Chimeric antigen receptor factories and methods of use thereof

Abstract

To provide a CAR T cell therapy against solid tumors such as renal cell carcinoma.SOLUTION: Provided herein is a chimeric antigen receptor of a CAR T cell targeting CAIX and CD70 as well as a cell comprising the same. Further the cell secretes a monoclonal antibody locally at a tumor site.SELECTED DRAWING: Figure 28

Description

[Technical Field]

[0001] This application claims priority from U.S. Provisional Application No. 62 / 773,885, filed November 30, 2018, and U.S. Provisional Application No. 62 / 826,462, filed March 29, 2019, the entire contents of each of which are incorporated herein by reference.

[0002] All patents, patent applications, and publications cited herein are incorporated by reference in their entirety. The disclosures of these publications are incorporated by reference into this application in order to more fully describe the state of the art known to those skilled in the art at the date of the invention described and claimed herein.

[0003] This patent disclosure contains material that is subject to copyright protection. The copyright owner has no objection to the facsimile reproduction of either the patent document or the patent disclosure, as it appears in the U.S. Patent and Trademark Office patent file or records, but otherwise reserves all copyright rights whatsoever.

[0004] Incorporation by reference of sequence listing The contents of the text file named "[ ]" created at [ ] is incorporated herein by reference in its entirety.

[0005] FIELD OF THE INVENTION The present invention is directed to chimeric antigen receptors and cells containing same, which cells further secrete monoclonal antibodies locally at the tumor site. [Background technology]

[0006] Background of the Invention Clear cell renal cell carcinoma (ccRCC) is the major type of RCC and is one of the 10 most common cancers in both men and women. Other types of renal cancer include papillary renal cell carcinoma, pigmented renal cell carcinoma, and other or unclassified types of renal cell carcinoma. See, for example, Lancet 373 (2009) 1119-32 (Non-Patent Document 1). [Prior art documents] [Non-patent literature]

[0007] [Non-Patent Document 1] Lancet 373(2009)1119-32 Summary of the Invention

[0008] Other objects and advantages of the present invention will become readily apparent from the following description.

[0009] Aspects of the present invention are directed to engineered cells comprising a chimeric antigen receptor. In embodiments, the chimeric antigen receptor comprises an extracellular ligand-binding domain specific for a first antigen and a second antigen on the surface of a cancer cell, wherein the first antigen comprises CAIX and the second antigen comprises CD70.

[0010] In embodiments, the CAR further comprises a transmembrane polypeptide, an intracellular signaling domain, and / or a costimulatory domain.

[0011] In embodiments, the extracellular ligand-binding domain comprises an antibody or fragment thereof. For example, the antibody comprises a VH and / or VL according to Table 2, or any combination thereof. For example, the antibody comprises a VH and / or VL according to Table 4, or any combination thereof. For example, the extracellular binding domain comprises a VH and / or VL from Table 2 and Table 4, or any combination thereof. For example, the antibody comprises a CDR1, CDR2, and / or CDR3 from Table 1, or any combination thereof. For example, the antibody comprises a CDR1, CDR2, and / or CDR3 from Table 3, or any combination thereof. For example, the extracellular binding domain comprises a CDR1, CDR2, and / or CDR3 from Table 1 and Table 3, or any combination thereof.

[0012] In embodiments, the engineered cells express and secrete the recombinant polypeptide.

[0013] In embodiments, the recombinant polypeptide comprises an antibody or a fragment thereof, or a cytokine. For example, the recombinant polypeptide comprises an antibody or a fragment thereof specific for TIGIT, GITR, PD-L1, PD-L2, PD-1, CTLA-4, VISTA, CD70, TIM-3, LAG-3, CD40L, or CCR4. For example, the recombinant polypeptide comprises a cytokine including IL-12, IL-15, or IL-18.

[0014] In embodiments, the recombinant polypeptide modulates the immune system of a subject, for example, the recombinant polypeptide is an immune checkpoint blockade antibody.

[0015] In embodiments, the recombinant polypeptide modulates tumor angiogenesis. For example, the recombinant polypeptide can be specific for VEGF, VEGFR1, VEGFR2, PDGF, Ang-1, or AT1.

[0016] In some embodiments, the engineered cells are T cells, NK cells, or NKT cells. For example, the T cells are CD4+, CD8+, CD3+ panT cells, or any combination thereof. For example, the T cells are a mixed population of CD4+ T cells and CD8+ T cells.

[0017] Aspects of the present disclosure are further directed to nucleic acid constructs encoding chimeric antigen receptors. In embodiments, the chimeric antigen receptor comprises an extracellular ligand-binding domain specific for a first antigen and a second antigen on the surface of a cancer cell, wherein the first antigen comprises CAIX and the second antigen comprises CD70.

[0018] In embodiments, the nucleic acid construct further encodes a transmembrane polypeptide, an intracellular signaling domain, and / or a costimulatory domain.

[0019] In embodiments, the nucleic acid construct further encodes a recombinant polypeptide.

[0020] Aspects of the present disclosure are further directed to vectors comprising the nucleic acid constructs described herein.

[0021] Additionally, aspects of the present disclosure are directed to cells comprising the vectors described herein.

[0022] Aspects of the present disclosure are also directed to methods for treating a subject suffering from cancer, in embodiments, the method comprises administering to the subject a therapeutically effective amount of the engineered cells described herein.

[0023] Aspects of the invention are also directed to methods of reducing the progression of or promoting the regression of cancer in a subject, in embodiments, the methods comprising administering to the subject a therapeutically effective amount of the engineered cells described herein.

[0024] Furthermore, aspects of the invention are directed to methods of reducing cell proliferation of cancer cells in a subject. In embodiments, the method comprises administering to the subject a therapeutically effective amount of the engineered cells described herein.

[0025] In embodiments, the cancer comprises renal cell carcinoma.

[0026] Embodiments of the present disclosure are directed to chimeric antigen receptors (CARs) comprising an extracellular ligand-binding domain specific for a first antigen and a second antigen on the surface of a cancer cell, wherein the first antigen comprises CAIX and the second antigen comprises CD70.

[0027] In embodiments, the CAR further comprises a transmembrane polypeptide, an intracellular signaling domain, and / or a costimulatory domain.

[0028] In embodiments, the extracellular ligand-binding domain comprises an antibody or a fragment thereof.

[0029] Additionally, aspects of the present invention are directed to cells comprising the chimeric antigen receptors (CARs) described herein.

[0030] Further aspects of the present invention are directed to engineered cells comprising a first chimeric antigen receptor and a second chimeric antigen receptor, wherein the first chimeric antigen receptor comprises an extracellular ligand-binding domain specific for CAIX and the second chimeric antigen receptor comprises an extracellular ligand-binding domain specific for CD70.

[0031] In embodiments, the engineered cells express and secrete the recombinant polypeptide.

