RU2787521C2 - Binding of fused protein to cd47 protein and its use - Google Patents

Abstract

FIELD: biotechnology.

SUBSTANCE: present invention relates to the field of biotechnology, specifically to binding CD47 proteins; it can be used in medicine for therapy of CD47-positive hematological tumors and CD47-positive solid tumors, as well as autoimmune diseases, such as Crohn’s disease, allergic asthma, and rheumatoid arthritis.

EFFECT: proposed invention provides obtainment of protein, which has following characteristics: 1) specific binding to CD47 protein with relatively high affinity; 2) specific blocking of interaction of CD47 protein with SIRPalpha; 3) it does not cause blood clotting reaction; 4) inhibition of growth and/or proliferation of tumors or tumor cells; 5) blocking of an apoptotic signal induced by interaction of CD47/SIRPalpha; and/or 6) it is safe for a human and does not have side effects causing damage to the body.

15 cl, 20 dwg, 2 tbl, 14 ex

Description

Technical field

[0001] The present application relates to the binding of a fusion protein to the CD47 protein and its use. Said fusion protein can specifically block the interaction between CD47 protein and SIRPα without inducing a blood coagulation reaction, and can inhibit the growth and/or proliferation of tumors or tumor cells.

State of the art

[0002] The CD47 protein is a transmembrane glycoprotein that is a member of the immunoglobulin superfamily and is expressed on the surface of various cells, including erythrocytes. CD47 ligands include integrins, thrombospondin-1, and signaling proteins (SIRPs). CD47 affects a number of biological functions, including cell migration, T cells, dendritic cell activation, axonal development, and more. In addition, by interacting with SIRPα, CD47 can inhibit macrophage phagocytosis and protect normal cells such as blood cells and the like from phagocytosis by macrophages. Studies have shown that, in addition to normal tissue cells expressing CD47, many tumor cells overexpress CD47 and prevent tumor cell phagocytosis by macrophages by combining with SIRPα on the macrophage surface. This is seen as a mechanism by which tumors elude the body's immune surveillance. Blocking the interaction between the CD47 protein and SIRPα can suppress tumor growth. (Theocharides APA, et al., 2012).

[0003] However, existing reagents for blocking the interaction between the CD47 protein and SIRPα have limited recognition activity. They tend to have insufficient affinity for the CD47 protein and limited ability to inhibit tumors. In another aspect, existing drugs with antibodies targeting CD47 have side effects that cause anemia or thrombocytopenia (Yinpeng Bai et al., Chin J Clin Oncol., 2017 Vol 44. No. 7). There is an urgent need to develop a new therapy that can effectively block the interaction between the CD47 protein and SIRPα with fewer side effects.

SUMMARY OF THE INVENTION

[0004] The present application relates to the binding of a fusion protein to the CD47 protein and its use. The fusion protein can specifically bind to the CD47 protein. The fusion protein of the present application has at least one of the following characteristics: 1) specific binding to the CD47 protein with relatively high affinity; 2) specific blocking of the interaction of the CD47 protein with SIRPα; 3) does not cause a blood coagulation reaction; 4) inhibition of growth and/or proliferation of tumors or tumor cells; 5) blocking the apoptotic signal induced by the CD47/SIRPα interaction; and/or 6) is safe for humans and does not have side effects harmful to the body. The present application further relates to a method for producing and using a fusion protein.

[0005] In one aspect, the present application relates to a fusion protein capable of specifically binding to the CD47 protein, and having at least one of the following characteristics: 1) binding to the CD47 protein with a KD value of 1×10 ^-8 M or smaller; 2) specific blocking of the interaction of the CD47 protein with SIRPα; 3) does not cause a blood coagulation reaction; and 4) inhibition of growth and/or proliferation of tumors or tumor cells.

[0006] In some embodiments, said CD47 protein is a human CD47 protein.

[0007] In some embodiments, said CD47 protein is a CD47 protein expressed on the surface of cells.

[0008] In some embodiments, said tumors or tumor cells are CD47 positive.

[0009] In some embodiments, said tumors are selected from the group consisting of CD47 positive hematologic tumors and/or CD47 positive solid tumors.

[0010] In some embodiments, said fusion protein comprises a human SIRPα domain that can specifically bind to said CD47 protein and an immunoglobulin Fc region, wherein said human SIRPα domain is directly or indirectly linked to an immunoglobulin Fc region.

[0011] In some embodiments, said human SIRPα domain comprises an extracellular domain of human SIRPα, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions. In some embodiments, said human SIRPα domain comprises a human SIRPα IgV domain, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions. In some embodiments, said human SIRPα domain comprises a variant 1 human SIRPα domain, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions. In some embodiments, said human SIRPα domain comprises a human SIRPα variant 1 IgV domain, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions. In some embodiments, said human SIRPα domain comprises amino acid residues at positions 33-149 of human SIRPα variant 1, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions.

[0012] In some embodiments, the implementation of the human SIRPα domain according to the present application contains the amino acid sequence specified in one of SEQ ID NO: 1-20, 62-65, and the amino acid sequence having at least 80% with it (for example, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or at least 100%) sequence homology.

[0013] In some embodiments, the implementation of the fusion protein according to the present application contains the amino acid sequence specified in one of SEQ ID NO: 21-61, and an amino acid sequence having with it at least 80% (for example, at least 85 %, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96% , at least 97%, at least 98%, at least 99%, or at least 100%) sequence homology.

[0014] In some embodiments, said human SIRPα domain, fragment, or variant thereof, which contains one or more amino acid substitutions, includes substitutions, deletions, or additions of one or more amino acid residues.

[0015] In some embodiments, said mutant contains amino acid substitutions at one or more residues selected from the group consisting of I61, V63, E77, Q82, K83, E84, V93, D95, D96, K98, N100, R107, G109, and V132.

[0016] In some embodiments, said mutant contains one or more amino acid substitutions selected from the group consisting of R22C, I29L, I61L/V/F, V63I, E77I/N/Q/K/H/M/R/N/ V/L, Q82S/R/G/N, K83R, E84K/H/D/R/G, V93L/A, D95H/R/E, D96S/T, K98R, N100G/K/D/E, R107N/ S, G109R/H and V132L/R/I/S.

[0017] In some embodiments, the implementation of the specified domain of human SIRPα contains the amino acid sequence specified in one of SEQ ID NO: 1-20, 62-65.

[0018] In some embodiments, said immunoglobulin Fc region comprises an IgG Fc region.

[0019] In some embodiments, said IgG is a human IgG. In some embodiments, said IgG is selected from the group consisting of IgG1 and/or IgG4.

[0020] In some embodiments, said human SIRPα domain is located at the N-terminus of said immunoglobulin Fc region.

[0021] In some embodiments, said human SIRPα domain is linked to an immunoglobulin Fc region via a linker.

[0022] In some embodiments, the implementation of the specified Fc-region of the immunoglobulin contains the amino acid sequence specified in one of SEQ ID NO: 67-68.

[0023] In some embodiments, the implementation of the specified fusion protein contains the amino acid sequence specified in one of SEQ ID NO: 21-61.

[0024] In another aspect, the present application relates to a nucleic acid molecule encoding a fusion protein according to the present application.

[0025] In another aspect, the present application relates to a vector containing the nucleic acid molecule according to the present application.

[0026] In another aspect, the present application relates to a host cell containing the nucleic acid molecule according to the present application or the vector according to the present application.

[0027] In another aspect, the present application relates to a method for producing a fusion protein according to the present application, a method comprising culturing a host cell according to the present application under conditions that allow the fusion protein to be expressed.

[0028] In another aspect, the present application relates to a composition containing a fusion protein, a nucleic acid molecule, a vector and/or a host cell according to the present application, and optionally pharmaceutically acceptable adjuvants.

[0029] In another aspect, the present application relates to the use of a fusion protein, a nucleic acid molecule, a vector, a host cell and / or a composition according to the present application for the preparation of a drug and / or kit, where the specified drug and / or kit is intended for prevention or treatment of tumors or autoimmune diseases. In some embodiments, the tumors are selected from the group consisting of CD47 positive hematologic tumors and/or CD47 positive solid tumors. In some embodiments, the autoimmune diseases are selected from the group consisting of Crohn's disease, allergic asthma, and rheumatoid arthritis.

[0030] In another aspect, the present application relates to a method of blocking the interaction between the CD47 protein and SIRPα, a method comprising the introduction of a fusion protein or composition according to the present application.

[0031] In another aspect, the present application relates to a method of inhibiting the growth and/or proliferation of tumors or tumor cells, a method comprising contacting the fusion protein or composition of the present application with tumors or tumor cells. In some embodiments, contact occurs in vitro .

[0032] In another aspect, the present application relates to a method for preventing or treating tumors or autoimmune diseases in an individual, a method comprising administering to the individual an effective amount of a fusion protein or composition according to this application. In some embodiments, the tumors are selected from the group consisting of CD47 positive hematologic tumors and/or CD47 positive solid tumors. In some embodiments, the autoimmune diseases are selected from the group consisting of Crohn's disease, allergic asthma, and rheumatoid arthritis.

[0033] Those skilled in the art will readily recognize other aspects and advantages of the present invention from the detailed description that follows. The detailed description below shows and describes only illustrative embodiments of the present invention. As will be apparent to those skilled in the art, the scope of the present invention allows those skilled in the art to modify the specific embodiments described without deviating from the spirit and scope of the invention as set forth in this application. Accordingly, the drawings and description of the present application are illustrative only and not restrictive.

BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Specific features of the invention relevant to the present application are shown in the appended claims. Referring to the illustrative embodiments detailed below and the accompanying drawings, the features and advantages of the present invention can be better understood. A brief description of the attached drawings is as follows:

[0035] FIG. 1 shows a physical structural diagram of the pTM vector.

[0036] FIG. 2 shows a schematic representation of a method for detecting interactions between truncated SIRPα domains and their mutants and CD47.

[0037] FIG. 3 shows the results of an enrichment screen for SIRPα truncated domain mutants by flow cytometry.

[0038] FIG. 4 shows the sequence alignment of the truncated SIRPα domains and their variants.

[0039] FIG. 5 shows the results of recognition by the fusion protein of the present application of the CD47 protein.

[0040] FIG. 6A-6B show the specificity of the fusion protein of the present application for recognition of the human CD47 protein.

[0041] FIG. 7 shows recognition by the fusion protein of the present application of the human CD47 protein and other proteins.

[0042] FIG. 8 shows that the fusion protein of the present application and TTI-621 competitively block the binding of the CD47 protein to its SIRPα ligand.

[0043] FIG. 9A-9C show the results of recognition by the present fusion protein of Raji cells, Jurkat cells, and CD47 protein on the surface of A549 cells.

[0044] FIG. 10A-10C show the recognition results of the fusion protein of the present application and TTI-621 of Raji cells, Jurkat cells, and CD47 protein on the surface of A549 cells.

[0045] FIG. 11 shows the results of the fusion protein of the present application and Hu5F9-G4 in coagulation resistance.

[0046] FIG. 12A-12B show the tumor-inhibiting activity of the fusion protein of the present application.

[0047] FIG. 13A-13B show the effects of the fusion protein of the present application on erythrocytes and platelets compared to TTI-621.

[0048] FIG. 14 shows the results of recognition by the fusion protein of the present application of the CD47 protein.

[0049] FIG. 15 shows that the fusion protein of the present application competitively blocks the binding of the CD47 protein to its SIRPα ligand.

[0050] FIG. 16 shows the results of the fusion protein of the present application and Hu5F9-G4 in coagulation resistance.

[0051] FIG. 17 shows that the fusion protein of the present application effectively blocks CD47-Fc-induced apoptosis of Jurkat-CSR cells.

[0052] FIG. 18A shows the effects of the fusion protein of the present application on the level of erythrocytes in the peripheral blood of mice.

[0053] FIG. 18B shows the effects of the fusion protein of the present application on the level of platelets in the peripheral blood of mice.

[0054] FIG. 19 shows the inhibitory effect of the fusion protein of the present application on tumor growth in mice.

[0055] FIG. 20 shows the effects of the fusion protein of the present application on the body weight of mice.

DETAILED DESCRIPTION OF EMBODIMENTS

[0056] Hereinafter, embodiments of the invention used in this application are described by way of specific examples. Specialists in this field can easily understand other advantages and effects of the invention described in this application, from the content disclosed in the present description.

FUNCTION PROTEIN

[0057] In one aspect, the present application relates to a fusion protein that can specifically bind to the CD47 protein with a KD value of 1×10 ^-8 M or less, for example, with a KD value of not more than 9×10 ^-9 M, not more than 8×10 ^-9 M, not more than 7×10 ^-9 M, not more than 6.2×10 ^-9 M, not more than 6×10 ^-9 M, not more than 5×10 ^-9 M , not more than 4.8×10 ^-9 M, not more than 4.5×10 ^-9 M, not more than 2×10 ^-9 M, not more than 1.5×10 ^-9 M, or not more than 1×10 ^-10 M or less.

[0058] In some cases, the fusion protein of the present application may also specifically block the interaction between the CD47 protein and SIRPα, thereby activating macrophages for phagocytosis of tumor cells or inhibiting the apoptotic signal of certain specific cells. In addition, the fusion protein of the present application may not cause a blood coagulation reaction. For example, said fusion protein and erythrocyte solution were added to a hemagglutination test plate, and then the erythrocytes were sunk to the bottom of the well instead of flattening into a grid. The fusion protein may further inhibit the growth and/or proliferation of tumors or tumor cells, for example, it may reduce the area or size of a tumor or may increase the survival rate of an individual with a tumor. The fusion protein is also safe to administer to an individual because it will not adversely affect the body weight and/or mortality rate of the individual. In addition, the fusion protein of the present application is easy to prepare and obtain, not limited to the source of the Fc region of a specific immunoglobulin.

