la séquence d'acide nucléique appropriée.
PCT/US1998/023745 1997-11-12 1998-11-09 Plasmides d'expression utilises dans des vaccins a base d'acide nucleique a plusieurs epitopes Ceased WO1999024596A1 (fr)

Priority Applications (3)

Application Number Priority Date Filing Date Title
CA002309344A CA2309344A1 (fr) 1997-11-12 1998-11-09 Plasmides d'expression utilises dans des vaccins a base d'acide nucleique a plusieurs epitopes
EP98957671A EP1030927A1 (fr) 1997-11-12 1998-11-09 Plasmides d'expression utilises dans des vaccins a base d'acide nucleique a plusieurs epitopes
AU13872/99A AU1387299A (en) 1997-11-12 1998-11-09 Expression plasmids for multiepitope nucleic acid-based vaccines

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US6513497P 1997-11-12 1997-11-12
US60/065,134 1997-11-12

Publications (1)

Publication Number Publication Date
WO1999024596A1 true WO1999024596A1 (fr) 1999-05-20

Family

ID=22060567

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US1998/023745 Ceased WO1999024596A1 (fr) 1997-11-12 1998-11-09 Plasmides d'expression utilises dans des vaccins a base d'acide nucleique a plusieurs epitopes

Country Status (4)

Country Link
EP (1) EP1030927A1 (fr)
AU (1) AU1387299A (fr)
CA (1) CA2309344A1 (fr)
WO (1) WO1999024596A1 (fr)

Cited By (10)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2004018666A1 (fr) * 2002-08-20 2004-03-04 Mannkind Corporation Vecteurs d'expression codant pour des epitopes d'antigenes associes a une cible
WO2003063770A3 (fr) * 2001-11-07 2004-04-01 Mankind Corp Epitopes codant pour des vecteurs d'expression d'antigenes associes a des cibles et procedes permettant leur conception
US6861234B1 (en) 2000-04-28 2005-03-01 Mannkind Corporation Method of epitope discovery
JP2007529223A (ja) * 2004-03-15 2007-10-25 バイオジェン・アイデック・エムエイ・インコーポレイテッド 選択的スプライシングを用いて真核細胞においてポリペプチドマルチマーを発現するための方法および構築物
WO2007087851A3 (fr) * 2006-02-01 2007-12-21 Tgcbiomics Gmbh Procédé de production d'epitomeres et leur utilisation de ces derniers sur des micro-organismes porteurs
US7807780B2 (en) * 2000-07-21 2010-10-05 Revance Therapeutics, Inc. Multi-component biological transport systems
US8974774B2 (en) 2004-03-03 2015-03-10 Revance Therapeutics, Inc. Compositions and methods for topical diagnostic and therapeutic transport
US9180081B2 (en) 2005-03-03 2015-11-10 Revance Therapeutics, Inc. Compositions and methods for topical application and transdermal delivery of botulinum toxins
US9211248B2 (en) 2004-03-03 2015-12-15 Revance Therapeutics, Inc. Compositions and methods for topical application and transdermal delivery of botulinum toxins
US11564998B2 (en) 2012-04-02 2023-01-31 Modernatx, Inc. Modified polynucleotides for the production of cytoplasmic and cytoskeletal proteins

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1991002805A2 (fr) * 1989-08-18 1991-03-07 Viagene, Inc. Retrovirus de recombinaison apportant des constructions de vecteur a des cellules cibles
WO1994024870A1 (fr) * 1993-01-20 1994-11-10 Biotransplant, Inc. Vecteurs retroviraux capables d'exprimer des proteines multimeres a partir de sites multiples d'initiation de traduction
WO1996003144A1 (fr) * 1994-07-27 1996-02-08 The Council Of The Queensland Institute Of Medical Research Vaccins a base de polyepitopes
DE19514310A1 (de) * 1995-04-18 1996-10-24 Univ Ludwigs Albert Vektoren zur Transfektion von eukaryotischen Zellen, deren Verwendung und damit transfizierte Zielzellen
WO1997035021A2 (fr) * 1996-03-20 1997-09-25 The Government Of The United States Of America, Represented By The Secretary, Department Of Health And Human Services Peptide d'oligo-epitope de l'antigene prostatique specifique