[0032] In embodiments, the first chimeric antigen receptor and the second chimeric antigen receptor are expressed from a single nucleic acid construct. [The present invention 1001] 1. An engineered cell comprising a chimeric antigen receptor, the chimeric antigen receptor comprising an extracellular ligand-binding domain specific for a first antigen and a second antigen on the surface of a cancer cell, the first antigen comprising CAIX and the second antigen comprising CD70. [The present invention 1002] 1001. The engineered cell of claim 1001, wherein said CAR further comprises a transmembrane polypeptide and an intracellular signaling domain. [The present invention 1003] 1002. The engineered cell of claim 1002, wherein the CAR further comprises a costimulatory domain. [The present invention 1004] 1001. The engineered cell of claim 1001, wherein said extracellular ligand-binding domain comprises an antibody or a fragment thereof. [The present invention 1005] 1005. The engineered cell of claim 10, wherein said antibody comprises a VH and / or VL according to Table 2, or any combination thereof. [The present invention 1006] 1005. The engineered cell of the present invention, wherein said antibody comprises a VH and / or VL according to Table 4, or any combination thereof. [The present invention 1007] 1005. The engineered cell of the present invention, wherein said extracellular binding domain comprises a VH and / or VL of Table 2 and Table 4, or any combination thereof. [The present invention 1008] 1005. The engineered cell of claim 1004, wherein said antibody comprises CDR1, CDR2, and / or CDR3 of Table 1, or any combination thereof. [The present invention 1009] 1005. The engineered cell of claim 1004, wherein said antibody comprises CDR1, CDR2, and / or CDR3 of Table 3, or any combination thereof. [The present invention 1010] 1005. The engineered cell of claim 1004, wherein said extracellular binding domain comprises CDR1, CDR2, and / or CDR3 of Table 1 and Table 3, or any combination thereof. [The present invention 1011] 1001. An engineered cell of the present invention that expresses and secretes a recombinant polypeptide. [The present invention 1012] 1011. The engineered cell of claim 10, wherein said recombinant polypeptide comprises an antibody or fragment thereof, or a cytokine. [The present invention 1013] 1011. The engineered cell of claim 1011, wherein said recombinant polypeptide modulates the immune system of a subject. [The present invention 1014] 1011. The engineered cell of the present invention, wherein said recombinant polypeptide is an immune checkpoint blocking antibody. [The present invention 1015] 1011. The engineered cell of claim 1011, wherein said recombinant polypeptide regulates tumor angiogenesis. [The present invention 1016] 1015. The engineered cell of claim 10, wherein said recombinant polypeptide is specific for VEGF, VEGFR1, VEGFR2, PDGF, Ang-1, or AT1. [The present invention 1017] 1011. The engineered cell of the present invention, wherein the recombinant polypeptide comprises an antibody or fragment thereof specific for TIGIT, GITR, PD-L1, PD-L2, PD-1, CTLA-4, VISTA, CD70, TIM-3, LAG-3, CD40L, or CCR4. [The present invention 1018] 1012. The engineered cell of claim 10, wherein said cytokine comprises IL-12, IL-15, or IL-18. [The present invention 1019] 1001. The engineered cell of claim 1001, wherein said cell comprises a T cell, an NK cell, or an NKT cell. [The present invention 1020] 1019. The engineered cell of the present invention, wherein said T cells are CD4+, CD8+, CD3+ pan T cells, or any combination thereof. [The present invention 1021] 1019. The engineered cell of the present invention, wherein said T cells are a mixed population of CD4+ T cells and CD8+ T cells. [The present invention 1022] 1. A nucleic acid construct encoding a chimeric antigen receptor, wherein the chimeric antigen receptor comprises an extracellular ligand-binding domain specific for a first antigen and a second antigen on the surface of a cancer cell, wherein the first antigen comprises CAIX and the second antigen comprises CD70. [The present invention 1023] 1022. The nucleic acid construct of claim 1022, wherein the chimeric antigen receptor further comprises a transmembrane polypeptide and an intracellular signaling domain. [The present invention 1024] The nucleic acid construct of claim 1023, wherein the chimeric antigen receptor further comprises a costimulatory domain. [The present invention 1025] The nucleic acid construct of the invention 1023, further encoding a recombinant polypeptide. [The present invention 1026] 1025. The nucleic acid construct of claim 1025, wherein said recombinant polypeptide is capable of being secreted from an engineered cell. [The present invention 1027] 1025. The nucleic acid construct of claim 1025, wherein said recombinant polypeptide comprises an antibody or a fragment thereof, or a cytokine. [The present invention 1028] 1025. The nucleic acid construct of claim 1025, wherein said recombinant polypeptide modulates the immune system of a subject. [The present invention 1029] 1025. The nucleic acid construct of the present invention, wherein the recombinant polypeptide is an immune checkpoint blocking antibody. [The present invention 1030] 1025. The nucleic acid construct of claim 1025, wherein said recombinant polypeptide regulates tumor angiogenesis. [The present invention 1031] 1025. The nucleic acid construct of the present invention, wherein the recombinant polypeptide comprises an antibody or fragment thereof specific for TIGIT, GITR, PD-L1, PD-L2, PD-1, CTLA-4, VISTA, CD70, TIM-3, LAG-3, CD40L, or CCR4. [The present invention 1032] 1027. The nucleic acid of claim 1027, wherein the cytokine comprises IL12, IL15, or IL18. [The present invention 1033] 1030. The nucleic acid of claim 10, wherein said recombinant polypeptide is specific for VEGF, VEGFR1, VEGFR2, PDGF, Ang-1, or AT1. [The present invention 1034] A vector comprising the nucleic acid construct of the present invention. [This invention 1035] A cell containing the vector of the present invention. [The present invention 1036] A method of treating a subject suffering from cancer, comprising administering to said subject a therapeutically effective amount of engineered cells of the present invention. [This invention 1037] 10. A method of reducing the progression or promoting the regression of cancer in a subject, comprising administering to said subject a therapeutically effective amount of an engineered cell of the present invention. [The present invention 1038] 10. A method of reducing cell proliferation of cancer cells in a subject, comprising administering to said subject a therapeutically effective amount of an engineered cell of the present invention. [This invention 1039] The method of any one of claims 1036 to 1038, wherein the cancer comprises renal cell carcinoma. [The present invention 1040] A chimeric antigen receptor (CAR) comprising an extracellular ligand-binding domain, wherein the extracellular ligand-binding domain is specific for a first antigen and a second antigen on the surface of a cancer cell, wherein the first antigen comprises CAIX and the second antigen comprises CD70. [The present invention 1041] The CAR of the present invention 1040, further comprising a transmembrane polypeptide and an intracellular signaling domain. [The present invention 1042] The CAR of the present invention 1041, further comprising a costimulatory domain. [This invention 1043] The CAR of the present invention 1040, wherein the extracellular ligand-binding domain comprises an antibody or a fragment thereof. [This invention 1044] A cell comprising the chimeric antigen receptor (CAR) of the present invention. [This invention 1045] An engineered cell comprising a first chimeric antigen receptor and a second chimeric antigen receptor, wherein the first chimeric antigen receptor comprises an extracellular ligand-binding domain specific for CAIX and the second chimeric antigen receptor comprises an extracellular ligand-binding domain specific for CD70. [The present invention 1046] 1045. An engineered cell of the present invention that expresses and secretes a recombinant polypeptide. [This invention 1047] 1046. The engineered cell of claim 1046, wherein said recombinant polypeptide comprises an antibody or fragment thereof, or a cytokine. [This invention 1048] 1046. The engineered cell of claim 1046, wherein said recombinant polypeptide modulates the immune system of a subject. [This invention 1049] 1046. The engineered cell of claim 1046, wherein said recombinant polypeptide is an immune checkpoint blockade antibody. [The present invention 1050] 1046. The engineered cell of claim 1046, wherein said recombinant polypeptide regulates tumor angiogenesis. [This invention 1051] 1046. The engineered cell of claim 1046, wherein said recombinant polypeptide comprises an antibody or fragment thereof specific for TIGIT, GITR, PD-L1, PD-L2, PD-1, CTLA-4, VISTA, CD70, TIM-3, LAG-3, CD40L, or CCR4. [This invention 1052] 1047. The engineered cell of claim 1047, wherein said cytokine comprises IL12, IL15, or IL18. [This invention 1053] 1050. The nucleic acid of claim 1050, wherein said recombinant polypeptide is specific for VEGF, VEGFR1, VEGFR2, PDGF, Ang-1, or AT1. [This invention 1054] 1045. The engineered cell of claim 1045, wherein said cell comprises a T cell or an NK cell. [This invention 1055] 1054. The engineered cell of claim 1054, wherein said T cells are CD4+, CD8+, CD3+ pan T cells, or any combination thereof. [The present invention 1056] 1054. The engineered cell of claim 10, wherein said T cells are a mixed population of CD4+ T cells and CD8+ T cells. [This invention 1057] 1045. The engineered cell of claim 1045, wherein said first chimeric antigen receptor and said second chimeric antigen receptor are expressed from a single nucleic acid construct. [Brief explanation of the drawings]