[0059] As used herein, the term "fusion protein" generally refers to a complex polypeptide, i.e., a single continuous amino acid sequence consisting of two or more polypeptides. The fusion protein can generally be produced artificially by recombinant nucleic acid or by chemical synthesis.

[0060] In this application, the term "CD47 protein" is also known as integrin-associated protein (IAP), which refers to the immunoglobulin superfamily. The CD47 protein can bind to membrane integrins, thrombospondin-1 (TSP-1) or signal regulatory protein alpha (SIRPα). The CD47 protein can be expressed on the surface of the cell membrane. Said CD47 protein may be a supramolecular complex consisting of certain IAPs, G proteins and cholesterol. In the present application, said CD47 protein may be human CD47 protein with accession number CEJ95640.1 in the GenBank database. In the present application, the CD47 protein may include the amino acid sequence shown in SEQ ID NO: 66.

[0061] As used herein, the term "CD47-positive" generally refers to the expression in the body or on the surface of cells of a characteristic of a CD47 protein, fragment or variant thereof that contains one or more substitutions. CD47 positive cells may be CD47 overexpressing cells. A CD47-positive cell can mainly serve as a sign of diseases. For example, in the case of a disease, the density of said CD47 protein on the surface of a CD47 positive cell will exceed the density of the CD47 protein of the same type in cells under normal conditions. In some embodiments, said tumors or tumor cells may be CD47 positive. For example, said tumors may be selected from the group consisting of CD47 positive hematologic tumors and/or CD47 positive solid tumors.

[0062] In this application, the term " K _D " can be used interchangeably with "KD", and generally refers to the dissociation constant of a specific antigen-antibody interaction per unit M (mol/l). KD can be calculated from the concentration of material AB and dissociated materials A and B: KD=c(A)×c(B)/c(AB). It can be seen from the formula that the larger the KD value, the greater the dissociation and the weaker the affinity of materials A and B; on the contrary, the lower the KD value, the smaller the dissociation and the stronger the affinity of materials A and B.

[0063] As used herein, the term "SIRPα" generally refers to a regulatory membrane glycoprotein from the SIRP family. This SIRPα can recognize the CD47 protein and can serve as a ligand for the CD47 protein. SIRPα is a transmembrane protein that can have three regions in its extracellular region, similar to regions of the immunoglobulin family, where the region at the N-terminus mediates binding to CD47. Said SIRPα is natively expressed on the surface of macrophages, dendritic cells and nerve cells. The cytoplasmic region of said SIRPα is highly conserved in rats. This SIRPα exhibits a polymorphism, which, however, does not affect its recognition and binding to the CD47 protein.

[0064] As used herein, the term "human SIRPα domain" generally includes human SIRPα, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions. As used herein, said human SIRPα domain may comprise an extracellular domain of human SIRPα, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions. Said human SIRPα domain may comprise a human SIRPα IgV domain, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions. Said human SIRPα domain may comprise a variant 1 human SIRPα domain, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions. Said human SIRPα domain may include amino acid residues at positions 33-149 of human SIRPα variant 1, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions. In humans, the SIRPα protein is originally of two types, one type (human SIRPα variant 1 or Type V1) has an amino acid sequence with GeneBank accession number NP_542970.1 (its amino acid sequence is set forth in SEQ ID NO: 62, where amino acid residues at positions 31 -504 may constitute the mature SIRPα domain). The other type (Variant 2 or Type V2) differs by 13 amino acids from Variant 1 or Type V1 and its amino acid sequence has GeneBank accession number CAA71403.1.

[0065] As used herein, said human SIRPα domain may comprise an extracellular domain of human SIRPα, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions.

[0066] As used herein, the term "extracellular domain" generally refers to the functional structural region of a protein located outside the cell membrane. In some embodiments, said extracellular domain may refer to a human SIRPα domain, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions. For example, said extracellular domain from human SIRPα domains may contain 3 immunoglobulin superfamily (IgSF) domains and multiple glycosylation sites. Said extracellular domain from human SIRPα domains can bind to a specific ligand (eg, CD47 protein), including a signal transduction function. This extracellular domain from human SIRPα domains can also be activated by a number of mitogens such as serum, insulin, growth factors, EGF, PDGF and neurotrophic factors and phosphorylated.

[0067] As used herein, said human SIRPα domain may comprise a human SIRPα IgV domain, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions. As used herein, said human SIRPα domain may include human SIRPα variant 1, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions. For example, said human SIRPα domain may include amino acid residues at positions 33-149 of human SIRPα variant 1, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions.

[0068] As used herein, the term "IgV domain" generally refers to an IgV-like domain that is analogous to the variable domain of an antibody. The immunoglobulin domain can be subdivided into four classes: IgV, IgC1, IgC2 and IgI. The IgV domain can be present in various protein families, and contains the light chain and heavy chain, the immunoglobulin T-cell receptor. Human SIRPα may be highly polymorphic in the IgV domain. For example, said human SIRPα variant 1 IgV domain can mediate binding of the human SIRPα domain to human CD47 protein (Seiffert, M. et al. (2001) Blood 97,2741-9; Vernon-ffilson, E. F. et al. (2000) Eur J Immunol 30, 2130-7).

[0069] For example, said human SIRPα variant 1, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions may comprise the amino acid sequence set forth in SEQ ID NO: 62. For example, said human SIRPα domain may comprise an IgV domain of human SIRPα variant 1, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions. For example, said IgV domain from human SIRPα variant 1, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions, may contain the amino acid sequence set forth in SEQ ID NO: 65 (i.e., residues at amino acid positions 38-145 sequence set forth in SEQ ID NO: 62). Alternatively, for example, said human SIRPα domain may comprise a truncated human SIRPα variant 1 domain, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions. Said variant 1 human SIRPα truncated domain, fragment thereof, or variant thereof which contains one or more amino acid substitutions may comprise the amino acid sequence set forth in SEQ ID NO: 63 (i.e., residues at positions 33-149 of the amino acid sequence set forth in in SEQ ID NO: 62).

[0070] The human SIRPα domain of the present application may contain the amino acid sequence set forth in any one sequence selected from the group consisting of SEQ ID NOs: 1-20 and 62-65.

[0071] As used herein, the term "mutant" generally refers to a protein, polypeptide, or amino acid sequence in which a mutation occurs. Said mutation may refer to a difference from the wild type. For example, said mutation may be a structural amino acid sequence change that occurs on a wild type basis. For example, the specified wild type may be a typical phenotype without the specified structural change. For example, said mutant can be obtained by mutating a human SIRPα domain, a fragment thereof (comprising a human SIRPα IgV domain, a fragment thereof, a human SIRPα variant 1 domain, a fragment thereof, or amino acid residues at positions 33-149 of a human SIRPα variant 1, fragment thereof) that considered as wild type.

[0072] In this application, the mutant may contain amino acid substitutions at one or more residues selected from the group consisting of I61, V63, E77, Q82, K83, E84, V93, D95, D96, K98, N100, R107, G109, and V132. The indicated amino acid residue positions for amino acid substitutions may represent the exact residue number when used as a reference for the amino acid sequence set forth in SEQ ID NO: 62, where "residue X _n " refers to an X residue that corresponds to position n in the amino acid sequence set forth in SEQ ID NO: 62. SEQ ID NO: 62, where n is a positive integer, X is an abbreviation for any amino acid residue. For example, "residue I61" refers to amino acid residue I, which corresponds to position 61 in the amino acid sequence set forth in SEQ ID NO:62.

[0073] As used herein, “amino acid substitution X _n ” refers to a substitution of the amino acid found at amino acid residue X at position n of the amino acid sequence set forth in SEQ ID NO: 62, where n is a positive integer, X is an abbreviation for any amino acid residue . For example, "I61 amino acid substitutions" refers to an amino acid substitution found at amino acid residue I, which corresponds to position 61 in the amino acid sequence set forth in SEQ ID NO:62.

[0074] As used herein, a specific amino acid residue in a specific amino acid sequence "corresponding" to a specific amino acid residue in another amino acid sequence generally refers to the corresponding amino acid residue relatedness obtained by amino acid sequence alignment under optimal conditions. Said sequence alignment can be performed in a manner that is understood by those skilled in the art, eg using BLAST, BLAST-2, ALIGN, NEEDLE or Megalign (DNASTAR) software, etc. Those skilled in the art can determine suitable parameters to apply for alignment, including any algorithm necessary to achieve optimal alignment in the compared full-length sequence.

[0075] The amino acid substitutions of this application may be non-conservative substitutions. These non-conservative substitutions may include substituting amino acid residues in the target protein or polypeptide in a non-conservative manner, such as substituting an amino acid residue having a particular side chain size or a particular characteristic (e.g., hydrophilic) with an amino acid residue having a different side chain size or other characteristic (e.g., , hydrophobic).

[0076] These amino acid substitutions may also be conservative substitutions. These conservative substitutions may include replacing amino acid residues in the target protein or polypeptide in a conservative manner, for example, replacing an amino acid residue having a particular side chain size or a particular characteristic (e.g., hydrophilic) with an amino acid residue having the same or similar side chain size or the same or a similar characteristic (eg, still hydrophilic). Such conservative substitutions generally do not significantly affect the structure or function of the protein produced. As used herein, an amino acid sequence variant that is a mutant of a fusion protein, a fragment thereof, or a variant thereof that contains one or more amino acid substitutions may contain conservative amino acid substitutions that cannot appreciably change the structure or function of the protein (e.g., the ability to block binding CD47 with its ligand).

[0077] For example, conservative substitutions in this application can be considered as conservative substitutions between amino acids in each of the following groups:

A group of amino acids with non-polar side chain(s): alanine, valine, leucine, isoleucine, proline, phenylalanine, tryptophan and methionine.

A group of uncharged amino acids with polar side chains: glycine, serine, threonine, cysteine, tyrosine, asparagine and glutamine.

A group of negatively charged amino acids with polar side chains: aspartic acid and glutamic acid.

A group of positively charged basic amino acids: lysine, arginine and histidine.

A group of amino acids with phenyl: phenylalanine, tryptophan and tyrosine.

[0078] In some embodiments, said mutant may contain one or more amino acid substitutions selected from the group consisting of I61L/V/F, V63I, E77I/N/Q/K/H/M/R/N/V/L , Q82S/R/G/N, K83R, E84K/H/D/R/G, V93L/A, D95H/R/E, D96S/T, K98R, N100G/K/D/E, R107N/S, G109R /H and V132L/R/I/S.

[0079] As used herein, an amino acid substitution "XnY/Z" means that the X residue corresponding to position n in the amino acid sequence set forth in SEQ ID NO: 62 is replaced by a Y amino acid residue or a Z amino acid residue, where n is a positive integer , X, Y, and Z are independently abbreviations for any amino acid residue, respectively, and X is different from Y or Z. ID NO: 62, substituted by amino acid residue L, V or F.

[0080] For example, the fusion protein of the present application may contain a substitution of an amino acid group selected from the group consisting of:

(1) I61L, V63I, E77I, E84K, V93L, L96S, K98R, N100G and V132 L;

(2) I61V, E77N, Q82S, K83R and E84H;

(3) I61F, V63I, K83R, E84K and V132I;

(4) I61L, E77Q, E84D, R107N and V132I;

(5) I61L, V63I, E77K, K83R, E84D and N100G;

(6) I61V, E77H, Q82R, K83R, E84H and R107S;

(7) I61L, E77I, Q82G, E84R, V93L, L96T, N100G, R107S, G109R and V132R;

(8) I61L, E77M, Q82G, K83R, E84D and V132L;

(9) I61L;

(10) I61F, D95H, L96S, G109H and V132S;

(11) I61F, D95H, L96S, K98R, G109H and V132S;

(12) I61L, E77Q, E84D, V93A, R107N and V132I;

(13) E77K, L96S, N100K, G109H and V132L;

(14) I61L, V63I, Q82G, E84G, D95R, L96S, N100D and V132I;

(15) I61L, E77R, Q82N, K83R, E84G, V93L, D95E, L96T, K98R, N100D and V132L;

(16) I61V, E77N, Q82S, K83R, E84H and V93A;

(17) I61V, V63I, E77V, K83R, E84D, D95E, L96T, K98R and N100E;

(18) I61L, V63I, E77V, K83R, D95E, L96S, K98R, N100D and G109R;

(19) I61V, E77L, Q82G, E84G, V93L, D95E, L96T, K98R and N100G; and

(20) I61L, V63I, E77N, Q82G and E84G.

[0081] In this application, variants of the SIRPα domain, respectively containing one group of amino acid substitutions from the above (1) to (20) based on the truncated domain of human SIRPα variant 1 (the amino acid sequence set forth in SEQ ID NO: 63, that is, the residues at positions 33-149 of the amino acid sequence set forth in SEQ ID NO: 62) can be sequentially named M1, M5, M12, M35, M37, M41, M57, M67, M81, M82, M84, M91, M99, M102, M111 , M122, M126, M130, M135 and M145. These mutants may sequentially contain the amino acid sequences shown in one of SEQ ID NOs: 1 to 20.