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1991002805A2 (fr) * 1989-08-18 1991-03-07 Viagene, Inc. Retrovirus de recombinaison apportant des constructions de vecteur a des cellules cibles
WO1994024870A1 (fr) * 1993-01-20 1994-11-10 Biotransplant, Inc. Vecteurs retroviraux capables d'exprimer des proteines multimeres a partir de sites multiples d'initiation de traduction
WO1996003144A1 (fr) * 1994-07-27 1996-02-08 The Council Of The Queensland Institute Of Medical Research Vaccins a base de polyepitopes
DE19514310A1 (de) * 1995-04-18 1996-10-24 Univ Ludwigs Albert Vektoren zur Transfektion von eukaryotischen Zellen, deren Verwendung und damit transfizierte Zielzellen
WO1997035021A2 (fr) * 1996-03-20 1997-09-25 The Government Of The United States Of America, Represented By The Secretary, Department Of Health And Human Services Peptide d'oligo-epitope de l'antigene prostatique specifique

Cited By (19)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6861234B1 (en) 2000-04-28 2005-03-01 Mannkind Corporation Method of epitope discovery
US7807780B2 (en) * 2000-07-21 2010-10-05 Revance Therapeutics, Inc. Multi-component biological transport systems
US7232682B2 (en) 2001-11-07 2007-06-19 Mannkind Corporation Expression vectors encoding epitopes of target-associated antigens and methods for their design
CN1313617C (zh) * 2001-11-07 2007-05-02 曼康公司 编码靶相关抗原表位的表达载体及其设计方法
WO2003063770A3 (fr) * 2001-11-07 2004-04-01 Mankind Corp Epitopes codant pour des vecteurs d'expression d'antigenes associes a des cibles et procedes permettant leur conception
US8252916B2 (en) 2001-11-07 2012-08-28 Mannkind Corporation Expression vectors encoding epitopes of target-associated antigens and methods for their design
US8637305B2 (en) 2001-11-07 2014-01-28 Mannkind Corporation Expression vectors encoding epitopes of target-associated antigens and methods for their design
WO2004018666A1 (fr) * 2002-08-20 2004-03-04 Mannkind Corporation Vecteurs d'expression codant pour des epitopes d'antigenes associes a une cible
US8974774B2 (en) 2004-03-03 2015-03-10 Revance Therapeutics, Inc. Compositions and methods for topical diagnostic and therapeutic transport
US10172877B2 (en) 2004-03-03 2019-01-08 Revance Therapeutics, Inc. Compositions and methods for topical diagnostic and therapeutic transport
US9211248B2 (en) 2004-03-03 2015-12-15 Revance Therapeutics, Inc. Compositions and methods for topical application and transdermal delivery of botulinum toxins
EP1733034A4 (fr) * 2004-03-15 2010-01-20 Biogen Idec Inc Methodes et constructions permettant d'exprimer des multimeres polypeptidiques dans des cellules eucaryotes au moyen de l'epissage alternatif
JP2007529223A (ja) * 2004-03-15 2007-10-25 バイオジェン・アイデック・エムエイ・インコーポレイテッド 選択的スプライシングを用いて真核細胞においてポリペプチドマルチマーを発現するための方法および構築物
US9180081B2 (en) 2005-03-03 2015-11-10 Revance Therapeutics, Inc. Compositions and methods for topical application and transdermal delivery of botulinum toxins
US9314416B2 (en) 2005-03-03 2016-04-19 Revance Therapeutics, Inc. Compositions and methods for topical application and transdermal delivery of botulinum toxins
US10080786B2 (en) 2005-03-03 2018-09-25 Revance Therapeutics, Inc. Methods for treating pain by topical application and transdermal delivery of botulinum toxin
US10744078B2 (en) 2005-03-03 2020-08-18 Revance Therapeutics, Inc. Compositions and methods for topical application and transdermal delivery of botulinum toxins
WO2007087851A3 (fr) * 2006-02-01 2007-12-21 Tgcbiomics Gmbh Procédé de production d'epitomeres et leur utilisation de ces derniers sur des micro-organismes porteurs
US11564998B2 (en) 2012-04-02 2023-01-31 Modernatx, Inc. Modified polynucleotides for the production of cytoplasmic and cytoskeletal proteins

Also Published As

Publication number Publication date
EP1030927A1 (fr) 2000-08-30
AU1387299A (en) 1999-05-31
CA2309344A1 (fr) 1999-05-20