[0033] [Figure 1]Figure 1 shows a schematic diagram of bispecific tandem CAR T anti-CD70 and anti-CAIX scFvs combined in different permutations by changing the order of two targeting scFvs with various linkers and hinges connected to costimulatory domains (non-limiting examples include CD28, 41BB, CD28-41BB, or 41BB-CD28) and activation domains (such as CD3). Second-generation CAR T cell factories can be engineered with the introduction of a second cassette anti-CD70 scFv to assess efficacy (e.g., address heterogeneity) and safety (e.g., limit on-target off-tumor effects). [Figure 2] This study demonstrates that second-generation CAR T cell factories can be established by combining a series of CARs (such as different scFvs, linkers, and hinges) with immune checkpoint blockade payloads. Based on these data, eight constructs were established using G36 as the anti-CAIX scFv and B7 as the anti-CD70 scFv. Those skilled in the art will recognize that any scFv can be used according to the present invention. Dual CAR engineering is the most important part, and the payload was replaced with zsgreen to demonstrate conversion efficiency. [Figure 3] This image shows CAIX and CD70 being upregulated and coexpressed in ccRCC. IHC staining of ccRCC primary cell lines generated from patient samples indicates that both CAIX and CD70 are highly expressed and coexpressed in ccRCC. Without wishing to be bound by theory, CAIX and CD70 are two potential targets for ccRCC therapy. To validate the targets, IHC was performed for CAIX and CD70 staining in primary cell lines generated from patients with ccRCC. These images show that CAIX and CD70 staining was 100% positive, demonstrating that CAIX and CD70 are highly and coexpressed in ccRCC. This is currently being validated in a study measuring CAIX and CD70 expression in patients with ccRCC in at least 150 samples. [Figure 4]Figure 1 shows CAIX and CD70 are upregulated and co-expressed in ccRCC. IHC staining of ccRCC patient samples shows that both CAIX and CD70 are highly expressed and co-expressed on ccRCC. [Figure 5-1] Establishment of CRISPR skrc-59 cell lines is shown. Four CRISPR-engineered skrc-59 cell lines with four distinct phenotypes (i) CAIX+CD70+, (ii) CAIX+CD70-, (iii) CAIX-CD70+, and (iv) CAIX-CD70- were established for further in vitro evaluation. Four CRISPR skrc-59 cell lines with four distinct phenotypes (i) CAIX+CD70+, (ii) CAIX+CD70-, (iii) CAIX-CD70+, and (iv) CAIX-CD70- were used in the in vitro assays described herein. The corresponding table shows the quantified expression levels of CAIX and CD70 in the four different cell lines. [Figure 5-2] See the description of Figure 5-1. [Figure 6] This graph shows anti-CD70 minibodies that showed selective binding to CD70+ SKRC59 cells. Phage display (panned against CD70+ SKRC-59 cells and subtracted against CD70- SKRC-59 cells) shows that a series of anti-CD70 minibodies showed binding to CD70-positive ccRCC SKRC-59 cells. Anti-CD70 minibodies were expressed on Expi293 cells in 6-well plates. Three days after transfection, supernatants were collected and an IgG quantification ELISA (Bethyl) was performed to estimate the minibody concentration in the supernatant. This approximate concentration was used to normalize the supernatant for FACS binding curves. Staining was performed via a standard FACS staining protocol using an anti-hFc-APC secondary. As can be seen, one of the killer hits (#9) is highly nonspecific. The other two killer monoCARs (#3, #7) show good specificity for CD70. [Figure 7]We demonstrate that anti-CD70 CAR T cells exhibit killing activity in a Celigo killing assay. These anti-CD70 scFv candidates were cloned into vectors, packaged into lentivirus, and transduced into primary T cells. The CAR T cells were evaluated for antiproliferative activity in a Celigo assay. Results showed that #3, #7, and #9 had efficacy compared with CD70 ligand CD27 CAR T cells. These hits were also cloned into a pHAGE vector and subjected to a Celigo killing assay. These graphs show that anti-CD70 CAR T cells exhibited killing activity in a Celigo killing assay, with an effector:target ratio of 2:1. Cell numbers at different time points were compared and normalized to untreated cells at the corresponding time points. Therefore, the Y axis represents the percentage of treated cells / untreated cells, and the X axis represents the different treatments. After 24 hours of co-culture with T cells, the number of SKRC59 cells in the treated group was significantly reduced compared to the untreated and untransduced T cell groups, demonstrating the anti-proliferative activity of these CAR T cells. After 48 hours of co-culture, we found three candidates that matched CD27, the ligand for CD70. Therefore, 3, 7, and 9 were used in further chromium 51 killing assays. [Figure 8] We demonstrate that anti-CD70 CAR T cells exhibit killing activity in a chromium 51 killing assay against CAIX+CD70+ cells. CAR T cells were also evaluated for killing activity in a chromium 51 release assay. The results showed that CAR T cells #7 had enhanced efficacy compared to CD27 (CD70 ligand) CAR T cells. A 4-hour chromium 51 release assay was performed. After 4 hours of incubation with chromium 51-labeled target cells, B7 was verified to have killing activity against SKRC59 CD70+ cells. [Figure 9]Second-generation CAR-T cells are shown. CAR T cells can be generated using zsGreen instead of immune checkpoint blockade. Based on these data, eight constructs were established using G36 as the anti-CAIX scFv and B7 as the anti-CD70 scFv. Dual CAR engineering is the most important part, and the payload was replaced with zsGreen to demonstrate transduction efficiency. [Figure 10] Figure 1 shows transfected 293T cells binding to CAIX-PE. 293T cells were transfected with different constructs of the dual CAR and subjected to binding assays with PE-labeled CAIX protein. All dual CARs bind to CAIX, and the different orientations of the two scFvs affect the EC50 of anti-CAIX scFv binding. Anti-CAIX scFv G36 favors the second cassette after the linker. For example, 293T cells were transfected with these eight bispecific constructs and the corresponding mono-CARs and stained with CAIX-PE. After normalization by transfection efficiency, the different orientations of the two scFvs affect the EC50 of anti-CAIX scFv binding. Anti-CAIX scFv G36 favors the second cassette after the linker. [Figure 11] Figure 1 shows transfected 293T cells binding to CD70-APC. 293T cells were transfected with different constructs of the dual CAR and subjected to binding assays using PE-labeled CAIX protein. All dual CARs bind to CD70, and the different orientations of the two scFvs do not affect the EC50 of anti-CD70 scFv binding. Anti-CD70 scFv B7 does not have a significant preference. For example, 293T cells were transfected with these eight bispecific constructs and the corresponding mono-CARs and stained with CD70-PE. After normalization by transfection efficiency, the different orientations of the two scFvs do not affect the EC50 of anti-CD70 scFv binding. Anti-CD70 scFv B7 does not have a significant preference. [Figure 12]Figure 1 shows a B7-GGGGS3-G36 killing assay using Celigo. For example, B7-GGGGS3-G36 CAR was used to perform selective killing assays against CAIX+ or CD70+ single-positive cells or CAIX+CD70+ double-positive cells mixed with CAIX-CD70- cells. B7-GGGGS3-G36 CAR T cells had greater killing activity against target cells (CAIX+CD70+) than non-target cells (CAIX+CD70-, CAIX-CD70+, CAIX-CD70-). [Figure 13] B7-GGGGS3-G36 killing assay by FACS. Four different CRISPR-engineered skrc-59 cells were transduced with BFP fluorescent groups. The cells were mixed at a 1:1:1:1 ratio and treated with B7-GGGGS3-G36 CAR T cells or medium. After treatment, the cells were stained with PE-labeled anti-CD70 antibody and APC-labeled anti-CAIX antibody and analyzed by flow cytometry. B7-GGGGS3-G36 demonstrated selective killing of CAIX+CD70+ cells, reducing the population from 26.7% to 18.5%. [Figure 14] Schematic diagram of fine-tuned anti-CAIX CAR T. Anti-CAIX scFvs with various KDs were generated as CARs, and the corresponding killing activities were evaluated. To limit on-target off-tumor effects, a second-generation CAR T cell factory was engineered by introducing an anti-CD70 scFv in a second cassette. [Figure 15] This is a diagram rendering of fine-tuned anti-CAIX CAR-T cells. In validation experiments, CAR T cells were generated using zsGreen instead of immune checkpoint blockade. Eight constructs were established by using G36 as the anti-CAIX scFv and B7 as the anti-CD70 scFv. Dual CAR engineering was the key part, and the payload was replaced with zsgreen to demonstrate transduction efficiency. A series of antibodies against CAIX were identified with different KD values ​​ranging from 1.49 nM to 99.58 nM (see also Figure 16). Binding experiments classified the 13 antibodies into four groups. [Figure 16-1] 1 depicts a table showing the 19 anti-CAIX ScFvs identified and the corresponding binding. [Figure 16-2] See the description of Figure 16-1. [Figure 17] Figure 1 shows a graph of anti-CAIX monoCAR T killing against CAIX+CD70+. These anti-CAIX CAR T cells were evaluated for antiproliferative activity in a Celigo assay. Results showed that the 19 CARs could be divided into four groups based on killing activity: G37, G39, G125(++++)>G10, G21, G36, G40, G45, G57, G62, G98, G106, G119(+++)>G6, G9, G17, G27, G28(++)>G104(++). [Figure 18] Figure 1 shows a graph of killing of anti-CAIX monoCAR T cells against CAIX+CD70+ cells. These anti-CAIX CAR T cells were evaluated for antiproliferative activity in a Celigo assay. Results showed that the 19 CARs could be divided into four groups based on killing activity: G37, G39, G125 (++++) > G10, G21, G36, G40, G45, G57, G62, G98, G106, G119 (+++) > G6, G9, G17, G27, G28 (++) > G104 (++). [Figure 19] Figure 1 shows the killing of anti-CAIX monoCAR T cells against CAIX+CD70+ cells. These anti-CAIX CAR T cells were evaluated for antiproliferative activity in the Celigo assay. The results showed that the 19 CARs could be divided into four groups based on killing activity: G37, G39, G125 (++++) > G10, G21, G36, G40, G45, G57, G62, G98, G106, G119 (+++) > G6, G9, G17, G27, G28 (++) > G104 (++). [Figure 20]Schematic of selective killing assay (e.g., using either Celigo assay or FACS assay). Two different selectivity assays were performed to examine the selectivity of bispecific CARs for four different cell lines. For example, CAIX+CD70+ cells were mixed with mono-positive cells, and then CART cells were added. After a certain period of incubation, the number of double-positive and single-positive cells was measured by Celigo. Alternatively, the four different cell lines were mixed at a 1:1:1:1 ratio, and CART cells were added. After 24 hours of coculture, all cells were collected and stained for CAIX-APC and CD70-PE by FACS. [Figure 21] Graph showing tandem CAR killing against skrc-59 mixed cells. Selective killing assays were performed on CAIX+ or CD70+ single-positive cells or CAIX+CD70+ double-positive cells mixed with CAIX-CD70- cells. Bispecific CAR T cells had greater killing activity against target cells (CAIX+CD70+) than non-target cells (CAIX+CD70-, CAIX-CD70+, CAIX-CD70-) at an E:T ratio of 5:1. [Figure 22] Graph showing tandem CAR killing against skrc-59 mixed cells. Selective killing assays were performed on CAIX+ or CD70+ single-positive or CAIX+CD70+ double-positive cells mixed with CAIX-CD70- cells. Bispecific CAR T cells had greater killing activity against target cells (CAIX+CD70+) than non-target cells (CAIX+CD70-, CAIX-CD70+, CAIX-CD70-) at an E:T ratio of 5:1. [Figure 23] Graph showing tandem CAR killing in skrc-59 mixed cells. Selective killing assays were performed on CAIX+ or CD70+ single-positive or CAIX+CD70+ double-positive cells mixed with CAIX-CD70- cells. Bispecific CAR T cells had greater killing activity against target cells (CAIX+CD70+) than non-target cells (CAIX+CD70-, CAIX-CD70+, CAIX-CD70-) at an E:T ratio of 10:1. [Figure 24] CAIX and CD70 are shown to be upregulated and co-expressed in ccRCC. [Figure 25] We demonstrate the design of CARs to address undesirable side effects previously associated with CAR T cell therapy. To evaluate efficacy and safety profiles for CAR T cell therapy in solid tumors, we first developed a CAR T cell factory to restore effective anti-cancer immunity. It can locally secrete human anti-immune checkpoint blockade monoclonal antibodies (mAbs) at tumor sites to modulate the tumor microenvironment by reversing T cell exhaustion. To limit on-target off-tumor side effects, fine-tuned CARs were engineered with reduced-affinity scFvs to expand the therapeutic window by limiting tumor-associated antigen recognition on normal tissues. Furthermore, bispecific CARs were engineered by incorporating a second cassette, such as an anti-CD70 scFv, to increase preferential killing in the dual-positive population and improve safety profiles. [Figure 26] We demonstrate that CAR-T cell factories exhibit enhanced killing. We designed a bicistronic lentiviral vector to express an anti-CAIX scFv linked to CD28 and CD3z signaling domains in one cassette and an anti-PDL1 mAb in the second. Thus, the CAR-T cell factory can target CAIX and secrete a checkpoint blockade inhibitor at the tumor site, transforming the suppressive tumor microenvironment. In an orthotopic RCC mouse model, 1E7 CAR-T cells were injected intravenously on day 0, and 2.5E6 CAR-T cells were injected on day 17. In our orthotopic RCC mouse model, the CAR-T cell factory demonstrated enhanced killing compared with CAR-T cells secreting an irrelevant antibody. [Figure 27] A schematic diagram of the evaluation of CAR-T on a humanized orthotopic ccRCC mouse model is shown. [Figure 28] MRI images of a mouse model of RCC are shown. [Figure 29]A schematic diagram of the experiments designed to establish a stress model and compare second and third generation CARs and CD8 versus CD4 / 8 on an orthotopic ccRCC mouse model is shown. [Figure 30] These results demonstrate that 41BB exhibited superior tumor killing in vivo. At a dose of 1E7, efficient tumor regression was observed in the G36-41BB and G36-CD28-41BB-treated groups. The mixed CD4 / CD8 G36-41BB CAR-T exhibited superior tumor killing even at a dose of 3E6. Comparing two second-generation CAR constructs and one third-generation CAR construct, G36-41BB was more efficient than G36-CD28 and G36-CD28-41BB. [Figure 31] 1 shows the G36-41BB stress model. 