[0082] As used herein, a fusion protein comprising a truncated human SIRPα variant 1 domain (amino acid sequence set forth in SEQ ID NO: 63, namely, residues at positions 33-149 of the amino acid sequence set forth in SEQ ID NO: 62) and The human IgG1 Fc (amino acid sequence set forth in SEQ ID NO: 67) can be sequentially referred to as SS002, which contains the amino acid sequence set forth in SEQ ID NO: 61.

[0083] In this application, the term "Immunoglobulin Fc region" generally refers to the main region of the Y-shaped structure of the antibody structure, which is also called the region of the crystallizing fragment (Fc region). In IgG, IgA, and IgD antibody isotypes, the Fc region may consist of two identical protein fragments derived from the second and third constant domains of the two antibody heavy chains. The Fc regions of IgM and IgE may contain three heavy chain constant domains in each polypeptide chain. The IgG Fc region has a highly conserved N-glycosylation site. In some embodiments, said immunoglobulin Fc region may comprise an IgG Fc region. In some embodiments, said immunoglobulin Fc region may include the CH2 and CH3 regions of the heavy chain constant region. In some embodiments, said immunoglobulin Fc region may comprise a hinge region. For example, said immunoglobulin Fc region may comprise an amino acid sequence selected from any of the following sequences: SEQ ID NOs: 67-68.

[0084] In this application, the term "IgG" mainly refers to immunoglobulin G (Immunoglobulin G). IgG is one of the human immunoglobulins. According to the difference in antigenicity of the gamma chain in IgG molecules, human IgG has four subtypes: IgG1, IgG2, IgG3 and IgG4. In this application, the term "IgG1" generally refers to the single subtype with the highest proportion of IgG, which has a relatively high affinity for the Fc receptor. For example, said IgG may be human IgG. Alternatively, for example, said IgG may be selected from the group consisting of IgG1 and/or IgG4.

[0085] As used herein, said fusion protein comprises a human SIRPα domain that can specifically bind to CD47 protein and an immunoglobulin Fc region, where the human SIRPα domain can be directly or indirectly linked to an immunoglobulin Fc region. For example, the human SIRPα domain may be located at the N-terminus of the Fc region of an immunoglobulin. For example, said C-terminus of a human SIRPα domain may be directly or indirectly linked to the N-terminus of an immunoglobulin Fc. For example, said human SIRPα domain can be linked to said immunoglobulin Fc via a linker. In the present application, said linker may be a peptide linker.

[0086] In the present application, the fusion protein of the present application, respectively containing one group of amino acid substitutions from the above (1)-(20) based on the specified SS002, respectively, can be sequentially named SS002M1, SS002M5, SS002M12, SS002M35, SS002M37, SS002M41 . These fusion proteins may sequentially include the amino acid sequence set forth in SEQ ID NO: 40.

[0087] In the present application, the fusion protein of the present application, respectively containing one group of amino acid substitutions from the above (1)-(20) and human IgG4 Fc (amino acid sequence set forth in SEQ ID NO: 68) based on a truncated human SIRPα variant 1 (comprising the amino acid sequence of SEQ ID NO: 63, i.e., residues at positions 33-149 in the amino acid sequence set forth in SEQ ID NO: 62) can be sequentially named SS002M1G4, SS002M5G4, SS002M12G4, SS002M35G4, SS002M37G4, SS002M41G4, SS002M57G4, SS002M67G4, SS002M81G4, SS002M82G4, SS002M84G4, SS002M91G4, SS002M99G4, SS002M102G4, SS002M111G4, SS002M122G4, SS002M126G4, SS002M130G4, SS002M135G4 и SS002M145G4. These fusion proteins may sequentially include the amino acid sequence set forth in one of SEQ ID NO: 41 to SEQ ID NO: 60.

[0088] In some embodiments, the fusion protein of the present application may contain the amino acid sequence specified in one of SEQ ID NO: 21-SEQ ID NO: 61.

[0089] The protein, polypeptide and/or polypeptide included in the present application can also be understood as covering at least the following scope: variants or homologues with the same or similar function as the specified protein or polypeptide.

[0090] As used herein, said variants may be a protein or polypeptides resulting from a substitution, deletion, or addition of one or more amino acids compared to the amino acid sequence of said protein and said polypeptide (e.g., a human SIRPα domain, a fragment thereof, or a variant that contains one or more amino acid substitutions; or said fusion protein). For example, said functional variant may include proteins or polypeptides with amino acid changes by substitution or insertion of at least one amino acid, such as 1-30, 1-20, or 1-10, alternatively, such as 1, 2, 3, 4 or 5 substitutions, deletions and/or insertions. Said functional variant may substantially retain the biological characteristics of said protein or said polypeptide prior to change (eg, substitution, deletion, or insertion). For example, said functional variant may retain at least 60%, 70%, 80%, 90%, or 100% of the biological activity (e.g., ability to specifically bind to the CD47 protein) of said protein or said polypeptide prior to change.

[0091] As used herein, said homologue may be a protein or polypeptide that has at least about 80% (e.g., at least about 85%, about 90%, about 91%, about 92%, about 93 %, approximately 94%, approximately 95%, approximately 96%, approximately 97%, approximately 98%, approximately 99% or more) sequence homology with the amino acid sequence of the specified protein and/or specified polypeptide (for example, the human SIRPα domain, its fragment, or a variant thereof that contains one or more amino acid substitutions; or said fusion protein).

[0092] As used herein, said homology generally refers to similarity, analogy, or association between two or more sequences. "Percent sequence homology" can be calculated by comparing two sequences to be aligned in the comparison window to determine the number of positions that have the same nucleic acid base (e.g., A, T, C, G, I) or the same amino acid residue (e.g., Ala, Pro, Ser, Thr, Gly, Val, Leu, Ile, Phe, Tyr, Trp, Lys, Arg, His, Asp, Glu, Asn, Gln, Cys, and Met), and thus , get the number of matching positions, then divide the number of matching positions by the total number of positions in the comparison window (i.e., window size) and multiply the result by 100 to get the percent sequence homology. Alignment to determine percent sequence homology can be performed by a number of methods known in the art, for example using publicly available computer programs such as BLAST, BLAST-2, ALIGN or Megalign (DNASTAR). Suitable parameters for sequence alignment can be determined by those skilled in the art, including any algorithm necessary to achieve maximum alignment within the full length sequence being compared or within the region of the target sequence. Said homology can also be determined by the following methods: FASTA and BLAST. The FASTA algorithm is described, for example, in W. R. Pearson and D. J. Lipman's "Improved Tool for Biological Sequence Comparison", Proc. Natl. Acad. Sci. 85: 2444-2448, 1988; and D, J. Lipman and W. R. Pearson's "Fast and Sensitive Protein Similarity Search", Science, 227:1435-1441, 1989. For a description of the BLAST algorithm, please refer to S. Altschul, W. Gish, W. Miller, E. W. Myers and D. Lipman, "A Basic Local Alignment Search Tool", Journal of Molecular Biology, 215: 403-410, 1990.

NUCLEIC ACID MOLECULE, VECTOR, HOST CELL

[0093] In another aspect, the present application relates to one or more nucleic acid molecules capable of encoding a fusion protein according to the present application.

[0094] In some embodiments, said nucleic acid molecule may fully encode a fusion protein of the present application. For example, said fusion protein can be made using only one type of nucleic acid molecule. In some embodiments, said nucleic acid molecule may encode a portion of a fusion protein of the present application. For example, said fusion protein can be made using more than two different types of said nucleic acid molecules. For example, said nucleic acid molecule may encode said human SIRPα domains of the fusion protein of the present application (e.g., said human SIRPα extracellular domain, fragment thereof, or variant thereof that contains one or more amino acid substitutions, said human SIRPα IgV domain, fragment thereof, or a variant thereof which contains one or more amino acid substitutions, said variant 1 human SIRPα domain, a fragment thereof, or a variant thereof which contains one or more amino acid substitutions). Alternatively, for example, said nucleic acid molecule may encode an immunoglobulin Fc region from a fusion protein.

[0095] In another aspect, the present application relates to one or more vectors, which may contain one or more nucleic acid molecules according to the present application. In another aspect, the present application relates to a cell (eg, a host cell), which may contain a nucleic acid molecule according to the present application or a vector according to the present application.

[0096] As used herein, the term "nucleic acid molecule" generally refers to an isolated form of a nucleotide, deoxyribonucleotide, or ribonucleotide, or analogs thereof, of any length, isolated from their natural environment or artificially synthesized. The nucleic acid molecules of the present application can be isolated. For example, they can be prepared or synthesized by the following methods: (i) in vitro amplification, such as polymerase chain reaction (PCR) amplification, (ii) clonal recombination, (iii) purification, such as fractionation by restriction enzyme digestion and gel electrophoresis , or (iv) synthesis, eg chemical synthesis. In some embodiments, said isolated nucleic acid is a nucleic acid molecule produced using recombinant DNA technology. In the present application, the nucleic acid encoding the specified antibody or antigennegative fragment can be obtained by a number of methods known in this field. These methods include, but are not limited to, overlap-fitting PCR using restriction fragment manipulation or synthetic oligonucleotides. Specific manipulations can be found in Sambrook et al., Molecular Cloning, A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, 1989; and Ausube et al. Current Protocols in Molecular Biology, Greene Publishing and Wiley-Interscience, New York NY, 1993.

[0097] As used herein, the term "vector" generally refers to a nucleic molecule capable of self-replication in a suitable host that carries the inserted nucleic acid molecule into a host cell and/or between host cells. Said vector may include a vector that is mainly used for inserting DNA or RNA into cells, a vector that is mainly used for DNA or RNA replication, and a vector that is mainly used for DNA or RNA transcription and/or translation expression. Said vector also includes a vector with several of the functions described above. Said vector may be a polynucleotide that can be decoded and translated into a polypeptide when introduced into a suitable host cell. In general, when a suitable host cell containing said vector is cultured, said vector can produce the desired expression product. In the present application, the specified vector may include one or more of the specified nucleic acid molecules. For example, said vector may contain all of the nucleic acid molecules required to encode said fusion protein. In this case, only one vector is required to obtain the fusion protein of the present application. In some embodiments, said vector may contain a nucleic acid molecule encoding a portion of said fusion protein, for example, a nucleic acid molecule encoding a human SIRPα domain in the fusion protein of the present application. Alternatively, said vector may contain, for example, a nucleic acid molecule encoding said immunoglobulin Fc region from said fusion protein. Currently, two or more different vectors are needed to obtain the fusion protein of the present application.

[0098] In addition, the specified vector may also contain other genes, such as a marker gene, which allows the selection of the vector in a suitable host cell and in a suitable state. In addition, said vector may also include an expression control element that allows the coding region to be correctly expressed in a suitable host. Such a control element is well known to those skilled in the art. For example, they may contain promoters, ribosomal binding sites, enhancers, and other control elements that regulate gene transcription or mRNA translation. In some embodiments, said expression control sequence is a regulatory element. The specific structure of said expression control sequence may vary depending on the function of the species or cell types, but typically contains 5'-untranscribed sequences and 5'- and 3'-untranslated sequences involved in transcription and translation initiation, such as TATA- boxes, capping sequences, CAAT sequences, etc. For example, a 5' non-transcribed expression control sequence may contain a promoter region, and a promoter region may contain a promoter sequence for transcriptional control of the operably linked nucleic acid. In the present application, said vector may be a pTM vector.

[0099] In this application, the terms "host cell", "cell" and "host" are used interchangeably, and in general they refer to a plasmid or vectors, which may include or include a nucleic acid molecule according to this application, or may refer to individual cells, cell lines or cell cultures with the fusion protein of the present application, fragments or variants thereof. Said host cell may include progeny of a single host cell. Due to natural, accidental or intentional mutations, progeny cells and original parental cells may not necessarily be completely identical in morphology or genome, provided that they can express the antibodies of the present application or their antigen-binding fragments. Said host cell can be obtained by transfecting the cell in vitro with the vector of the present application. Said host cell may be a prokaryotic cell (e.g., Escherichia coli ) or a eukaryotic cell (e.g., yeast cells, e.g., COS cells, Chinese Hamster (CHO) cells, HeLa cells, HEK293 cells, COS-1 cells, NS0 cells, or myeloma cells. cells). In the present application, said host cell may be a CHO cell.

COMPOSITION, METHOD OF OBTAINING AND APPLICATION

[00100] In another aspect, the present application relates to a method for producing said fusion protein, comprising culturing a host cell under conditions that allow expression of said fusion protein.

[00101] In another aspect, the present application relates to a composition containing the specified fusion protein, the specified nucleic acid molecule, the specified vector and/or the specified host cell, and optionally a pharmaceutically acceptable adjuvant.

[00102] As used herein, the term "pharmaceutically acceptable adjuvant" may include buffers, antioxidants, preservatives, low molecular weight polypeptides, proteins, hydrophilic polymers, amino acids, sugars, chelating agents, counterions, metal complexes, and/or nonionic surfactants. , etc.

[00103] Said pharmaceutically acceptable may include buffers, antioxidants, preservatives, low molecular weight polypeptides, proteins, hydrophilic polymers, amino acids, sugars, chelating agents, counterions, metal complexes and/or nonionic surfactants, etc.

[00104] In this application, the specified pharmaceutical composition can be formulated with a pharmaceutically acceptable carrier or diluent and any other known adjuvants and excipients in accordance with generally accepted technical methods in this field, for example, following the methods in Remington: The Science and Practice of Pharmacy, nineteenth edition, published by Gennaro, Mack Publishing Co., Easton, PA, 1995.