Similar Documents

Publication Publication Date Title
Dietrich et al. Delivery of antigen-encoding plasmid DNA into the cytosol of macrophages by attenuated suicide Listeria monocytogenes
US9950081B2 (en) Vectors and methods for genetic immunization
Ciernik et al. Induction of cytotoxic T lymphocytes and antitumor immunity with DNA vaccines expressing single T cell epitopes.
EP1543020B1 (fr) Plasmides synthetiques optimises par codons
JP3881014B2 (ja) ポリヌクレオチドツベクローシスワクチン
KR101454842B1 (ko) 개선된 알파바이러스 레플리콘 및 헬퍼 구축물
JP2004528012A (ja) 改善された免疫応答を誘導し得る発現ベクターおよびこのベクターの使用方法
EP1435387A1 (fr) Micro-organismes oncolytiques exprimant les proteines hsp, et leurs utilisations
AU2054799A (en) Delivery of polypeptide-encoding plasmid dna into the cytosol of macrophages by attenuated suicide bacteria
CA2257137A1 (fr) Genes synthetiques de l'hepatite c
WO1999024596A1 (fr) Plasmides d'expression utilises dans des vaccins a base d'acide nucleique a plusieurs epitopes
JP2001516210A (ja) 遺伝子治療による炎症細胞の自己調節されたアポトーシス
ES2834329T3 (es) Vacuna de ADN tolerogénica
AU2006259041A1 (en) Agents and methods based on the use of the EDA domain of fibronectin
EP1844142B1 (fr) Vecteurs et procédés pour l'immunisation génétique
Azevedo et al. Main features of DNA-based immunization vectors
JP2780961B2 (ja) 単純ヘルペスウイルスタンパク質およびそれを含む ワクチン
AU761497B2 (en) Self-regulated apoptosis of inflammatory cells by gene therapy
KR20220038755A (ko) 한타바이러스 항원성 조성물
JP4411213B2 (ja) ネコ白血病ウィルスによるネコ感染症のようなオンコウィルス感染症に対するワクチン
CN101379078A (zh) 爱泼斯坦-巴尔病毒相关疾病的治疗
JPH09509562A (ja) 新規なタンパク質／ポリペプチド及びコトランスフェクションプラスミド並びにその組換え生キャリヤー
WO2002055104A2 (fr) Vaccin contre l'acne
WO2000028048A1 (fr) Nouveaux plasmides destines a etre utilises en medecine et leur technique de production
US20220257747A1 (en) Methods and compositions of astrovirus replicons

Legal Events

Date Code Title Description
AK Designated states

Kind code of ref document: A1

Designated state(s): AL AM AT AU AZ BA BB BG BR BY CA CH CN CU CZ DE DK EE ES FI GB GD GE GH GM HR HU ID IL IS JP KE KG KP KR KZ LC LK LR LS LT LU LV MD MG MK MN MW MX NO NZ PL PT RO RU SD SE SG SI SK SL TJ TM TR TT UA UG US UZ VN YU ZW

AL Designated countries for regional patents

Kind code of ref document: A1

Designated state(s): GH GM KE LS MW SD SZ UG ZW AM AZ BY KG KZ MD RU TJ TM AT BE CH CY DE DK ES FI FR GB GR IE IT LU MC NL PT SE BF BJ CF CG CI CM GA GN GW ML MR NE SN TD TG

121 Ep: the epo has been informed by wipo that ep was designated in this application
DFPE Request for preliminary examination filed prior to expiration of 19th month from priority date (pct application filed before 20040101)
WWE Wipo information: entry into national phase

Ref document number: 13872/99

Country of ref document: AU

ENP Entry into the national phase

Ref document number: 2309344

Country of ref document: CA

Ref country code: CA

Ref document number: 2309344

Kind code of ref document: A

Format of ref document f/p: F

ENP Entry into the national phase

Ref country code: JP

Ref document number: 2000 519590

Kind code of ref document: A

Format of ref document f/p: F

NENP Non-entry into the national phase

Ref country code: KR

WWE Wipo information: entry into national phase

Ref document number: 1998957671

Country of ref document: EP

WWP Wipo information: published in national office

Ref document number: 1998957671

Country of ref document: EP

REG Reference to national code

Ref country code: DE

Ref legal event code: 8642

WWW Wipo information: withdrawn in national office

Ref document number: 1998957671

Country of ref document: EP