3E6 administration can be used in the G36-41BB stress model. [Figure 32] This shows that 41BB exhibited superior killing in vivo. [Figure 33] 41BB CAR-T cells showed the highest proliferation in vivo. [Figure 34] CD8 versus CD4 / CD8 is shown. [Figure 35] 1 shows the in vivo proliferation of CD4 and CD8 T cells. [Figure 36] A schematic diagram of a fine-tuned anti-CAIX CAR-T designed to limit on-target off-tumor effects is shown. Without wishing to be bound by theory, reducing the affinity of the anti-CAIX scFv allows the CAR to recognize only the high-density CAIX on ccRCC. A series of anti-CAIX scFvs with various KDs were cloned into lentiviral vectors, and corresponding CAR-T cells were generated and evaluated for killing activity. [Figure 37]Detailed diagram of the "therapeutic window." The term "therapeutic window," originally derived from pharmaceutical toxicology, refers to the dose range between efficacy and toxicity, achieving the best therapeutic effect without unacceptable toxicity. It is the range between the minimum effective dose (MED) and the maximum tolerated dose (MTD). This concept has been applied to optimizing CAR-T therapy. To expand the therapeutic window, the CAR affinity to the antigen can be fine-tuned by assembling scFvs with different KDs within the CAR construct, such as from 1 nm to 100 nm. Ideally, after optimization, the CAR will recognize only high-density antigens on tumor cells, not low-density antigens on normal cells. Targeting antigens expressed only on tumor cells or non-critical tissues widens the therapeutic window because it avoids direct toxicity in key tissues. On the other hand, targeting antigens expressed in critical normal tissues / cells narrows the therapeutic window by decreasing the MTD. [Figure 38] FIG. 1 is a schematic illustrating widening the therapeutic window to address on-target off-tumor side effects. [Figure 39] IHC double staining of CAIX and CD70 on patient samples is shown. CD70 and CAIX were found to be highly expressed and co-expressed on ccRCC. Therefore, CD70 was selected as the second target. [Figure 40] IHC double staining for CAIX and CD70 on patient samples is shown. [Figure 41] As shown in the table, a panel of scFvs against CAIX with different KD values ​​ranging from 1.49 nM to 99.58 nM were generated and screened by the Celigo killing assay. [Figure 42]Demonstrating a correlation between scFv affinity and CART cell killing, 19 CARs were divided into four groups according to cytotoxicity and tested against skrc-59 cells with different levels of CAIX expression: G37, G39, G125>G10, G21, G36, G40, G45, G57, G62, G98, G106, G119>G6, G9, G17, G27, G28>G104. [Figure 43] "Or" gating is shown, which captures tumor cell heterogeneity but does not kill healthy cells at low target density. [Figure 44] CD70 is highly expressed on kidney cancer, especially clear cell renal cell carcinoma, making it an ideal second target for bispecific CARs. [Figure 45] Figure 1 shows IHC double staining of CAIX and CD70 on ccRCC patient samples. CD70 and CAIX are highly expressed and co-expressed on ccRCC. Therefore, CD70 is selected as the second target. [Figure 46] Figure 1 shows IHC double staining of CAIX and CD70 on ccRCC patient samples. CD70 and CAIX are highly expressed and co-expressed on ccRCC. Therefore, CD70 was selected as the second target. [Figure 47] CAIX and CD70 are shown to be upregulated and co-expressed on ccRCC. [Figure 48]This figure shows that anti-CD70 minibodies exhibited selective binding to CD70+ SKRC-59 cells. Phage display (panned against CD70+ SKRC-59 cells and subtracted against CD70- SKRC-59 cells) identified a series of anti-CD70 minibodies that showed promising binding to CD70-positive ccRCC SKRC-59 cells. Anti-CD70 minibodies were expressed on Expi293 cells in 6-well plates. Three days after transfection, supernatants were collected and IgG quantification ELISA (Bethyl) was performed to estimate the minibody concentration in the supernatant. This approximate concentration was used to normalize the supernatant for FACS binding curves. Staining was performed via a standard FACS staining protocol using an anti-hFc-APC secondary. One of the killer hits (#9) was highly nonspecific. The other two killer monoCARs (#3, #7) showed good specificity for CD70. [Figure 49] We demonstrate that anti-CD70 B7 CAR-T cells demonstrated promising killing of CD70+ skrc-59 cells. Phage display (panned against CD70+ skrc-59 cells and subtracted against CD70- skrc-59 cells) identified a series of anti-CD70 minibodies that showed promising binding to CD70-positive ccRCC skrc-59 cells. These hits were then cloned into lentiviral vectors and subjected to Celigo killing assays with the corresponding CAR-T cells. From the screening, we identified B7 as a candidate. [Figure 50] Figure 1 shows bispecific CAR constructs. A series of constructs were established in different rotations (or orientations) with different linkers by using G36 as the anti-CAIX scFv and B7 as the anti-CD70 scFv. [Figure 51]This shows that anti-scFv G36 prefers the second cassette after the linker. 293T cells were transfected with these eight bispecific constructs and the corresponding monoCARs and stained with CAIX-PE. After normalization by transfection efficiency, this shows that the different orientations of the two scFvs affect the EC50 of anti-CAIX scFv binding. Anti-CAIX scFv G36 prefers the second cassette after the linker. [Figure 52] Anti-CD70 scFv B7 has no preference. 293T cells were transfected with different constructs of the dual CAR, and binding assays were performed with APC-labeled CD70 protein. After normalization by transfection efficiency, it was shown that the different orientations of the two scFvs did not affect the EC50 of anti-CD70 scFv binding. Anti-CD70 scFv B7 has no significant preference. [Figure 53] Establishment of four CRISPR skrc-59 cell lines. Four CRISPR-engineered skrc-59 cell lines with four distinct phenotypes were established for further evaluation in vitro: CAIX+CD70+, CAIX+CD70-, CAIX-CD70+, and CAIX-CD70-. [Figure 54] We demonstrate that B7-GGGGS3-G36 exhibits preferential killing. Using the B7-GGGGS3-G36 CAR as an example, we performed selective killing assays against CAIX+ or CD70+ single-positive cells, or CAIX+CD70+ double-positive cells mixed with CAIX-CD70- cells. B7-GGGGS3-G36 CAR-T cells were shown to have preferential killing activity against target cells (CAIX+CD70+) over non-target cells (CAIX+CD70-, CAIX-CD70+, and CAIX-CD70-). After incubation for a set period, the numbers of double-positive and single-positive cells were measured using Celigo. Furthermore, we found that B7-GGGGS3 has a restricted selectivity index for CAIX+CD70+ cells when mixed with CAIX+ cells. [Figure 55]This shows that B7-GGGGS3-G36 exhibits preferential killing. Four different CRISPR-engineered skrc-59 cells were transduced with BFP fluorescent groups. The cells were mixed at a 1:1:1:1 ratio and treated with B7-GGGGS3-G36 CAR-T cells or medium. After treatment, the cells were stained with PE-labeled anti-CD70 antibody and APC-labeled anti-CAIX antibody and analyzed by flow cytometry. B7-GGGGS3-G36 demonstrated selective killing of CAIX+CD70+ cells, reducing the population from 26.7% to 18.5%. This selective killing data from FACS showed that B7-GGGGS3-G36 exhibited selective killing of CAIX+CD70+ cells, reducing the population from 26.7% to 18.5%. [Figure 56] 1 shows a schematic diagram of a bispecific split CAR T. Anti-CD70 and anti-CAIX scFvs were expressed on the cell surface with different costimulatory domains. See, e.g., dual(split)CAR T, Nat Rev Cancer 16(9):566-81. [Figure 57] Split CAR killing in ccRCC primary cancer cells. Split CAR T cells were evaluated for killing activity against primary ccRCC cancer cells using mono-CAR T and tandem CAR T cells. It showed that split CAR achieved excellent killing at low E:T ratios, such as a 1:1 ratio. [Figure 58] 1 provides a multiple alignment of the amino acid sequences of anticarbonic anhydrase IX (G250) scFv clones. [Figure 59] 1 provides an alignment of human and mouse CAIX amino acid sequences. [Figure 60] The structure of carbonic anhydrase IX (G250) is provided. [Figure 61] CAR T cell killing assay of Skrc-59 transduced cells. Celigo image cytometry is shown. [Figure 62]Homology study of 10 different species. Tree: The distance of each branch is equal to the number of differences between the sequences (e.g., 0.1 means that there is 10% difference between the two sequences), and the distance between two species is equal to the total length of all branches connecting them. Homology > 60% → potential cross-reactivity. [Figure 63] Establishment of skrc-59 stable cell lines expressing CAIX or CD70 from different species. CRISPR knockout CAIX- / CD70-skrc-59 cells were transduced with 10 different constructs (CD70 or CAIX from 5 different species) and sorted by FACS. Half-transduced with BFP (for Celigo). Stained with commercially available antibodies. [Figure 64] Binding assay of anti-CAIX(G36) scFv is shown. Binding data was normalized to that of a commercial antibody and analyzed using nonlinear regression and a logarithmic (agonist) vs. response model. [Figure 65] Killing assay of anti-CAIX (G36) CAR T cells: 100% in monkeys (same killing as in humans, cross-reactivity), 50% in mice and hamsters (significant killing). [Figure 66] 1 shows a killing assay of anti-CD70 (B7) CAR T cells. [Figure 67] Killing assays of 20 different anti-CAIX CAR T cells are shown. [Figure 68] Fold change killing assay G36 E:T 10:1 is shown. [Figure 69] Killing assay G36 E:T 5:1 is shown. [Figure 70] Killing assay G36 n=2 is shown. [Figure 71] Killing Assays Twenty anti-CAIX scFvs are shown. [Figure 72] All candidate monkeys are shown. [Figure 73] All candidate mice are shown. [Figure 74] All candidate hamsters are shown. [Figure 75] Fold change B7 killing assay is shown. [Figure 76-1] 1 shows the amino acid sequence and germline alignment of anti-cd70 antibodies. [Figure 76-2] See the description of Figure 76-1. [Figure 77] 1 shows the nucleic acid construct of split car. [Figure 78] The amino acid sequences and germline alignments of the anti-PDL1 and anti-PD1 sequences are shown. [Figure 79] 1 shows the amino acid sequences and germline alignments of anti-CAIX antibodies. [Figure 80] Figure 1 shows the cytotoxicity of the tandem CAR B7-GGGGS5-G36. [Figure 81-1] 1 shows the amino acid sequences and germline alignments of anti-TIGIT antibodies. [Figure 81-2] See the description of Figure 81-1. [Figure 81-3] See the description of Figure 81-1. [Figure 82] Anti-PD-L1 amino acid sequence is shown. [Figure 83-1] Anti-PD1 nucleic acid and amino acid sequences are shown. [Figure 83-2] See the description of Figure 83-1. [Figure 83-3] See the description of Figure 83-1. [Figure 83-4] See the description of Figure 83-1. [Figure 83-5] See the description of Figure 83-1. DETAILED DESCRIPTION OF THE INVENTION