[00105] As used herein, said composition may be formulated for oral administration, intravenous administration, intramuscular administration, in situ administration at a tumor site, inhalation, rectal administration, vaginal administration, transdermal administration, or the drug is administered via a subcutaneous depot.

[00106] In this application, the specified composition can be used to inhibit tumor growth. For example, the composition of the present application may inhibit or delay the development or progression of diseases (eg, tumors or autoimmune diseases) (eg, reduce tumor size or even substantially eliminate tumors) and/or may reduce and/or stabilize the condition of the disease.

[00107] The pharmaceutical composition according to the present application may contain a therapeutically effective amount of the specified fused protein. Said therapeutically effective amount is the dose required to prevent and/or treat (at least in part) diseases (eg, tumors or autoimmune diseases) and/or any of their complications in an individual with or at risk of the disease.

[00108] In another aspect, the present application relates to the use of a fusion protein, a nucleic acid molecule, a vector, a host cell and/or a composition according to the present application for the preparation of a drug and/or kit for use in the prevention and/or treatment of tumors or autoimmune diseases .

[00109] In this application, the term "tumor", as a rule, refers to neoplasms resulting from the proliferation of cells of local tissues of the body. Since such neoplasms are mainly presented in the form of voluminous bulges, they are also called neoplasias. According to the cellular characteristics of the neoplasms and the degree of damage to the body, tumors are divided into two classes, that is, benign tumors and malignant tumors. Cancer is a general term for malignant tumors. Tumors according to the present application can be selected from the group of CD47-positive hematological tumors and/or CD47-positive solid tumors.

[00110] In this application, the term "CD47-positive hematological tumor" generally refers to hematological tumors overexpressing CD47, which may include a number of leukemias, lymphomas and myelomas. Said "leukemia" basically refers to a malignant tumor of the blood in which too many white blood cells are produced, which are ineffective in fighting infection and crowd out other parts of the blood, such as platelets and red blood cells. Leukemias can be divided into acute and chronic leukemias. Some forms of leukemia may be, for example, acute lymphocytic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), chronic myelogenous leukemia (CML), myeloproliferative disorder/tumor (MPDS), and myelodysplastic syndrome. Said "lymphoma" may refer to Hodgkin's lymphoma, indolent and aggressive non-Hodgkin's lymphoma, Burkitt's lymphoma, follicular lymphoma (small cell and large cell), etc. Said myeloma may refer to multiple myeloma (MM), giant cell myeloma, heavy chain myeloma, light chain myeloma, or Bence-Jones myeloma.

[00111] As used herein, the term "CD47-positive solid tumor" generally refers to a solid tumor or visible tumor that overexpresses CD47 as can be detected by clinical examination such as X-ray, CT scan, B-ultrasound, or palpation. Major categories may include carcinomas and sarcomas. For example, these CD47-positive solid tumors may include Ewing's sarcoma, osteosarcoma, rhabdomyosarcoma, bladder cancer, ovarian cancer, prostate cancer, lung cancer, colon cancer, breast cancer, pancreatic cancer, astrocytic carcinoma, glioblastoma, renal cell carcinoma. cells, etc.

[00112] In this application, these autoimmune diseases may include Crohn's disease, allergic asthma and rheumatoid arthritis.

[00113] In this application, the term "Crohn's disease" generally refers to an inflammatory disease of the small intestine of unknown origin, which can occur in any part of the gastrointestinal tract. Said Crohn's disease and chronic ulcerative colitis are collectively referred to as inflammatory bowel disease (IBD).

[00114] In this application, the term "allergic asthma" generally refers to chronic inflammation of the airways, which involves a number of cells, in particular, mast cells, eosinophils and T-lymphocytes.

[00115] As used herein, the term "rheumatoid arthritis" generally refers to a chronic systemic autoimmune disease dominated by joint involvement.

[00116] In another aspect, the fusion protein, nucleic acid molecule, vector, host cell and/or composition of the present application can be used to prevent or treat said tumors or said autoimmune diseases.

[00117] In another aspect, the present application relates to a method for the prevention or treatment of tumors or autoimmune diseases, including the introduction of a fusion protein, a nucleic acid molecule, a vector, a host cell and / or a composition according to the present application to an individual.

[00118] In another aspect, the present application relates to a method of blocking the interaction of a CD47 protein and SIRPα, comprising the introduction of a fusion protein or composition according to the present application (for example, the introduction of an individual or a cell or biological sample that needs it).

[00119] In another aspect, the present application relates to a method of inhibiting the growth and/or proliferation of tumors or tumor cells, comprising contacting the fusion protein or composition of the present application with tumors or tumor cells. For example, said contact may occur in vitro .

[00120] In this application, the term "individual" generally refers to any human or non-human animal. The term "non-human animal" can include all vertebrates, such as mammals and non-mammals, for example, non-human primates, goats, sheep, dogs, cows, chickens, amphibians, reptiles, etc.

[00121] In this application, the term "approximately" generally refers to deviations within 0.5-10% of the specified value, for example, within 0.5%, 1%, 1.5%, 2%, 2.5 %, 3%, 3.5%, 4%, 4.5%, 5%, 5.5%, 6%, 6.5%, 7%, 7.5%, 8%, 8.5%, 9%, 9.5% or 10% of the indicated value.

[00122] As used herein, the term "comprising" generally means "including", "consisting of", "having", or "encompassing". In some cases, it also refers to the meaning "being" or "consisting of".

[00123] Without wishing to be bound by theory, the following examples are provided for the purpose of illustrating the modes of operation of the apparatus, method, and system of the present application, and not to limit the scope of the invention as claimed herein.

Examples

Example 1: Variant Screening

[00124] A truncated human SIRPα domain, variant 1 (NP_542970.1) with the amino acid sequence set forth in SEQ ID NO: 63 (i.e. residues at positions 33-149 in SEQ ID NO: 62) was taken using the Discovery Studio software (Neotrident) to construct a structure in a truncated domain that interacts with human CD47 (CEJ95640.1). The sites of interaction in the two proteins and their modes of interaction were theoretically analyzed to determine that the amino acid sites in the truncated domains that were directly or indirectly involved in interaction with CD47 were I61, V63, E77, Q82, K83, E84, V93, D95, D96, K98, N100, R107, G109, and V132 (in which the positions of the amino acid residues in the amino acid substitutions were numbered using the amino acid sequence set forth in SEQ ID NO: 62 as a reference). These sites of action were randomly mutated and a mutant library was constructed. The mutant library was then cloned into the pTM vector. Said pTM vector contained signal peptide and transmembrane region sequences (as shown in FIG. 1) which could display the gene cloned in the vector on the cell surface.

[00125] The mutant library engineered expression vector was transfected into CHO cells (ATCC) to display and express the mutant library on the cell surface. Then, CD47 protein (Yiqiao Shenzhou) was fluorescently labeled with FITC to obtain CD47-FITC. Consistent with the difference in binding activity between CD47-FITC and CHO cell surface truncated domain mutants, CD47-FITC binding mutants were enriched and examined by flow cytometry. The specific screening principle can be seen in FIG. 2 where the truncated domain and the mutant are bound to the CD47 protein with a fluorescent molecule, so that the binding results can be reflected by the level of the fluorescent molecule.

[00126] After four rounds of screening and enrichment, cells were harvested that strongly bound to CD47-FITC (as shown in FIG. 3). The mRNA was then isolated and reverse transcription performed to obtain cDNA, and the mutant truncated domain was sequenced (as shown in FIG. 4). The sequencing results showed that the indicated positions I61, V63, E77, Q82, K83, E84, V93, D95, D96, K98, N100, R107, G109, V132 have different combinations of mutation sites.

[00127] It can be seen from the results that a human SIRPα truncated domain mutant, variant 1, which can specifically recognize CD47 can be obtained by introducing various combinations of mutation sites at residues I61, V63, E77, Q82, K83, E84, V93, D95, D96 . , K98, N100, R107, G109 and/or V132.

[00128] Further analysis gave possible types of mutated amino acids at the indicated mutation sites: I61L/V/F, V63I, E77I/N/Q/K/H/M/R/N/V/L, Q82S/R/G/N , K83R, E84K/H/D/R/G, V93L/A, D95H/R/E, D96S/T, K98R, N100G/K/D/E, R107N/S, G109R/H, V132 L/R/ I/S.

[00129] Mutants containing these mutations were named M1, M5, M12, M35, M37, M41, M57, M67, M81, M82, M84, M91, M99, M102, M111, M122, M126, M130, M135 and M145 , and contained the following amino acid sequences listed sequentially in one of SEQ ID NOs: 1-20, respectively. At the same time, these mutants included in sequence the following combinations of amino acid mutations based on the amino acid sequence set forth in SEQ ID NO: 63:

(1) I61L, V63I, E77I, E84K, V93L, L96S, K98R, N100G and V132 L;

(2) I61V, E77N, Q82S, K83R and E84H;

(3) I61F, V63I, K83R, E84K and V132I;

(4) I61L, E77Q, E84D, R107N and V132I;

(5) I61L, V63I, E77K, K83R, E84D and N100G;

(6) I61V, E77H, Q82R, K83R, E84H and R107S;

(7) I61L, E77I, Q82G, E84R, V93L, L96T, N100G, R107S, G109R and V132R;

(8) I61L, E77M, Q82G, K83R, E84D and V132L;

(9) I61L;

(10) I61F, D95H, L96S, G109H and V132S;

(11) I61F, D95H, L96S, K98R, G109H and V132S;

(12) I61L, E77Q, E84D, V93A, R107N and V132I;

(13) E77K, L96S, N100K, G109H and V132L;

(14) I61L, V63I, Q82G, E84G, D95R, L96S, N100D and V132I;

(15) I61L, E77R, Q82N, K83R, E84G, V93L, D95E, L96T, K98R, N100D and V132L;

(16) I61V, E77N, Q82S, K83R, E84H and V93A;

(17) I61V, V63I, E77V, K83R, E84D, D95E, L96T, K98R and N100E;

(18) I61L, V63I, E77V, K83R, D95E, L96S, K98R, N100D and G109R;

(19) I61V, E77L, Q82G, E84G, V93L, D95E, L96T, K98R and N100G; and (20) I61L, V63I, E77N, Q82G and E84G.

Example 2 Measurement of Fusion Protein Binding Activity

[00130] The truncated human SIRPα domain variant 1 (or the truncated so-called wild-type SIRPα domain) in Example 1 and the human SIRPα domain mutant obtained in Example 1 (i.e. M1, M5, M12, M35, M37, M41, M57, M67, M81, M82, M84, M91, M99, M102, M111, M122, M126, M130, M135 and M145) were fused and expressed with human IgG1-Fc (the amino acid sequence of which is set forth in SEQ ID NO: 67), respectively to obtain the corresponding fusion protein from the SIRPα mutant 1, consisting of a truncated human Fc domain (shortly, fusion protein). Эти слитые белки были названы SS002, SS002M1, SS002M5, SS002M12, SS002M35, SS002M37, SS002M41, SS002M57, SS002M67, SS002M81, SS002M82, SS002M84, SS002M91, SS002M99, SS002M102, SS002M111, SS002M122, SS002M126, SS002M130, SS002M135 и SS002M145, соответственно. These fusion proteins consisted of the amino acid sequence shown in one of SEQ ID NOs: 61 and 21-40, respectively.

Table 1. SIRPα mutant 1 truncated domain and mutant and corresponding fusion protein SIRPα mutant 1 truncated domain or mutant thereof Fusion protein Shortened wild-type SIRPα domain SS002 M1 SS002M1 M5 SS002M5 M12 SS002M12 M35 SS002M35 M37 SS002M37 M41 SS002M41 M57 SS002M57 M67 SS002M67 M81 SS002M81 M82 SS002M82 M84 SS002M84 M91 SS002M91 M99 SS002M99 M102 SS002M102 M111 SS002M111 M122 SS002M122 M126 SS002M126 M130 SS002M130 M135 SS002M135 M145 SS002M145

[00131] The fusion proteins SS002, SS002M12, SS002M5, SS002M82, SS002M84, SS002M91, SS002M102 and SS002M130 were selected as an example for biological activity analysis.

[00132] The affinity of various fusion proteins including SS002, SS002M5, SS002M12, SS002M82, SS002M84, SS002M91, SS002M102, SS002M130 and the like for CD47 was determined using an ELISA method.

[00133] The ELISA plate was coated with 1 g/ml of target CD47-His antigen at 4°C overnight. After washing with PBST, 10% fetal calf serum was added and the mixture was blocked at 37° C. for 1 hour. Then, SS002, SS002M5, SS002M82, SS002M84, SS002M91, SS002M102, and SS002M130 were added, respectively, and reacted at 37°C for 1 hour. The reaction mixture was then washed with PBST and horseradish peroxidase labeled goat anti-human IgG (Thermo Fisher Scientific) was added and reacted at room temperature for 30 minutes. The plate was then re-washed 5 times with PBST and dried to remove residual liquid, if possible with blotting paper. Then 100 ml of TMB (eBioscience) was added to each well and kept at a temperature (20±5°C) in the dark for 1-5 minutes. 100 ml 2N H2SO4 was added to each well to quench the substrate reaction. The OD value was read at 450 nm with a microplate reader, and the affinity between each fusion protein and the CD47 molecule was analyzed (as shown in FIG. 5).

[00134] Results in FIG. 5 showed that fusion proteins including SS002, SS002M5, SS002M12, SS002M82, SS002M84, SS002M91, SS002M102, SS002M130 and the like. can effectively recognize the CD47 molecule.

Example 3: Affinity Analysis

[00135] As an example, the Biacore method was used to measure the affinity of each fusion protein, including SS002, SS002M5, SS002M12, SS002M82, SS002M84, SS002M91, SS002M102, SS002M130, and the like. to the CD47 molecule and the results are shown in Table 2 below.

Table 2 Binding affinity for CD47 Fusion protein Ka (10 ⁶ 1/Ms) Kd (10 ^-4 1/s) KD( ^10-9M ) SS002 1.32±0.09 63.00±1.35 4.80±0.20 SS002M5 1.72±0.12 86.20±9.65 5.00±0.22 SS002M12 1.87±0.05 86.10±2.94 4.62±0.08 SS002M82 1.29±0.05 21.13±0.42 1.65±0.03 SS002M84 1.10±0.02 14.30±0.20 1.30±0.02 SS002M91 1.56±0.13 2.72±0.44 0.18±0.04 SS002M102 1.79±0.20 28.97±7.83 1.60±0.24 SS002M130 1.43±0.09 88.20±8.67 6.20±0.99

[00136] The results in Table 2 show that SS002, SS002M5, SS002M12, SS002M82, SS002M84, SS002M91, SS002M102, SS002M130 and other fusion proteins can recognize CD47 molecules with high affinity.

Example 4 Species Recognition Specificity for a Fusion Protein

[00137] The fusion proteins SS002 and SS002M91 were taken as an example for analysis of specific recognition activity.

[00138] For species analysis of fusion proteins, 1 mg/ml human CD47 and mouse CD47 ELISA plates (Beijing Yiqiao Shenzhou Biotechnology Co., Ltd.), respectively, were coated and kept at 4°C overnight. After washing with PBST, 10% fetal calf serum was added and the mixture was blocked at 37° C. for 1 hour. Then SS002 and SS002M91 were added, respectively, and the reaction was carried out at 37°C for 1 hour. After washing with PBST, horseradish peroxidase labeled goat anti-human IgG (Thermo Fisher Scientific) was added and reacted at room temperature for 30 minutes. The plate was then re-washed 5 times with PBST and dried to remove residual liquid droplets, if possible with blotting paper. Then 100 ml of TMB (eBioscience) was added to each well and kept at room temperature (20±5°C) in the dark for 1-5 minutes. 100 ml 2N H ₂ SO ₄ was added to each well to quench the substrate reaction. The OD value was read at 450 nm with a microplate reader, and the binding ability of the fusion protein to various CD47 species was analyzed (experimental results for the fusion protein SS002 and SS002M91 are shown in FIGS. 6A and 6B, respectively).

[00139] The results in FIG. 6A-6B show that both SS002 and SS002M91 can specifically recognize human CD47 molecules, but cannot recognize mouse CD47 molecules.

Example 5 Fusion Proteins Specifically Recognize Target Antigen

[00140] Fusion proteins SS002 and SS002M91 were taken as an example. Plates were coated for ELISA 1 mg/ml SS002, SS002M91, as well as milk (Beijing Bomed Biotechnology Co., Ltd.), BSA (BOVOGEN), CD19 (Beijing Yiqiao Shenzhou Biotechnology Co., Ltd.), TROP2 (Beijing Yiqiao Shenzhou Biotechnology Co., Ltd.), CD47 (Beijing Magppel Biotech Co., Ltd.), CD38 (Beijing Yiqiao Shenzhou Biotechnology Co., Ltd.), Gas6 (R&D) and other proteins, and AXL (ACRO Biosystems), respectively, and left at 4°C overnight. After washing with PBST, 10% fetal calf serum was added and the mixture was blocked at 37° C. for 1 hour. Then SS002 and SS002M91 were added, respectively, and the reaction was carried out at 37°C for 1 hour. After washing with PBST, horseradish peroxidase labeled goat anti-human IgG (Thermo Fisher Scientific) was added and reacted at room temperature for 30 minutes. The plate was then re-washed 5 times with PBST and dried to remove residual liquid droplets, if possible with blotting paper. Then 100 ml of TMB (eBioscience) was added to each well and kept at room temperature (20±5°C) in the dark for 1-5 minutes. 100 ml 2N H ₂ SO ₄ was added to each well to quench the substrate reaction. The OD value was read at 450 nm with a microplate reader, and the ability of the fusion protein to bind to various of the above proteins was analyzed (as shown in FIG. 7).

[00141] Results in FIG. 7 showed that the fusion proteins SS002 and SS002M91 readily recognize the CD47 molecule and do not cross-react with various other proteins.

Example 6 Fusion Proteins Specifically Block CD47/SIRP α Interaction

[00142] SS002M91 was used as an example for specific blocking activity of the CD47/SIRPα interaction, and the expression of the American analog TTI-621 (see CN105073780A) was used as a positive control.

[00143] ELISA plates were coated with 1 mg/ml SIRPα-His and left at 4°C overnight. After washing with PBST, 10% fetal calf serum was added and the mixture was blocked at 37° C. for 1 hour. SS002M91 and TTI-621 were serially diluted with 10% fetal calf blood, respectively, and Biotin-Fc-CD47 was added to the samples to a final concentration of 2 μg/ml. The mixture was pre-incubated at 37°C for 30 minutes for use as a primary antibody. After the ELISA plate was washed with PBST, the primary antibody was added and incubated at 37° C. for 1 hour. Then, after washing 5 times with PBST, horseradish peroxidase labeled avidin (Streptavidin-HRP, Jiaxuan Bio) was added and incubated at 37° C. for 30 minutes. After washing 5 times with PBST, 100 μl of TMB (eBioscience) was added to each well and left at room temperature (20±5°C) in the dark for 1-5 minutes. 100 ml 2N H ₂ SO ₄ was added to each well to quench the substrate reaction. The OD value was read at 450 nm with a microplate reader, and the blocking effect of the SIRPα fusion protein on CD47/SIRPα was analyzed (as shown in FIG. 8).

[00144] Results in FIG. 8 showed that both SS002M91 and TTI-621 can competitively block CD47 binding to its SIRPα ligand. However, the SS002M91 fusion protein has a significantly higher blocking activity than TTI-621. The IC50 value for SS002M91 is 5.47µg/ml and the IC50 value for TTI-621 is 493.5µg/ml.

Example 7 Fusion Protein Specifically Recognizes the CD47 Molecule on the Surface of Tumor Cells

[00145] Fusion proteins SS002 and SS002M91 were taken as examples for assaying the recognition activity of the CD47 molecule on the surface of tumor cells.

[00146] Flow cytometry (BD Calibur) was used to appropriately detect the activity of SS002 and SS002M91, which specifically recognize CD47 molecules on the surface of Raji cells, Jurkat cells, and A549 cells. The above cells were collected in the logarithmic growth phase, respectively, with the cell density adjusted to 5×10 ⁶ cells/ml, and pre-cooled on ice for 10 minutes. The SIRPα SS002 and SS002M91 fusion proteins were diluted to various concentrations with pre-chilled saline containing 2% FBS. 100 μl of cells were added to an equal volume of the above diluted SIRPα fusion protein and reacted at 4° C. in the dark for 30 minutes. Upon completion, the cells were washed twice with pre-chilled saline containing 2% FBS. Cells were resuspended in 100 μl of diluted PE-labeled goat anti-human IgG-Fc secondary antibody (eBioscience) and reacted at 4° C. in the dark for 30 minutes. After completion of the reaction, the cells were washed twice with pre-chilled saline containing 2% FBS and resuspended in 400 μl of 1% paraformaldehyde. Flow cytometry (BD Calibur) was used to analyze the binding capacity of the fusion protein to CD47 on the cell surface (as shown in Figures 9A-9C, where SS002 and SS002M91 fusion proteins specifically recognize surface CD47 on Raji cells, Jurkat cells, and A549 cells, respectively) .

[00147] The results showed that the SS002M91 fusion protein could specifically recognize CD47 on the surface of Raji cells, Jurkat cells and A549 cells with a recognition activity that was significantly higher than that of SS002 and was dose dependent. Among them, the binding value of EC ₅₀ to Raji cells was 197.0 ng/ml for SS002M91 and 1140.0 ng/ml for SS002 (as shown in FIG. 9A). The binding value of EC ₅₀ to Jurkat cells for SS002M91 was 796.0 ng/ml and for SS002 was 4529.0 ng/ml (as shown in FIG. 9B). The binding value of EC ₅₀ to A549 cells for SS002M91 was 321.9 ng/ml and for SS002 was 1655.0 ng/ml (as shown in FIG. 9C).

[00148] Similarly, the method of this example was used to compare the CD47 recognition activity of SS002M91 and TTI-621 on the surfaces of Raji cells (as shown in FIG. 10A), Jurkat cells (as shown in FIG. 10B), and A549 cells (as shown in Fig. 10C). The results showed that in one aspect, the maximum fluorescence intensity for SS002M91, which specifically recognizes CD47 molecules on the surface of tumor cells, was significantly higher than for TTI-621: the maximum fluorescence intensity of SS002M91 binding to Raji cells was approximately 1200, while how the maximum fluorescence intensity of TTI-621 binding to Raji cells was approximately 600; the maximum fluorescence intensity of SS002M91 binding to Jurkat cells was approximately 5000, while the maximum fluorescence intensity of TTI-621 binding to Jurkat cells was approximately 3000; the maximum fluorescence intensity of SS002M91 binding to A549 cells was approximately 180, while the maximum fluorescence intensity of TTI-621 binding to A549 cells was approximately 60. In another aspect, the semi-optimal dose of SS002M91, which specifically recognizes CD47 molecules on the surface of tumor cells, was significantly better than TTI-621: the EC ₅₀ value for SS002M91 binding to Raji cells was 13.06 ng/ml, while the EC 50 value for TTI-621 binding to Raji cells was 40.37 ng/ml; the EC ₅₀ value for SS002M91 binding to Jurkat cells was 28.09 ng/ml, while the EC ₅₀ value for TTI-621 binding to Jurkat cells was 53.92 ng/ml; the EC ₅₀ value for SS002M91 binding to A549 cells was 26.95 ng/ml and the EC ₅₀ value for TTI-621 binding to A549 cells was 1003 ng/ml. It can be seen that SS002M91, which specifically recognizes CD47 molecules on the surface of tumor cells, is significantly better than TT1-621.

Example 8: Detection of blood coagulation reaction

[00149] SS002 and SS002M91 were taken as examples for a clotting activity assay using the anti-CD47 antibody Hu5F9-G4 as a control (see Guerriero JL, Sotayo A, Ponichtera HE, et al. Class IIa HDAC inhibition reduces breast tumors and metastases through anti-tumour macrophages [J] Nature, 2017, 543(7645): 428.432 and Gholamin S, Mitra SS, Feroze AH et al Disrupting the CD47-SIRPα antiphagocytic axis by a humanized anti-CD47 antibody is an efficacious treatment for malignant pediatric brain tumors Sci Transl Med 2017).

[00150] Whole blood from healthy donors was used to obtain human erythrocytes (collected from the peripheral blood of volunteers). Whole blood was diluted 5-fold with PBS, washed 3 times, and a fresh 1% erythrocyte solution was prepared. 50 µl of various concentrations of SIRPα SS002, SS002M91 fusion proteins and anti-CD47 Hu5F9-G4 antibody were added to each well of the hemagglutination plate, 50 µl of 1% erythrocyte solution was added to each well. The mixture was gently stirred until homogeneous and incubated at 37°C in an atmosphere of 5% CO ₂ during the night. The plate was then photographed for interpretation using standards that all erythrocytes coagulated, sank to the bottom of the well, and flattened into a mesh shape at 100% coagulation (++++), and the phenomenon that erythrocytes sank to the bottom and looked like dots taken as no coagulation (-) (as shown in FIG. 11).

[00151] Results in FIG. 11 showed that the fusion proteins SS002M91 and SS002 do not cause erythrocyte agglutination, while the CD47 Hu5F9-G4 antibody can significantly cause erythrocyte agglutination in a certain dose range.

Example 9 Detection of Tumor Inhibiting Activity in vivo

[00152] As an example for assaying tumor inhibitory activityin vivoused fusion protein SS002M91.

[00153] B-NSG mice were inoculated with Raji-Luc cells to create a tumor model to evaluate the tumor-inhibiting activity of the SS002M91 antibody. 8-week-old female B-NSG mice (Beijing Biocytogene Biotechnology Co., Ltd.) were taken as experimental animals, and Raji-Luc cells (Beijing Biocytogene Biotechnology Co., Ltd.) were selected for testing. Raji-Luc cells were transferred to a stable cell line derived from a fluorescein reporter gene. After resuscitation and cultivation to the required number, the cells were harvested in the log phase of growth and suspended to 5×10 ⁶ cells/0.2 ml. B-NSG mice were then inoculated with cells via the tail vein at a concentration of 0.2 ml/mouse. After vaccination, tumor growth and body weight were observed on Day 0 and Day 3 using a small animal imager. And on Day 3, 12 mice with a moderate tumor imaging signal (approximately 1.00×10 ⁶ P/S) were selected and randomly divided into 2 groups (6 mice per group), i.e., a vehicle control group (G1, saline ) and the experimental group (G2, SS002M91). The experimental group was dosed at 10 mg/kg on Days 0 and D3 after group assignment, twice in total. Tumor growth and survival rates of mice were observed (as shown in FIG. 12A and 12B, respectively).