[0034] Chimeric antigen receptor (CAR) T-cell therapy represents an exciting area of ​​discovery that has already revolutionized the treatment of several blood-borne cancers. For example, in acute lymphoblastic leukemia (ALL), remission rates of 80–90% have been demonstrated, leading to FDA approval. This technology utilizes a patient's own immune cells to fight cancer by engineering them to better recognize specific proteins located on cancer cells. For example, after the modifications are made in the laboratory, the immune cells can be grown extensively in the lab, doubling their cancer-killing potential, and finally infused into the patient, where their increased efficacy and numbers allow them to attack cancer anywhere in the body.

[0035] CAR T cell therapy has been largely elusive in solid tumors such as renal cell carcinoma (RCC), where cancer cells create a tumor microenvironment that shuts off immune infiltration. We have successfully created an advanced mouse model that can provide critical information for this technology. Our approach relies on creating humanized RCC in mice, which can be studied to better understand its immunological fingerprint. By examining how human RCC behaves in mice, our laboratory has been able to identify antigens that can be targeted using CAR T cell therapy in humans and other mammals, such as carbonic anhydrase IX [CAIX] and CD70. To a large extent, these two antigens, CAIX and CD70, are found together only on renal cancer tumor cells, allowing the immune system to attack them locally with minimal impact on healthy tissue. Progress has also been made in counteracting the immunosuppressive effects within the microtumor environment by engineering T cells to produce and / or secrete checkpoint-blocking antibodies. Together, these approaches will enable dramatically improved efficacy within the tumor microenvironment.

[0036] Described herein are, for example, CAR T cells targeting CAIX and CD70, which can be administered to animals while closely monitoring changes in the tumor microenvironment and subsequent antitumor effects. Those skilled in the art will recognize that these approaches can also be combined with currently available technologies that have already positively impacted the clinical setting of RCC, such as the use of immune checkpoint blockade.

[0037] A detailed description of one or more embodiments is provided herein. However, it is understood that the present invention can be embodied in various forms. Therefore, the specific details disclosed herein should not be construed as limiting, but as a basis for the claims and as a representative basis for teaching one skilled in the art how to use the present invention in any suitable manner.

[0038] The singular forms "a," "an," and "the" include plural references unless the context clearly dictates otherwise. The use of the word "a" or "an" when used in conjunction with the word "comprising" in the claims and / or specification can mean "one," but is also consistent with the meanings of "one or more," "at least one," and "one or more than one."

[0039] Whenever the phrases "for example," "such as," "including," and the like are used herein, unless expressly stated otherwise, it is understood that they are accompanied by the phrase "without limitation." Similarly, "one example," "exemplary," and the like are understood to be non-limiting.

[0040] The term "substantially" permits deviations from the descriptor that do not adversely affect the intended purpose. It is understood that a descriptor is modified by the term "substantially" even if the word "substantially" is not expressly recited.

[0041] The terms "comprising" and "including," as well as "having" and "involving" (and similarly, "comprises," "includes," "has," and "involves"), etc., are used interchangeably and have the same meaning. Specifically, each term is defined consistent with the general U.S. patent law definition of "comprising," and therefore shall be construed as having the open term meaning "at least the following," and as not excluding additional features, limitations, embodiments, etc. Thus, for example, "a process involving steps a, b, and c" means that the process includes at least steps a, b, and c. Whenever the terms "a" and "an" are used, they shall be understood to mean "one or more," unless such interpretation is meaningless in the context.

[0042] The term "about" as used herein can refer to approximately, roughly, approximately, or within a range. When the term "about" is used in conjunction with a numerical range, it modifies that range by extending the boundaries above and below the stated numerical values. In general, the term "about" is used herein to modify a numerical value above and below the stated value by a variance of 20 percent (up and down).

[0043] Chimeric antigen receptor (CAR) T-cell therapy Chimeric antigen receptor (CAR) T cell therapy redirects a patient's T cells to kill tumor cells through the exogenous expression of a CAR. For example, referring to Figure 1, a CAR is a transmembrane fusion protein that links the antigen recognition domain of an antibody or fragment to the intracellular signaling domain of a T cell receptor and co-receptor. For example, a chimeric antigen receptor fuses an antigen-specific antibody fragment to a T cell co-stimulatory domain and a CD3 zeta intracellular signaling domain, allowing the redirection of T cells to antigens presented on target cells, such as tumor cells.

[0044] The term "antibody" herein is used in the broadest sense and can refer to immunoglobulin molecules and immunologically active portions of immunoglobulin (Ig) molecules, i.e., molecules that contain an antigen-binding site that specifically binds (immunoreacts with) an antigen. "Specifically binds" or "immunoreacts with" can refer to antibodies that react with one or more antigenic determinants of a desired antigen and not with other polypeptides. Antibodies of the present invention include, but are not limited to, polyclonal, monoclonal, humanized, fully human, bispecific, multispecific, chimeric, dAb (domain antibodies), single-chain antibodies, Fab, Fab', and F(ab')2 fragments, scFv, diabodies, minibodies, scFv-Fc fusions, and Fab expression libraries. Unless expressly stated to the contrary, reference herein to "antibody" or "antibodies" encompasses, for example, any (or all) of these molecules, so long as they exhibit the desired antigen-binding activity.

[0045] Single-chain Fv ("scFv") polypeptide molecules are covalently linked VH::VL heterodimers and can be expressed from gene fusions containing VH- and VL-encoding genes connected by a peptide-encoding linker. (See Huston et al. (1988) Proc Nat Acad Sci USA 85(16):5879-5883.) Numerous methods have been described for identifying chemical structures for converting naturally aggregated but chemically separated light and heavy polypeptide chains derived from antibody V regions into scFv molecules that fold into a three-dimensional structure substantially similar to that of an antigen-binding site. See, e.g., U.S. Patent Nos. 5,091,513, 5,132,405, and 4,946,778.

[0046] Solid tumors present unique challenges for CAR-T therapy. Unlike hematological cancers, tumor-associated target proteins are overexpressed between tumors and healthy tissues, resulting in on-target / off-tumor T cell killing of healthy tissues. Furthermore, immune suppression in the tumor microenvironment (TME) limits tumor-killing CAR-T cell activation. Disclosed aspects address these issues. For example, embodiments include (a) T cells comprising a bispecific CAR that targets two antigens on cancer cells and reduces on-target / off-tumor T cell killing. See Figure 12, for example, for example, B7-GGGGS-G36 CAR T cells had greater killing activity against target renal cancer cells (CAIX+CD70+) than non-target cells (CAIX+CD70-, CAIX-CD70+, CAIX-CD70-). In embodiments, the bispecific CAR can also (b) secrete checkpoint blockade antibodies that remove suppression in the tumor microenvironment. Other embodiments include fine-tuned CARs that recognize only high-density antigens on tumor cells, but not low-density antigens on normal cells. Without wishing to be bound by theory, this can be achieved, for example, by reducing the affinity of one or more antibodies associated with the CAR.

[0047] For example, Figure 1 provides a schematic diagram of a bispecific CAR that targets two antigens, such as CAIX and CD70. A bispecific CAR can refer to a CAR that has binding specificities for at least two different antigens. For example, a bispecific CAR can include a monoclonal antibody, such as a human or humanized antibody, or a fragment thereof. In this case, one of the binding specificities is CAIX and / or CD70. The second binding target is any other antigen, preferably a cell surface protein or receptor or receptor subunit. For example, one of the binding specificities is for CAIX, and the second binding specificity is for CD70.

[0048] As reported (see, e.g., J Clin Oncol 24(2006)20-22; Molecular Therapy 21(2013)4), the initial clinical trial of first-generation anti-CAIX G250 CAR-T cells in patients with renal cell carcinoma (RCC) failed to address the on-target, off-tumor side effects. Two of the first three patients developed hepatitis due to CAIX expression in the bile duct. The introduction of a secondary antibody, such as a secondary scFv, may enhance patient safety by reducing or eliminating on-target / off-tumor effects with dual-targeting CAR T. For example, Figure 24 shows that CAIX and CD70 are upregulated and co-expressed in ccRCC. Meanwhile, CAIX is expressed in the bile duct (mainly in the cytoplasm), whereas CD70 is not expressed in the bile duct. See, e.g., British Journal of Cancer 103(2010)676-684.

[0049] Bispecific antibodies can be prepared as full-length antibodies or antibody fragments (e.g., F(ab')2 bispecific antibodies). Methods for making bispecific antibodies are known in the art. See, for example, U.S. Patent No. 8,329,178, which is incorporated herein by reference in its entirety.

[0050] Antibody molecules obtained from humans belong to one of the classes IgG, IgM, IgA, IgE, and IgD, which differ from each other in the nature of the heavy chains present in the molecule. Certain classes also have subclasses, such as IgG1, IgG2, and others. Furthermore, in humans, the light chains can be kappa or lambda chains.

[0051] The term "antigen-binding site" or "binding portion" can refer to the portion of an immunoglobulin molecule involved in antigen binding. The antigen-binding site is formed by amino acid residues in the N-terminal variable ("V") regions of the heavy ("H") and light ("L") chains. Three highly divergent stretches within the V regions of the heavy and light chains, called "hypervariable regions," are interposed between more conserved adjacent stretches known as "framework regions" or "FRs." Thus, the term "FR" refers to the amino acid sequences naturally found between and adjacent to the hypervariable regions of immunoglobulins. In antibody molecules, the three hypervariable regions of the light chain and the three hypervariable regions of the heavy chain are positioned relative to each other in three-dimensional space to form an antigen-binding surface. The antigen-binding surface is complementary to the three-dimensional surface of a bound antigen, and the three hypervariable regions of each of the heavy and light chains are referred to as "complementarity-determining regions" or "CDRs." See, for example, Tables 1 and 3, which provide the CDRs of anti-CAIX and anti-CD70 antibodies.

[0052] An emerging mechanism associated with tumor progression is the immune checkpoint pathway, which involves cellular interactions that prevent excessive T cell activation under normal conditions and allow T cell function in a self-limiting manner. As an evasion mechanism, many tumors can stimulate the expression of immune checkpoint molecules, resulting in an anergic phenotype of T cells that cannot suppress tumor progression. For example, emerging clinical data highlight the importance of one inhibitory ligand-receptor pair, programmed death ligand 1 (PD-L1, B7-H1, and CD274) and programmed death receptor 1 (PD-1, CD279), as an immune checkpoint that prevents cytotoxic T lymphocytes from killing cancer cells. The PD1 receptor is expressed on many cell types, including T cells, B cells, natural killer cells (NK), and host tissues. PD-L1-expressing tumors and antigen-presenting cells (APCs) can block T cell receptor (TCR) signaling of cytotoxic T lymphocytes through binding to the receptor PD-1, reducing cytokine production and T cell proliferation. Overexpression of PD-L1 is found in many tumor types and mediates immunosuppressive functions through interactions with other proteins, including CD80 (B7.1), and blocks their ability to activate T cells via binding to CD28.

[0053] Genetic engineering of human lymphocytes to express tumor-targeting chimeric antigen receptors (CARs) can generate anti-tumor effector cells that circumvent tumor immune evasion mechanisms due to abnormalities in protein antigen processing and presentation. Furthermore, these transgenic receptors can target tumor-associated antigens that are not protein-derived. Certain embodiments of the present disclosure include lymphocytes engineered to contain at least a CAR (CART), and in certain embodiments of the present invention, a single CAR targets two or more antigens (e.g., bispecific CARs). In some embodiments, the cells contain split CARs, such as anti-CD70 and anti-CAIX scFvs, expressed on the cell surface with distinct costimulatory domains. Furthermore, some embodiments include fine-tuned CARs. In some embodiments, the CART is further engineered to express and secrete one or more polypeptides, such as antibodies or cytokines, such as IL-12, IL-15, or IL-18. Such CARTs are referred to herein as armed CARTs or CAR factories. Armed CART allows for the simultaneous secretion of polypeptides locally at the target site (i.e., tumor site).

[0054] For example, referring to Figure 56, a split CAR comprises two or more CARs on the surface of a cell, such as a T cell or NK cell. The CARs can be specific for two or more antigens, such as CD70 and CAIX. Figure 77 provides an example of a nucleic acid construct encoding a split CAR. In this example, the first CAR is specific for CAIX, and the second CAR is specific for CD70. As described herein, the CARs can be in any desired orientation. For example, the first CAR can be specific for CD70, and the second CAR can be specific for CAIX. As shown in the example, the first and second CARs can be expressed from a single nucleic acid construct. In such an example, a nucleic acid encoding a cleavable linker can be positioned between the nucleic acid encoding the first CAR and the nucleic acid encoding the second CAR. In other embodiments, the two CARs can be expressed in the same cell but from two separate nucleic acid constructs.

[0055] Chimeric antigen receptors (CARs), engineered TCRs containing single-chain variable antibody fragments (scFvs) previously selected for their high affinity for specific tumor-associated antigens, represent a powerful new approach to cancer treatment. The scFvs displayed on the CARs are linked to intracellular signaling blocks, including CD3ζ, to induce T cell activation before antigen binding. This structure, characteristic of first-generation CARs, has been improved in second-generation CARs, which link the signaling costimulatory endodomains of CD28, 4-1BB, or OX40 to CD3, or in third-generation CARs, which link these two elements in tandem to CD3ζ. These endodomains are required for full T cell activation during TCR recognition by antigen-presenting cells (APCs) and improve cytokine production and proliferation of CAR-T cells. Due to the difficulty of finding specific tumor-associated antigens, inefficient T cell homing to tumor sites, poor T cell persistence in the body, and the immunosuppressive microenvironment of solid tumors, the efficacy of CAR-T cells in the treatment of solid tumors has been modest.

[0056] In certain cases, lymphocytes may comprise receptors that are chimeric, non-natural, and at least partially engineered by human hands. In certain cases, engineered chimeric antigen receptors (CARs) have one, two, three, four, or more components, and in some embodiments, one or more components facilitate targeting or binding of lymphocytes to one or more tumor antigen-containing cancer cells.

[0057] A CAR according to the present invention comprises at least one transmembrane polypeptide comprising at least one extracellular ligand-binding domain and one transmembrane polypeptide comprising at least one intracellular signaling domain, such that the polypeptides assemble together to form a chimeric antigen receptor. Exemplary CARS useful in embodiments of the present disclosure include, for example, those disclosed in PCT / US2006 / 046350, PCT / US2015 / 067178, PCT / US2015 / 067225, and PCT / US2019 / 022272, each of which is incorporated herein by reference in its entirety.

[0058] As used herein, the term "extracellular ligand-binding domain" can refer to an oligo- or polypeptide capable of binding to a ligand. The domain can interact with a cell surface molecule. For example, the extracellular ligand-binding domain can be selected to recognize a ligand that acts as a cell surface marker on target cells associated with a particular disease state.

[0059] In particular, the extracellular ligand-binding domain can include an antigen-binding domain or antigen-recognition domain derived from an antibody directed against a target antigen. The antigen-binding domain or antigen-recognition domain can be an antibody fragment. An "antibody fragment" can be a molecule other than an intact antibody that contains a portion of the intact antibody that binds to the antigen to which the intact antibody binds. Examples of antibody fragments include, but are not limited to, Fv, Fab, Fab', Fab'-SH, F(ab')2, diabodies, linear antibodies, single-chain antibody molecules (e.g., scFv), and multispecific antibodies formed from antibody fragments. For example, with reference to Figure 1, one embodiment includes a CAR with two scFvs as antigen-recognition domains. For example, with reference to Figure 14, one embodiment includes a CAR with one scFv as the antigen-recognition domain.

[0060] The antigen recognition domain can be directed to any antigen target of interest. In embodiments, the antigen target of interest is on the surface of a cell, such as the surface of a cancer cell. Non-limiting examples of antigen targets include CAIX and / or CD70.

[0061] In some embodiments, the CAR is specific for CAIX and / or CD70.

[0062] In embodiments, the extracellular ligand-binding domain is a single-chain antibody fragment (scFv) comprising the light chain (VL) and heavy chain (VH) variable fragments of a target antigen-specific monoclonal antibody joined by a flexible linker. Those skilled in the art will recognize that embodiments can include different linkers typically known in the art. See, for example, Chen, et al., "Fusion protein linkers: property, design, and functionality," Advanced drug delivery reviews 65.10 (2013):1357-1369, incorporated herein by reference in its entirety. For example, using different linkers allows for fine-tuning of dual-targeting CAR constructs. The length of the linker can vary depending on the antibodies of the dual-targeting CAR construct, their angle of approach to the target epitope, and the target's topography on the tumor cell membrane. For example, referring to Figure 2, flexible linkers can include GGGSI, GGGGS3, GGGGS5, or IgG1 hinges. In some embodiments, the number of Gs in the linker can be 2, 3, 4, 5, 6, or 7 in combination with any of S1, S2, S3, S4, S5, or S6. For example, the scFv antibody is specific for CAIX and / or CD70. The orientation of the scFv relative to the linker can vary, for example, as shown in Figures 10 and 50. In one nucleic acid construct shown in Figure 50, the anti-CAIX scFv can be in the first cassette (i.e., before the linker) and the anti-CD70 cassette can be in the second cassette (i.e., after the linker). Alternatively, the anti-CAIX scFv can be in the second cassette and the anti-CD70 scFv can be in the first cassette. For example, with reference to Figure 51, the anti-CAIX scFv G36 can be in the first cassette and the anti-CD70 B7 can be in the second cassette. Alternatively, anti-CAIX scFv G36 can be in the second cassette and anti-CD70 B7 can be in the first cassette. As described herein, linkers of various lengths and flexibilities can be utilized. As shown, different orientations of the two scFvs can affect binding.For example, G36 has higher avidity than the one engineered as the second cassette, whereas the anti-CD70 scFv B7 has no significant preference, as shown in Figure 11.

[0063] Examples of antibodies useful for constructing CARs according to the present disclosure include those detailed herein in Tables 1, 2, 3, or 4. See also, e.g., WO / 2007 / 065027 and WO / 2016 / 100985, the contents of which are incorporated herein by reference in their entireties.