[00154] The results showed that on Day 10 after grouping, the mean fluorescence intensity of the tumors in the control group was 6.75×10 ⁸ P/S, while the mean fluorescence intensity of the tumors in the dosing group was 1.76×10 ⁶ P/S, and the degree of inhibition is approximately 95%.

Example 10: Detection of effects on erythrocytes and platelets

[00155] The SS002M91 fusion protein was used as an example to conduct a preliminary in vivo safety assessment, and B-NSG mice were used as a model.

[00156] Eighteen female 8-week-old B-NSG mice (Beijing Biocytogene Biotechnology Co., Ltd.) were selected and randomly divided into 3 groups, i.e., vehicle control group (saline injected), experimental group (fused SS002M91 protein) and a positive control group (treated with TTI-621) (6 mice per group). Mice were dosed at 10 mg/kg on Day 0, Day 3 and Day 7 after group assignment, a total of 3 times. On the following day after the third injection (ie, Day 8), the levels of red blood cells and platelets were analyzed in the peripheral blood of mice (the level of red blood cells and the level of platelets are shown in FIG. 13A and FIG. 13B, respectively).

[00157] The results showed that compared to the control group, SS002M91 does not cause a significant decrease in red blood cells (P=0.4483) and platelets (P=0.9199); while TTI-621 had little effect on platelets (P=0.9447) but caused a decrease in red blood cells (P=0.0246).

Example 11: Effect of fusion with different IgG Fc subtypes on fusion protein activity

[00158] According to the method of constructing the fusion protein in Example 2, the mutants M1, M5, M12, M35, M37, M41, M57, M67, M81, M82, M84, M91, M99, M102, M111, M122, M126, M130 , M135 and M145 of the SIRPα domain obtained in Example 1 were fused and expressed with human IgG4-Fc (amino acid sequences of which were indicated in SEQ ID NO: 68), respectively, to obtain the corresponding fusion protein: truncated SIRPα domains of mutant 1-human Fc (referred to as a fusion protein). Эти слитые белки были названы SS002M1G4, SS002M5G4, SS002M12G4, SS002M35G4, SS002M37G4, SS002M41G4, SS002M57G4, SS002M67G4, SS002M81G4, SS002M82G4, SS002M84G4, SS002M91G4, SS002M99G4, SS002M102G4, SS002M111G4, SS002M122G4, SS002M126G4, SS002M130G4, SS002M135G4 и SS002M145G4 (аминокислотные последовательности которых были изложены in SEQ ID NO: 41-60, respectively).

[00159] The fusion protein SS002M91G4 was chosen as an example for the biological activity assay.

[00160] The binding activity of SS002M91G4 to the CD47 antigen was analyzed according to the method for measuring binding activity in Example 2, and the results are shown in FIG. 14. Results in FIG. 14 showed that SS002M91G4 has good binding activity to the CD47 antigen with an EC ₅₀ value of 0.0157 μg/ml, which is substantially consistent with the EC ₅₀ (0.0195 μg/ml) for SS002M91.

[00161] In accordance with the method of analysis of specific blocking of the CD47/SIRPα interaction in Example 6, the activity of SS002M91G4 to block the CD47 SIRPα interaction was analyzed, and the results are shown in FIG. 15. Results in FIG. 15 showed that SS002M91G4 has good activity in blocking the CD47/SIRPα interaction with an IC ₅₀ value of 3.46 μg/ml, which is substantially consistent with the IC 50 value of 5.47 μg/ml for SS002M91.

[00162] It can be seen from the above results that fusion with various IgG Fc subtypes does not significantly affect the activity of the fusion protein constructed according to the present application.

Example 12 Detection of blood clotting reaction for a fusion protein

[00163] The blood clotting response of the IgG4 Fc fusion protein was evaluated using SS002M91G4 as an example and according to the method for detecting blood clotting response in Example 8. Whole blood from healthy donors was used to obtain human erythrocytes. Whole blood was diluted 5-fold with PBS, washed 3 times, and a fresh 1% erythrocyte solution was prepared. 50 µl of various concentrations of SS002M91G4 fusion proteins, TTI-621 positive control, and anti-CD47 antibody Hu5F9-G4 were added to each well of the hemagglutination plate, and 50 µl of 1% erythrocyte solution was added to each well. The mixture was gently stirred until homogeneous and incubated at 37°C in an atmosphere of 5% CO ₂ during the night. The plate was then photographed for interpretation using standards that all erythrocytes coagulated, sank to the bottom of the well, and flattened into a mesh shape at 100% coagulation (++++), and the phenomenon that erythrocytes sank to the bottom and looked like dots taken as no coagulation (-).

[00164] The results are shown in FIG. 16. The results showed that the IgG4 Fc fusion proteins SS002M91G4 and TTI-621 did not cause erythrocyte agglutination, while the CD47 Hu5F9-G4 antibody significantly caused erythrocyte agglutination over a certain dose range.

Example 13 Fusion Protein Biological Activity Assay

[00165] To evaluate the biological activity of this IgG4 Fc fusion protein, SS002M91G4 was taken as an example, Jurkat-CSR cell system (Immune OncoBiomedical Technology (Shanghai) Co., Ltd.) was used, and TTI-621 was used as a positive control.

[00166] CD47-Fc protein (Catalog number 12283-H02H, Sino Biological) was diluted to 0.2 μg/ml, and 50 μl of diluent was added to each well. Then TTI-621 and SS002M91G4 were diluted to 0.4 mg/ml, respectively, and then sequentially diluted to different concentrations. Added 50 μl to each well and incubated with CD47-Fc in an incubator at 37°C, 5% CO ₂ for 45 minutes. Jurkat-CSR cells were taken and the density adjusted to 5×10 ⁵ /ml, 100 μl of cell suspension was added to each well, and a blank control group was established. The suspension was co-cultured with the protein mixture in an incubator at 37°C, 5% CO ₂ for 20 hours. After completion of cultivation, 20 μl of CCK-8 (Dojindo, Japan Institute of Chemistry) was added to each well and cultured in a 37°C, 5% CO ₂ incubator for 4 hours. The OD value was measured at a wavelength of 450 nm with a microspectrophotometer for reading plates. The cell growth inhibition rate was calculated according to the following formula: Percent inhibition = (OD450 (sample)-OD450 (blank)/(OD450 (Jurkat-CSR)-OD450 (blank))×100.

[00167] According to the principle of the Jurkat-CSR cell system, the CD47/SIRPα interaction can induce target cell apoptosis, and the addition of an inhibitor that inhibits the CD47/SIRPα interaction can block the apoptosis signal. The stronger the action of the inhibitor, the more completely the apoptotic signal is blocked. The result is shown in FIG. 17. The results showed that SS002M91G4 can significantly block CD47-Fc-induced apoptosis of Jurkat-CSR cells, the blocking effect is stronger than that of TTI-621, and the IC ₅₀ is about 1/100 of TTI-621. It can be seen that the biological activity of SS002M91G4 in blocking the CD47/SIRPα interaction is significantly better than that of TTI-621.

Example 14: Tumor Inhibiting Activity in vivo Fc of a fusion protein from various IgG subtypes and effects on erythrocytes and platelets

[00168] Fusion proteins SS002M91 and SS002M91G4 were taken as an example and tumor activity and effects on erythrocytes and platelets were analyzed in vivo .

[00169] NOD/SCID mice were subcutaneously inoculated with Raji cells and a transplanted subcutaneous tumor model from human lymphoma was created to evaluate tumor inhibition activity of SS002M91 and SS002M91G4 in vivo .

[00170] 6-7 week old female NOD/SCID mice (Shanghai Lingchang Biotechnology Co., Ltd.) were selected and Raji cells were cultured in RPMI1640 medium containing 10% fetal bovine serum. 1×10 ⁷ Raji cells in the exponential growth phase were harvested and resuspended to the appropriate concentration in PBS and then mixed with Matrigel (BD Matrigel™) at a 1:1 ratio for subcutaneous tumor inoculation in mice. After inoculation, when the average tumor volume was approximately 98.6 mm ³ , mice were randomly divided into 4 groups (6 mice per group) according to tumor size (i.e., vehicle control group, SS002M91 group, SS002M91G4 group, and TTI-621). Mice were intraperitoneally injected at a dose of 10 mg/kg at a time (where the above four groups were injected with a solution of PBS, SS002M91, SS002M91G4 and TTI-621, respectively) twice a week for two weeks.

[00171] The experiment ended on day 6 after the last injection and blood was taken for routine testing. The levels of erythrocytes and platelets in the peripheral blood of mice were analyzed (the level of erythrocytes and the level of platelets are shown in FIG. 18A and FIG. 18B, respectively). Tumor growth was monitored during administration. The therapeutic effect was evaluated according to relative tumor growth inhibition (TGI) (results shown in FIG. 19), and safety was evaluated based on animal weight change and mortality (results shown in FIG. 20). The formula for calculating TGI (%), that is, the relative inhibition of tumor growth, was as follows: TGI% = (1-T/C)×100%. Of these, T and C are the relative tumor volume (RTV) or tumor weight (TW) of the treatment groups (e.g., SS002M91 group, SS002M91G4 group, and TTI-621 group) and control group (i.e., vehicle control group) at a certain moment of time.

[00172] The results showed that the SS002M91 group, the SS002M91G4 group, and the TTI-621 (10 mg/kg) group showed a significant tumor inhibition effect on Day 6 after drug withdrawal and showed a relative tumor growth inhibition TGI (%), which was 74, 09%, 66.65% and 54.75%, respectively. Compared to the vehicle control group, there are statistically significant differences (all p values less than 0.01). The SS002M91 group and the SS002M91G4 group have a similar tumor inhibitory effect, better than the TTI-621 positive control group.

[00173] During treatment, no animals died or showed obvious drug toxicity, and they all tolerated the drug well. The results of the routine blood test showed that, compared with the TTI-621 positive control group, the erythrocytes and platelets of the SS002M91 group were reduced, the platelets of the TTI-621 group also slightly decreased, but the erythrocytes were essentially not affected by SS002M91G4.

[00174] The above detailed description has been provided by way of illustration and example, and is not intended to limit the scope of the appended claims. The various embodiments listed herein are now apparent to those skilled in the art and remain within the scope of the appended claims and their equivalents.

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SEQUENCE LIST

<110> HANGZHOU SUMGEN BIOTECH CO., LTD.; SUMGENMAB (BEIJING)

BIOTECH CO., LTD.

<120> FUNCTION PROTEIN BINDING TO CD47 PROTEIN AND ITS USE

<130> 0070-PA-009

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<170>PatentIn version 3.5

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Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Phe Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Glu Leu Ile Tyr

35 40 45

Asn Gln Lys Glu Gly His Phe Pro Arg Val Thr Thr Val Ser His Ser

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile His Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Ser Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser

115

<210> 11

<211> 117

<212> Protein

<213> Artificial sequence

<220>

<223> M84

<400> 11

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Phe Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Glu Leu Ile Tyr

35 40 45

Asn Gln Lys Glu Gly His Phe Pro Arg Val Thr Thr Val Ser His Ser

50 55 60

Thr Arg Arg Asn Asn Met Asp Phe Ser Ile Arg Ile His Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Ser Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser

115

<210> 12

<211> 117

<212> Protein

<213> Artificial sequence

<220>

<223> M91

<400> 12

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Gln Leu Ile Tyr

35 40 45

Asn Gln Lys Asp Gly His Phe Pro Arg Val Thr Thr Ala Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Asn Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Ile Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser

115

<210> 13

<211> 117

<212> Protein

<213> Artificial sequence

<220>

<223> M99

<400> 13

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Ile Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Lys Leu Ile Tyr

35 40 45

Asn Gln Lys Glu Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Ser

50 55 60

Thr Lys Arg Lys Asn Met Asp Phe Ser Ile Arg Ile His Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Leu Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser

115

<210> 14

<211> 117

<212> Protein

<213> Artificial sequence

<220>

<223> M102

<400> 14

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Ile Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Glu Leu Ile Tyr

35 40 45

Asn Gly Lys Gly Gly His Phe Pro Arg Val Thr Thr Val Ser Arg Ser

50 55 60

Thr Lys Arg Asp Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Ile Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser

115

<210> 15

<211> 117

<212> Protein

<213> Artificial sequence

<220>

<223> M111

<400> 15

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Arg Leu Ile Tyr

35 40 45

Asn Asn Arg Gly Gly His Phe Pro Arg Val Thr Thr Leu Ser Glu Thr

50 55 60

Thr Arg Arg Asp Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Leu Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser

115

<210> 16

<211> 117

<212> Protein

<213> Artificial sequence

<220>

<223> M122

<400> 16

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Val Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Asn Leu Ile Tyr

35 40 45

Asn Ser Arg His Gly His Phe Pro Arg Val Thr Thr Ala Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser

115

<210> 17

<211> 117

<212> Protein

<213> Artificial sequence

<220>

<223> M126

<400> 17

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Val Pro Ile Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Val Leu Ile Tyr

35 40 45

Asn Gln Arg Asp Gly His Phe Pro Arg Val Thr Thr Val Ser Glu Thr

50 55 60

Thr Arg Arg Glu Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser

115

<210> 18

<211> 117

<212> Protein

<213> Artificial sequence

<220>

<223> M130

<400> 18

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Ile Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Val Leu Ile Tyr