[0064] Antigen recognition domains useful for constructing CAR-Ts, e.g., scFVs directed against CAIX and / or CD70, can be synthesized, engineered, and / or produced using nucleic acids (e.g., DNA). The DNA encoding the antigen recognition domain can be cloned in frame with DNA encoding necessary CAR-T elements, such as, for example, but not limited to, the CD8 hinge region, transmembrane domain, costimulatory domain, and the like, of a molecule of immunological interest, such as, for example, but not limited to, CD28 and 41BB and CD3-zeta intracellular signaling domains. See, e.g., Figure 2.

[0065] By way of non-limiting example, binding domains other than scFvs, such as camelid single domain antibody fragments or receptor ligands, antibody binding domains, antibody hypervariable loops, or CDRs, can also be used for predefined targeting of lymphocytes.

[0066] In one embodiment, the transmembrane domain further comprises a stalk region between the extracellular ligand-binding domain and the transmembrane domain. As used herein, the term "stalk region" can refer to any oligo- or polypeptide that functions to link the transmembrane domain to the extracellular ligand-binding domain. In particular, the stalk region is used to provide more flexibility and accessibility to the extracellular ligand-binding domain. The stalk region can comprise up to 300 amino acids, e.g., 10-100 amino acids, e.g., 25-50 amino acids. The stalk region can be derived from all or part of a naturally occurring molecule, such as all or part of the extracellular region of CD8, CD4, or CD28, or all or part of an antibody constant region. Alternatively, the stalk region can be a synthetic sequence corresponding to a naturally occurring stalk sequence, or can be a completely synthetic stalk sequence. In one embodiment, the stalk region is a portion of the human CD8 alpha chain.

[0067] The signal transduction domain or intracellular signaling domain of the CAR of the present invention is involved in intracellular signal transduction after the extracellular ligand binding domain binds to a target, resulting in immune cell activation and immune response. In other words, the signal transduction domain is involved in activating at least one of the normal effector functions of the immune cell in which the CAR is expressed. For example, the effector function of a T cell can be cytolytic activity or helper activity, including cytokine secretion. Therefore, the term "signal transduction domain" can refer to the portion of a protein that converts effector signal function signals and instructs the cell to perform a specific function.

[0068] The signal transduction domain can comprise two distinct classes of cytoplasmic signaling sequences: those that initiate antigen-dependent primary activation and those that act antigen-independently to provide secondary or costimulatory signals. Primary cytoplasmic signaling sequences can comprise signaling motifs known as ITAMs (immunoreceptor tyrosine-based activation motifs). ITAMs are well-defined signaling motifs found in the cytoplasmic tails of various receptors that function as binding sites for the syk / zap70 class of tyrosine kinases. Examples of ITAMs used in the present invention can include, but are not limited to, those derived from TCR zeta, FcR gamma, FcR beta, FcR epsilon, CD3 gamma, CD3 delta, CD3 epsilon, CD5, CD22, CD79a, CD79b, and CD66d. In another embodiment, the signal transduction domain of the CAR can comprise the CD3 zeta signaling domain or the cytoplasmic domain of the Fc epsilon RI beta or gamma chain. In another embodiment, signaling is provided by CD3 zeta, with costimulation provided by CD28 and a tumor necrosis factor receptor (TNFr), such as, for example, 4-1BB or OX40.

[0069] In one embodiment, the intracellular signaling domain of the CAR of the present invention comprises a costimulatory signal molecule. In some embodiments, the intracellular signaling domain comprises two, three, four, or more costimulatory molecules in tandem. Costimulatory molecules are cell surface molecules other than antigen receptors or their ligands that are required for an efficient immune response.

[0070] A "costimulatory ligand" can refer to a molecule on an antigen-presenting cell that specifically binds to a cognate costimulatory molecule on a T cell, thereby providing a signal that mediates T cell responses, including, but not limited to, proliferation, activation, differentiation, etc., in addition to the primary signal provided by the binding of, for example, a peptide-loaded MHC molecule to the TCR / CD3 complex. Costimulatory ligands include CD7, B7-1 (CD80), B7-2 (CD86), PD-L1, PD-L2, 4-1BBL, OX40L, inducible costimulatory ligand (ICOS-L), intercellular adhesion molecules (ICAMs, CD30L, CD40, CD70, CD83, HLA-G, MICA, M1CB, HVEM, lymphotoxin beta receptor, 3 / TR6, ILT3, ILT4, agonists or antibodies that bind ligands to Toll ligand receptors, and the like. In particular, costimulatory ligands can include, but are not limited to, ligands that specifically bind to B7-H3. Costimulatory ligands also encompass antibodies that specifically bind to costimulatory molecules present on T cells, such as, but not limited to, ligands that specifically bind to CD27, CD28, 4-IBB, OX40, CD30, CD40, PD-1, ICOS, lymphocyte function-associated antigen-1 (LFA-1), CD2, CD7, LTGHT, NKG2C, B7-H3, and CD83.

[0071] A "costimulatory molecule" can refer to a cognate binding partner on a T cell that specifically binds to a costimulatory ligand, thereby mediating a costimulatory response by the cell, such as, but not limited to, proliferation. Costimulatory molecules include, but are not limited to, MHC class 1 molecules, BTLA, and Toll ligand receptors. Examples of costimulatory molecules include CD27, CD28, CD8, 4-1BB (CD137), OX40, CD30, CD40, PD-1, ICOS, lymphocyte function-associated antigen-1 (LFA-1), CD2, CD7, LIGHT, NKG2C, B7-H3, and ligands that specifically bind to CD83, and the like.

[0072] In another specific embodiment, the signal transduction domain is a TNFR-associated factor 2 (TRAF2) binding motif, which is the intracellular tail of the costimulatory TNFR family. The cytoplasmic tail of the costimulatory TNFR family members contains a TRAF2 binding motif consisting of a major conserved motif (P / S / A)X(Q / E)E) or a minor motif (PXQXXD), where X is any amino acid. TRAF proteins are recruited to the intracellular tails of many TNFRs in response to receptor trimerization.

[0073] A chimeric antigen receptor fuses an antigen recognition domain to a signaling domain (also called a stimulatory domain) that regulates (i.e., stimulates) cell signaling. Non-limiting examples of such stimulatory domains include those of CD28, 41BB, and / or CD3-zeta intracellular signaling domains. See, e.g., Figure 2.

[0074] The distinctive features of suitable transmembrane polypeptides include their ability to be expressed on the surface of immune cells, particularly lymphocytes or natural killer (NK) cells, and to interact with each other to induce a cellular response of immune cells against a specific target cell. The different transmembrane polypeptides of the CAR of the present invention, including extracellular ligand binding domains and / or signal transduction domains, interact together to participate in signal transduction after binding to the target ligand and induce an immune response. The transmembrane domain can be derived from either natural or synthetic origin. The transmembrane domain can be derived from any membrane-bound or transmembrane protein.

[0075] The term "portion" as used herein can refer to any subset of a molecule, i.e., a shorter peptide. Alternatively, functional variants of a polypeptide's amino acid sequence can be prepared by mutation of the DNA encoding the polypeptide. Such variants or functional variants include, for example, deletions from, or insertions or substitutions of, residues within the amino acid sequence. Any combination of deletions, insertions, and substitutions can be made to arrive at the final construct, provided that the final construct has the desired activity, particularly specific anti-target cellular immune activity. The functionality of the CAR of the present invention in host cells can be detected by assays suitable for demonstrating the signal transduction ability of the CAR upon binding of a specific target. Such assays are available to those skilled in the art. For example, these assays allow for the detection of signal transduction pathways triggered upon target binding, such as assays involving measurement of increased calcium ion release, intracellular tyrosine phosphorylation, inositol phosphate turnover, or the resulting production of interleukin (IL) 2, interferon gamma, GM-CSF, IL-3, and IL-4.

[0076] Carbonic anhydrase IX (CAIX) Several mAbs that react with surface antigens on RCC have been identified. These include mAbs that recognize differentiated and overexpressed antigens, as well as mAbs that identify RCC-associated antigens not expressed in normal kidneys (Michael, 2003; Yang, 2003). The gene for CAIX, also known as G250 and MN, is located on chromosome 9p12-13 and encodes a transmembrane protein that binds zinc and has CA activity (Zavada, 1997; Grabmaier, 2000). In HeLa cells and RCC cell lines derived from human cervical cancer, CAIX / G250 / MN / is found at the plasma membrane and as a nuclear protein with apparent molecular weights of 58 and 54 kDa. It is N-glycosylated, and under nonreducing conditions, it forms oligomers (Pastorekova, 1992). Sequence analysis of the predicted CAIX protein reveals that it contains a signal peptide (aa1-37), an extracellular (EC) portion (aa38-414), a 20-amino acid hydrophobic transmembrane region (aa415-434), and a small 25-amino acid C-terminal extracellular portion (aa435-459). The amino acid sequences of human and mouse CAIX are shown in Figure 59. The extracellular portion consists of two distinct domains. The region between the signal peptide and the CA domain (aa53-111) shows significant homology (38% identity) with the keratan sulfate attachment domain of aggrecan, a large aggregating proteoglycan in humans (Doege, 1991). The PG-like domain of CAIX contains seven repeats of a hexapeptide motif with the consensus EEDLPE (sequence number [ ]). The carbonic anhydrase domain (aa135-391) is located near the plasma membrane. CAIX antigen appears during malignant transformation and stains positive in approximately 95% of clear cell RCC specimens, as well as in most renal cell metastases.