35 40 45

Asn Gln Arg Glu Gly His Phe Pro Arg Val Thr Thr Val Ser Glu Ser

50 55 60

Thr Arg Arg Asp Asn Met Asp Phe Ser Ile Arg Ile Arg Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser

115

<210> 19

<211> 117

<212> Protein

<213> Artificial sequence

<220>

<223> M135

<400> 19

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Val Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Leu Leu Ile Tyr

35 40 45

Asn Gly Lys Gly Gly His Phe Pro Arg Val Thr Thr Leu Ser Glu Thr

50 55 60

Thr Arg Arg Gly Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser

115

<210> 20

<211> 117

<212> Protein

<213> Artificial sequence

<220>

<223> M145

<400> 20

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Ile Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Asn Leu Ile Tyr

35 40 45

Asn Gly Lys Gly Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser

115

<210> 21

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M1

<400> 21

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Ile Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Ile Leu Ile Tyr

35 40 45

Asn Gln Lys Lys Gly His Phe Pro Arg Val Thr Thr Leu Ser Asp Ser

50 55 60

Thr Arg Arg Gly Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Leu Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 22

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M5

<400> 22

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Val Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Asn Leu Ile Tyr

35 40 45

Asn Ser Arg His Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 23

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M12

<400> 23

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Phe Pro Ile Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Glu Leu Ile Tyr

35 40 45

Asn Gln Arg Lys Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Ile Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 24

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M35

<400> 24

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Gln Leu Ile Tyr

35 40 45

Asn Gln Lys Asp Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Asn Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Ile Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 25

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M37

<400> 25

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Ile Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Lys Leu Ile Tyr

35 40 45

Asn Gln Arg Asp Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Gly Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 26

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M41

<400> 26

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Val Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg His Leu Ile Tyr

35 40 45

Asn Arg Arg His Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Ser Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 27

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M57

<400> 27

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Ile Leu Ile Tyr

35 40 45

Asn Gly Lys Arg Gly His Phe Pro Arg Val Thr Thr Leu Ser Asp Thr

50 55 60

Thr Lys Arg Gly Asn Met Asp Phe Ser Ile Ser Ile Arg Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Arg Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 28

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M67

<400> 28

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Met Leu Ile Tyr

35 40 45

Asn Gly Arg Asp Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Leu Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 29

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M81

<400> 29

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Glu Leu Ile Tyr

35 40 45

Asn Gln Lys Glu Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 30

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M82

<400> 30

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Phe Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Glu Leu Ile Tyr

35 40 45

Asn Gln Lys Glu Gly His Phe Pro Arg Val Thr Thr Val Ser His Ser

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile His Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Ser Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 31

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M84

<400> 31

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Phe Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Glu Leu Ile Tyr

35 40 45

Asn Gln Lys Glu Gly His Phe Pro Arg Val Thr Thr Val Ser His Ser

50 55 60

Thr Arg Arg Asn Asn Met Asp Phe Ser Ile Arg Ile His Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Ser Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 32

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M91

<400> 32

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Gln Leu Ile Tyr

35 40 45

Asn Gln Lys Asp Gly His Phe Pro Arg Val Thr Thr Ala Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Asn Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Ile Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 33

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M99

<400> 33

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Ile Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Lys Leu Ile Tyr

35 40 45

Asn Gln Lys Glu Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Ser

50 55 60

Thr Lys Arg Lys Asn Met Asp Phe Ser Ile Arg Ile His Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Leu Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 34

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M102

<400> 34

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Ile Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Glu Leu Ile Tyr

35 40 45

Asn Gly Lys Gly Gly His Phe Pro Arg Val Thr Thr Val Ser Arg Ser

50 55 60

Thr Lys Arg Asp Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Ile Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 35

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M111

<400> 35

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Arg Leu Ile Tyr

35 40 45

Asn Asn Arg Gly Gly His Phe Pro Arg Val Thr Thr Leu Ser Glu Thr

50 55 60

Thr Arg Arg Asp Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Leu Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 36

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M122

<400> 36

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Val Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Asn Leu Ile Tyr

35 40 45

Asn Ser Arg His Gly His Phe Pro Arg Val Thr Thr Ala Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 37

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M126

<400> 37

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Val Pro Ile Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Val Leu Ile Tyr

35 40 45

Asn Gln Arg Asp Gly His Phe Pro Arg Val Thr Thr Val Ser Glu Thr

50 55 60

Thr Arg Arg Glu Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 38

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M130

<400> 38

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Ile Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Val Leu Ile Tyr

35 40 45

Asn Gln Arg Glu Gly His Phe Pro Arg Val Thr Thr Val Ser Glu Ser

50 55 60

Thr Arg Arg Asp Asn Met Asp Phe Ser Ile Arg Ile Arg Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 39

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M135

<400> 39

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Val Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Leu Leu Ile Tyr

35 40 45

Asn Gly Lys Gly Gly His Phe Pro Arg Val Thr Thr Leu Ser Glu Thr

50 55 60

Thr Arg Arg Gly Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 40

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M145

<400> 40

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Ile Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Asn Leu Ile Tyr

35 40 45

Asn Gly Lys Gly Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 41

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M1G4

<400> 41

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Ile Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Ile Leu Ile Tyr

35 40 45

Asn Gln Lys Lys Gly His Phe Pro Arg Val Thr Thr Leu Ser Asp Ser

50 55 60

Thr Arg Arg Gly Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Leu Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 42

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M5G4

<400> 42

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Val Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Asn Leu Ile Tyr

35 40 45

Asn Ser Arg His Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 43

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M12G4

<400> 43

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Phe Pro Ile Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Glu Leu Ile Tyr

35 40 45

Asn Gln Arg Lys Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Ile Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 44

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M35G4

<400> 44

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Gln Leu Ile Tyr

35 40 45

Asn Gln Lys Asp Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Asn Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Ile Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 45

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M37G4

<400> 45

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Ile Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Lys Leu Ile Tyr

35 40 45

Asn Gln Arg Asp Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Gly Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 46

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M41G4

<400> 46

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Val Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg His Leu Ile Tyr

35 40 45

Asn Arg Arg His Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Ser Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 47

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M57G4

<400> 47

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Ile Leu Ile Tyr

35 40 45

Asn Gly Lys Arg Gly His Phe Pro Arg Val Thr Thr Leu Ser Asp Thr

50 55 60

Thr Lys Arg Gly Asn Met Asp Phe Ser Ile Ser Ile Arg Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Arg Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 48

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M67G4

<400> 48

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Met Leu Ile Tyr

35 40 45

Asn Gly Arg Asp Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Leu Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 49

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M81G4

<400> 49

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Glu Leu Ile Tyr

35 40 45

Asn Gln Lys Glu Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 50

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M82G4

<400> 50

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Phe Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Glu Leu Ile Tyr

35 40 45

Asn Gln Lys Glu Gly His Phe Pro Arg Val Thr Thr Val Ser His Ser

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile His Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Ser Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 51

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M84G4

<400> 51

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Phe Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Glu Leu Ile Tyr

35 40 45

Asn Gln Lys Glu Gly His Phe Pro Arg Val Thr Thr Val Ser His Ser

50 55 60

Thr Arg Arg Asn Asn Met Asp Phe Ser Ile Arg Ile His Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Ser Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 52

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M91G4

<400> 52

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Gln Leu Ile Tyr

35 40 45

Asn Gln Lys Asp Gly His Phe Pro Arg Val Thr Thr Ala Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Asn Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Ile Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 53

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M99G4

<400> 53

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Ile Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Lys Leu Ile Tyr

35 40 45

Asn Gln Lys Glu Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Ser

50 55 60

Thr Lys Arg Lys Asn Met Asp Phe Ser Ile Arg Ile His Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Leu Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 54

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M102G4

<400> 54

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Ile Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Glu Leu Ile Tyr

35 40 45

Asn Gly Lys Gly Gly His Phe Pro Arg Val Thr Thr Val Ser Arg Ser

50 55 60

Thr Lys Arg Asp Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Ile Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 55

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M111G4

<400> 55

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Val Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Asn Leu Ile Tyr

35 40 45

Asn Ser Arg His Gly His Phe Pro Arg Val Thr Thr Ala Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 56

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M122G4

<400> 56

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Val Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Asn Leu Ile Tyr

35 40 45

Asn Ser Arg His Gly His Phe Pro Arg Val Thr Thr Ala Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 57

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M126G4

<400> 57

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Val Pro Ile Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Val Leu Ile Tyr

35 40 45

Asn Gln Arg Asp Gly His Phe Pro Arg Val Thr Thr Val Ser Glu Thr

50 55 60

Thr Arg Arg Glu Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 58

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M130G4

<400> 58

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Ile Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Val Leu Ile Tyr

35 40 45

Asn Gln Arg Glu Gly His Phe Pro Arg Val Thr Thr Val Ser Glu Ser

50 55 60

Thr Arg Arg Asp Asn Met Asp Phe Ser Ile Arg Ile Arg Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 59

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M135G4

<400> 59

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Val Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Leu Leu Ile Tyr

35 40 45

Asn Gly Lys Gly Gly His Phe Pro Arg Val Thr Thr Leu Ser Glu Thr

50 55 60

Thr Arg Arg Gly Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 60

<211> 346

<212> Protein

<213> Artificial sequence

<220>

<223> SS002M145G4

<400> 60

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Leu Pro Ile Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Asn Leu Ile Tyr

35 40 45

Asn Gly Lys Gly Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys

115 120 125

Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro

130 135 140

Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys

145 150 155 160

Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp

165 170 175

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu

180 185 190

Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu

195 200 205

His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn

210 215 220

Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly

225 230 235 240

Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu

245 250 255

Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr

260 265 270

Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn

275 280 285

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe

290 295 300

Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn

305 310 315 320

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr

325 330 335

Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

340 345

<210> 61

<211> 350

<212> Protein

<213> Artificial sequence

<220>

<223>SS002

<400> 61

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Ile Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Glu Leu Ile Tyr

35 40 45

Asn Gln Lys Glu Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr

115 120 125

Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe

130 135 140

Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro

145 150 155 160

Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val

165 170 175

Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr

180 185 190

Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val

195 200 205

Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys

210 215 220

Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser

225 230 235 240

Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro

245 250 255

Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val

260 265 270

Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly

275 280 285

Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp

290 295 300

Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp

305 310 315 320

Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His

325 330 335

Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys

340 345 350

<210> 62

<211> 504

<212> Protein

<213> Artificial sequence

<220>

<223> Human SIRP ¦B option 1

<400> 62

Met Glu Pro Ala Gly Pro Ala Pro Gly Arg Leu Gly Pro Leu Leu Cys

1 5 10 15

Leu Leu Leu Ala Ala Ser Cys Ala Trp Ser Gly Val Ala Gly Glu Glu

20 25 30

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

35 40 45

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Ile Pro Val Gly

50 55 60

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Glu Leu Ile Tyr

65 70 75 80

Asn Gln Lys Glu Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

85 90 95

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

100 105 110

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

115 120 125

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

130 135 140

Arg Ala Lys Pro Ser Ala Pro Val Val Ser Gly Pro Ala Ala Arg Ala

145 150 155 160

Thr Pro Gln His Thr Val Ser Phe Thr Cys Glu Ser His Gly Phe Ser

165 170 175

Pro Arg Asp Ile Thr Leu Lys Trp Phe Lys Asn Gly Asn Glu Leu Ser

180 185 190

Asp Phe Gln Thr Asn Val Asp Pro Val Gly Glu Ser Val Ser Tyr Ser

195 200 205

Ile His Ser Thr Ala Lys Val Val Leu Thr Arg Glu Asp Val His Ser

210 215 220

Gln Val Ile Cys Glu Val Ala His Val Thr Leu Gln Gly Asp Pro Leu

225 230 235 240

Arg Gly Thr Ala Asn Leu Ser Glu Thr Ile Arg Val Pro Pro Thr Leu

245 250 255

Glu Val Thr Gln Gln Pro Val Arg Ala Glu Asn Gln Val Asn Val Thr

260 265 270

Cys Gln Val Arg Lys Phe Tyr Pro Gln Arg Leu Gln Leu Thr Trp Leu

275 280 285

Glu Asn Gly Asn Val Ser Arg Thr Glu Thr Ala Ser Thr Val Thr Glu

290 295 300

Asn Lys Asp Gly Thr Tyr Asn Trp Met Ser Trp Leu Leu Val Asn Val

305 310 315 320

Ser Ala His Arg Asp Asp Val Lys Leu Thr Cys Gln Val Glu His Asp

325 330 335

Gly Gln Pro Ala Val Ser Lys Ser His Asp Leu Lys Val Ser Ala His

340 345 350

Pro Lys Glu Gln Gly Ser Asn Thr Ala Ala Glu Asn Thr Gly Ser Asn

355 360 365

Glu Arg Asn Ile Tyr Ile Val Val Gly Val Val Cys Thr Leu Leu Val

370 375 380

Ala Leu Leu Met Ala Ala Leu Tyr Leu Val Arg Ile Arg Gln Lys Lys

385 390 395 400

Ala Gln Gly Ser Thr Ser Ser Thr Arg Leu His Glu Pro Glu Lys Asn

405 410 415

Ala Arg Glu Ile Thr Gln Asp Thr Asn Asp Ile Thr Tyr Ala Asp Leu

420 425 430

Asn Leu Pro Lys Gly Lys Lys Pro Ala Pro Gln Ala Ala Glu Pro Asn

435 440 445

Asn His Thr Glu Tyr Ala Ser Ile Gln Thr Ser Pro Gln Pro Ala Ser

450 455 460

Glu Asp Thr Leu Thr Tyr Ala Asp Leu Asp Met Val His Leu Asn Arg

465 470 475 480

Thr Pro Lys Gln Pro Ala Pro Lys Pro Glu Pro Ser Phe Ser Glu Tyr

485 490 495

Ala Ser Val Gln Val Pro Arg Lys

500

<210> 63

<211> 117

<212> Protein

<213> Artificial sequence

<220>

<223> Human SIRP truncated domain B variant 1

<400> 63

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

1 5 10 15

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Ile Pro Val Gly

20 25 30

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Glu Leu Ile Tyr

35 40 45

Asn Gln Lys Glu Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

50 55 60

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

65 70 75 80

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

85 90 95

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

100 105 110

Arg Ala Lys Pro Ser

115

<210> 64

<211> 373

<212> Protein

<213> Artificial sequence

<220>

<223> Human SIRP extracellular domain B domain

<400> 64

Met Glu Pro Ala Gly Pro Ala Pro Gly Arg Leu Gly Pro Leu Leu Cys

1 5 10 15

Leu Leu Leu Ala Ala Ser Cys Ala Trp Ser Gly Val Ala Gly Glu Glu

20 25 30

Glu Leu Gln Val Ile Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly

35 40 45

Glu Thr Ala Thr Leu Arg Cys Thr Ala Thr Ser Leu Ile Pro Val Gly

50 55 60

Pro Ile Gln Trp Phe Arg Gly Ala Gly Pro Gly Arg Glu Leu Ile Tyr

65 70 75 80

Asn Gln Lys Glu Gly His Phe Pro Arg Val Thr Thr Val Ser Asp Leu

85 90 95

Thr Lys Arg Asn Asn Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr

100 105 110

Pro Ala Asp Ala Gly Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser

115 120 125

Pro Asp Asp Val Glu Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val

130 135 140

Arg Ala Lys Pro Ser Ala Pro Val Val Ser Gly Pro Ala Ala Arg Ala

145 150 155 160

Thr Pro Gln His Thr Val Ser Phe Thr Cys Glu Ser His Gly Phe Ser

165 170 175

Pro Arg Asp Ile Thr Leu Lys Trp Phe Lys Asn Gly Asn Glu Leu Ser

180 185 190

Asp Phe Gln Thr Asn Val Asp Pro Val Gly Glu Ser Val Ser Tyr Ser

195 200 205

Ile His Ser Thr Ala Lys Val Val Leu Thr Arg Glu Asp Val His Ser

210 215 220

Gln Val Ile Cys Glu Val Ala His Val Thr Leu Gln Gly Asp Pro Leu

225 230 235 240

Arg Gly Thr Ala Asn Leu Ser Glu Thr Ile Arg Val Pro Pro Thr Leu

245 250 255

Glu Val Thr Gln Gln Pro Val Arg Ala Glu Asn Gln Val Asn Val Thr

260 265 270

Cys Gln Val Arg Lys Phe Tyr Pro Gln Arg Leu Gln Leu Thr Trp Leu

275 280 285

Glu Asn Gly Asn Val Ser Arg Thr Glu Thr Ala Ser Thr Val Thr Glu

290 295 300

Asn Lys Asp Gly Thr Tyr Asn Trp Met Ser Trp Leu Leu Val Asn Val

305 310 315 320

Ser Ala His Arg Asp Asp Val Lys Leu Thr Cys Gln Val Glu His Asp

325 330 335

Gly Gln Pro Ala Val Ser Lys Ser His Asp Leu Lys Val Ser Ala His

340 345 350

Pro Lys Glu Gln Gly Ser Asn Thr Ala Ala Glu Asn Thr Gly Ser Asn

355 360 365

Glu Arg Asn Ile Tyr

370

<210> 65

<211> 108

<212> Protein

<213> Artificial sequence

<220>

<223> IgV domain

<400> 65

Gln Pro Asp Lys Ser Val Leu Val Ala Ala Gly Glu Thr Ala Thr Leu

1 5 10 15

Arg Cys Thr Ala Thr Ser Leu Ile Pro Val Gly Pro Ile Gln Trp Phe

20 25 30

Arg Gly Ala Gly Pro Gly Arg Glu Leu Ile Tyr Asn Gln Lys Glu Gly

35 40 45

His Phe Pro Arg Val Thr Thr Val Ser Asp Leu Thr Lys Arg Asn Asn

50 55 60

Met Asp Phe Ser Ile Arg Ile Gly Asn Ile Thr Pro Ala Asp Ala Gly

65 70 75 80

Thr Tyr Tyr Cys Val Lys Phe Arg Lys Gly Ser Pro Asp Asp Val Glu

85 90 95

Phe Lys Ser Gly Ala Gly Thr Glu Leu Ser Val Arg

100 105

<210> 66

<211> 293

<212> Protein

<213> Artificial sequence

<220>

<223> human CD47

<400> 66

Met Trp Pro Leu Val Ala Ala Leu Leu Leu Gly Ser Ala Cys Cys Gly

1 5 10 15

Ser Ala Gln Leu Leu Phe Asn Lys Thr Lys Ser Val Glu Phe Thr Phe

20 25 30

Cys Asn Asp Thr Val Val Ile Pro Cys Phe Val Thr Asn Met Glu Ala

35 40 45

Gln Asn Thr Thr Glu Val Tyr Val Lys Trp Lys Phe Lys Gly Arg Asp

50 55 60

Ile Tyr Thr Phe Asp Gly Ala Leu Asn Lys Ser Thr Val Pro Thr Asp

65 70 75 80

Phe Ser Ser Ala Lys Ile Glu Val Ser Gln Leu Leu Lys Gly Asp Ala

85 90 95

Ser Leu Lys Met Asp Lys Ser Asp Ala Val Ser His Thr Gly Asn Tyr

100 105 110

Thr Cys Glu Val Thr Glu Leu Thr Arg Glu Gly Glu Thr Ile Ile Glu

115 120 125

Leu Lys Tyr Arg Val Val Ser Trp Phe Ser Pro Asn Glu Asn Ile Leu

130 135 140

Ile Val Ile Phe Pro Ile Phe Ala Ile Leu Leu Phe Trp Gly Gln Phe

145 150 155 160

Gly Ile Lys Thr Leu Lys Tyr Arg Ser Gly Gly Met Asp Glu Lys Thr

165 170 175

Ile Ala Leu Leu Val Ala Gly Leu Val Ile Thr Val Ile Val Ile Val

180 185 190

Gly Ala Ile Leu Phe Val Pro Gly Glu Tyr Ser Leu Lys Asn Ala Thr

195 200 205

Gly Leu Gly Leu Ile Val Thr Ser Thr Gly Ile Leu Ile Leu Leu His

210 215 220

Tyr Tyr Val Phe Ser Thr Ala Ile Gly Leu Thr Ser Phe Val Ile Ala

225 230 235 240

Ile Leu Val Ile Gln Val Ile Ala Tyr Ile Leu Ala Val Val Gly Leu

245 250 255

Ser Leu Cys Ile Ala Ala Cys Ile Pro Met His Gly Pro Leu Leu Ile

260 265 270

Ser Gly Leu Ser Ile Leu Ala Leu Ala Gln Leu Leu Gly Leu Val Tyr

275 280 285

Met Lys Phe Val Glu

290

<210> 67

<211> 233

<212> Protein

<213> Artificial sequence

<220>

<223> IgG1-FC

<400> 67

Leu Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro

1 5 10 15

Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys

20 25 30

Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val

35 40 45

Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr

50 55 60

Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu

65 70 75 80

Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His

85 90 95

Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys

100 105 110

Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln

115 120 125

Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Ser Arg Asp Glu Leu

130 135 140

Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro

145 150 155 160

Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn

165 170 175

Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu

180 185 190

Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val

195 200 205

Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr Gln

210 215 220

Lys Ser Leu Ser Leu Ser Pro Gly Lys

225 230

<210> 68

<211> 229

<212> Protein

<213> Artificial sequence

<220>

<223> IgG4-FC

<400> 68

Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys Pro Ala Pro Glu Phe

1 5 10 15

Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr

20 25 30

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val

35 40 45

Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp Tyr Val Asp Gly Val

50 55 60

Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Phe Asn Ser

65 70 75 80

Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu

85 90 95

Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Gly Leu Pro Ser

100 105 110

Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro

115 120 125

Gln Val Tyr Thr Leu Pro Ser Gln Glu Glu Met Thr Lys Asn Gln

130 135 140

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala

145 150 155 160

Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr

165 170 175

Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Arg Leu

180 185 190

Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn Val Phe Ser Cys Ser

195 200 205

Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser

210 215 220

Leu Ser Leu Gly Lys

225

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Claims (39)

1. A human SIRPα domain mutant that specifically binds to the CD47 protein, wherein said human SIRPα domain contains amino acid residues at positions 33-149 of human SIRPα variant 1, said amino acid residues at positions 33-149 of human SIRPα variant 1 are shown as SEQ ID NO : 63, and said human SIRPα domain mutant contains a group of amino acid substitutions selected from the group consisting of: (1) I61L, V63I, E77I, E84K, V93L, L96S, K98R, N100G and V132L; (2) I61V, E77N, Q82S, K83R and E84H; (3) I61F, V63I, K83R, E84K and V132I; (4) I61L, E77Q, E84D, R107N and V132I; (5) I61L, V63I, E77K, K83R, E84D and N100G; (6) I61V, E77H, Q82R, K83R, E84H and R107S; (7) I61L, E77I, Q82G, E84R, V93L, L96T, N100G, R107S, G109R and V132R; (8) I61L, E77M, Q82G, K83R, E84D and V132L; (9) I61L; (10) I61F, D95H, L96S, G109H and V132S; (11) I61F, D95H, L96S, K98R, G109H and V132S; (12) I61L, E77Q, E84D, V93A, R107N and V132I; (13) E77K, L96S, N100K, G109H and V132L; (14) I61L, V63I, Q82G, E84G, D95R, L96S, N100D and V132I; (15) I61L, E77R, Q82N, K83R, E84G, V93L, D95E, L96T, K98R, N100D and V132L; (16) I61V, E77N, Q82S, K83R, E84H and V93A; (17) I61V, V63I, E77V, K83R, E84D, D95E, L96T, K98R and N100E; (18) I61L, V63I, E77V, K83R, D95E, L96S, K98R, N100D and G109R; (19) I61V, E77L, Q82G, E84G, V93L, D95E, L96T, K98R and N100G; and (20) I61L, V63I, E77N, Q82G and E84G. 2. The human SIRPα domain mutant of claim 1, wherein said human SIRPα domain mutant comprises the amino acid sequence shown in any one of SEQ ID NOs: 1-20. 3. A human SIRPα domain mutant according to any one of claims 1 to 2, having at least one of the following characteristics: (a) binds to the CD47 protein with a KD value of 1×10 ^-8 M or less; (b) specifically blocks the interaction between the CD47 protein and SIRPα; (c) does not cause a blood clotting reaction; and (d) inhibits the growth and/or proliferation of tumors or tumor cells. 4. A fusion protein that specifically binds to the CD47 protein, comprising said human SIRPα domain mutant according to any one of claims 1 to 3, and an immunoglobulin Fc region, wherein said human SIRPα domain is directly or indirectly linked to an immunoglobulin Fc region. 5. The fusion protein of claim 4, wherein said human SIRPα domain mutant is located at the N-terminus of the immunoglobulin Fc region. 6. The fusion protein of claim 4, wherein said immunoglobulin Fc region comprises a human IgG1 Fc region. 7. The fusion protein of claim 4, wherein said immunoglobulin Fc region comprises a human IgG4 Fc region. 8. The fusion protein of claim 4, wherein said immunoglobulin Fc region contains the amino acid sequence shown in one of SEQ ID NOs: 67-68. 9. A fusion protein according to any one of claims 4-8, containing the amino acid sequence indicated in one of SEQ ID NOs: 21-60. 10. A composition that specifically binds to the CD47 protein, comprising an effective amount of a human SIRPα domain mutant according to any one of claims 1 to 3 or an effective amount of a fusion protein according to any one of claims 4 to 9, and optionally a pharmaceutically acceptable adjuvant. 11. The use of a human SIRPα domain mutant according to any one of claims 1 to 3 or a fusion protein according to any one of claims 4 to 9 in the preparation of a drug, wherein said drug is for the prevention or treatment of tumors, where said tumors are selected from the group consisting of from CD47 positive hematologic tumors and/or CD47 positive solid tumors. 12. Use of a human SIRPα domain mutant according to any one of claims 1 to 3 or a fusion protein according to any of claims 4 to 9 in preparing a kit, wherein said kit is for the prevention or treatment of tumors, wherein said tumors are selected from the group consisting of CD47 -positive hematological tumors and/or CD47-positive solid tumors. 13. The use of a human SIRPα domain mutant according to any one of claims 1 to 3 or a fusion protein according to any of claims 4 to 9 in obtaining a drug, where said drug is for the prevention or treatment of autoimmune diseases, where said autoimmune diseases are selected from the group consisting of Crohn's disease, allergic asthma and/or rheumatoid arthritis. 14. The use of a human SIRPα domain mutant according to any one of claims 1-3 or a fusion protein according to any one of claims 4-9 in obtaining a kit, where said kit is for the prevention or treatment of autoimmune diseases, where said autoimmune diseases are selected from the group consisting from Crohn's disease, allergic asthma and/or rheumatoid arthritis. 15. A method for blocking an interaction between a CD47 protein and SIRPα, said method comprising administering a fusion protein according to any one of claims 4 to 9 or a composition according to claim 10 to an individual or cell or biological sample in need thereof.

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