[0077] Epitopes expressed on the cell surface of tumor cells, unlike intracellular antigens, are excellent targets for humoral anticancer therapy because they are accessible to circulating antibodies in vivo. Human monoclonal antibodies (mAbs) have become a well-tolerated treatment option for an increasing number of cancers. The concept of selective tumor targeting by antibodies is based on the avid interaction between antibodies and antigens expressed on malignant cells but not on normal tissues. Many mechanisms have been proposed for the ability of antibodies against tumors to mediate their effects in vivo. For example, engagement of the antibody Fc domain with effector cell FcγRs leads to antibody-dependent cell-mediated cytotoxicity (ADCC). Some (antagonistic or inhibitory) antibodies can block signaling on tumor cells and, in this way, may act synergistically with immune effector responses by making tumor cells more susceptible to apoptotic or lytic cell death triggered by immune effector cells (Baselga, 1998). Another way antibodies can be utilized is through the construction and functional expression of chimeric immune receptors or "T-bodies" on T lymphocytes, otherwise known as "designer T cells." The antigen-binding domain of the chimeric receptor can consist of an antigen-binding domain, e.g., a single-chain fragment (scFv), while the intracellular signaling domain is derived from the cell membrane portion of the membrane-bound receptor, which can induce cell activation (Maher, 2002; Pinthus, 2003). T lymphocytes transplanted with chimeric receptors possess the combined advantages of MHC-independent and antibody-based antigen binding, with efficient T cell activation upon specific binding to the receptor ligand. This activation leads to the production and secretion of cytokines such as IL-2, interferon, GM-CSF, and TNF-α. Antigen-specific lysis of tumor cells has been reported both in vitro and in vivo. T lymphocytes can be permanently transplanted with antigen-specific chimeric receptors by retroviral transduction of vector constructs encoding the receptor molecule of choice (reviewed by Riviere, 2004).

[0078] Embodiments of the present invention can include isolated human monoclonal antibodies or fragments thereof that immunospecifically bind to carbonic anhydrase IX (G250) protein. Such antibodies can reduce the carbonic anhydrase activity of the protein. For example, such anti-CAIX antibodies can include those described in WO 2007 / 065027 and US 8,466,263, each of which is incorporated herein by reference in its entirety. See, for example, Figure 58, which provides a multiple alignment of the amino acid sequences of anti-carbonic anhydrase IX (G250) scFv clones. For example, embodiments of the present invention include single-chain antibodies, such as scFv G6, G9, G10, G17, G21, G27, G28, G36, G37, G39, G40, G45, G57, G62, G98, G104, G119, or G125, as well as any other scFv identified according to the methods disclosed herein.

[0079] For example, the amino acids of the CDRs of anti-CAIX antibodies are shown in Table 1.

[0080] (Table 1) TIFF2025011142000002.tif211154TIFF2025011142000003.tif63154

[0081] For example, the amino acid sequences of the VH and VL regions of anti-CAIX antibodies are shown in Table 2.

[0082] (Table 2) TIFF2025011142000004.tif139154TIFF2025011142000005.tif231154TIFF2025011142000006.tif231154TIFF2025011142000007.tif105154

[0083] Embodiments can also include consensus sequences of any of the amino acid sequences described herein, for example, if four or more clones have the same amino acid at a given position, that position in the consensus is designated by that amino acid.

[0084] CD70 CD70 is found on the surface of tumor cells of the kidney (e.g., clear cell carcinoma and papillary carcinoma), pancreas, larynx or pharynx, melanoma, ovary, lung adenocarcinoma, colon, breast, and brain. See, e.g., British Journal of Cancer 103 (2010) 676-684.

[0085] Embodiments of the present invention can include isolated human monoclonal antibodies or fragments thereof that immunospecifically bind to CD70. For example, embodiments of the present invention include single chain antibodies such as scFv A20, B2, B3, B5, B7, B8, or B9, as well as any other scFv identified according to the methods disclosed herein.

[0086] For example, the amino acids of the CDRs of anti-CD70 antibodies are shown in Table 3.

[0087] (Table 3) TIFF2025011142000008.tif71154

[0088] For example, the amino acid sequences of the VH and VL regions of anti-cd70 antibodies are shown in Table 4.

[0089] (Table 4) TIFF2025011142000009.tif178154

[0090] Embodiments can also include consensus sequences of any of the amino acid sequences described herein, for example, if four or more clones have the same amino acid at a given position, that position in the consensus is designated by that amino acid.

[0091] cell Embodiments of the present disclosure include cells expressing a CAR (i.e., a CART). The cells can be of any type, including immune cells capable of expressing a CAR for cancer therapy, or cells such as bacterial cells harboring an expression vector encoding a CAR. As used herein, the terms "cell," "cell line," and "cell culture" can be used interchangeably. All of these terms also include their progeny, that is, any subsequent generations. All progeny may not be identical, for example, due to deliberate or inadvertent mutation. In the context of expressing a heterologous nucleic acid sequence, a "host cell" can refer to a eukaryotic cell capable of replicating the vector and / or expressing the heterologous gene encoded by the vector. Host cells can be and have been used as recipients of vectors. Host cells may be "transfected" or "transformed," which refers to the process by which exogenous nucleic acid is transferred or introduced into a host cell. Transformed cells include the primary subject cell and its progeny. As used herein, the terms "engineered" and "recombinant" or host cells can refer to cells into which an exogenous nucleic acid sequence, such as a vector, has been introduced. Recombinant cells are therefore distinguishable from naturally occurring cells that do not contain recombinantly introduced nucleic acids. In embodiments of the invention, the host cells are T cells, including cytotoxic T cells (also known as TCs, cytotoxic T lymphocytes, CTLs, T killer cells, cytolytic T cells, CD8+ T cells, or killer T cells); NK cells and NKT cells are also encompassed by the present disclosure.

[0092] Some vectors can use control sequences that allow them to be replicated and / or expressed in both prokaryotic and eukaryotic cells. Those skilled in the art will further understand the conditions for incubating all of the above host cells to maintain them and allow vector replication. Techniques and conditions that allow large-scale production of vectors and production of nucleic acids encoded by vectors and their cognate polypeptides, proteins, or peptides are also understood and known.

[0093] The cells may be autologous, syngeneic, allogeneic, or in some cases xenogeneic.

[0094] There are many situations in which it may be desirable to be able to kill modified CTLs, such as when it is desired to terminate treatment, when cells become tumorigenic, in studies where the absence of cells after their presence is of interest, or for other reasons. To this end, one can provide for the expression of specific gene products that can kill modified cells under controlled conditions, such as inducible suicide genes.

[0095] Armed CART The present invention further includes CARTs modified to secrete one or more polypeptides. Such CARTs may be referred to as CART factories, CAR T cell factories, or armed CARTs. The polypeptides may be, for example, antibodies or fragments thereof described herein. For example, the polypeptides may be antibodies or cytokines. In embodiments, the antibodies are specific for TIGIT, CAIX, GITR, PD-L1, PD-L2, PD-1, CCR4, CTLA-4, VISTA, or CD70. For example, the CAR T cell factory may secrete PD-L1 mAb locally at tumor sites to restore effective anti-cancer immunity and / or reverse T cell exhaustion. In embodiments, the armed CARTs secrete IL-12, IL-15, or IL-18.

[0096] For example, a second expression construct, which may be on the same DNA vector as that encoding the CAR (e.g., antigen recognition domain) or on a second separate vector, can be used to encode a minibody (scFv-Fc) or antibody or fragment thereof directed against a single or multiple antigens of interest and can be cloned after an internal ribosome entry site (IRES). Referring to the figure, the second expression cassette contains either a fluorescent molecule or an immunomodulatory minibody.

[0097] Armed CARTs have the advantage of simultaneously secreting polypeptides at target sites, such as tumor sites. For example, armed CARTs can secrete anti-TIGIT antibodies or fragments thereof. TIGIT is a T cell co-inhibitory receptor (TCR) that limits antitumor and other T cell-dependent chronic immune responses, such as CD8+ T cell-dependent immune responses. TIGIT is expressed on a subset of activated T cells and natural killer (NK) cells. For example, TIGIT is highly expressed on tumor-infiltrating T cells. In cancer models, antibody blockade of TIGIT contributed to improved CD8+ T cell effector function and tumor clearance.

[0098] In embodiments, the armed CART anti-TIGIT antibody comprises one or more of the anti-TIGIT antibody clones (or fragments thereof, e.g., FR1, FR2, FR3, FR4, CDR1, CDR2, CDR3, or any combination of framework and / or CDR regions therein) described in Figure 81.

[0099] For example, an anti-TIGIT antibody can include a CDR1 of a VH region having the sequence GYTF....TSYG (SEQ ID NO: [ ]), a CDR2 of a VH region having the sequence ISAY..NGNT (SEQ ID NO: [ ]), a CDR3 of a VH region having the sequence ARDPGLWFGLTHDYYFDY (SEQ ID NO: [ ]), a CDR1 of a VL region having the sequence SSNI....GSNT (SEQ ID NO: [ ]), a CDR2 of a VL region having the sequence RN.......N (SEQ ID NO: [ ]), and a CDR3 of a VL region having the sequence AAWDDSRSGPV (SEQ ID NO: [ ]).

[0100] For example, an anti-TIGIT antibody can include a CDR1 of a VH region having the sequence GFTF....SDYS (SEQ ID NO: [ ]), a CDR2 of a VH region having the sequence INSD..GSRT (SEQ ID NO: [ ]), a CDR3 of a VH region having the sequence ARGPGFFGFDI (SEQ ID NO: [ ]), a CDR1 of a VL region having the sequence RSNI....GRNS (SEQ ID NO: [ ]), a CDR2 of a VL region having the sequence SN.......N (SEQ ID NO: [ ]), and a CDR3 of a VL region having the sequence AAWDARLTGPL (SEQ ID NO: [ ]).

[0101] For example, an anti-TIGIT antibody can include a CDR1 of a VH region having the sequence GYSF....TNYW (SEQ ID NO: [ ]), a CDR2 of a VH region having the sequence INPV..NSRT (SEQ ID NO: [ ]), a CDR3 of a VH region having the sequence ARYYYYAMEV (SEQ ID NO: [ ]), a CDR1 of a VL region having the sequence SSNI....GSNT (SEQ ID NO: [ ]), a CDR2 of a VL region having the sequence RN.......N (SEQ ID NO: [ ]), and a CDR3 of a VL region having the sequence EAWDDSLNGPV (SEQ ID NO: [ ]).

[0102] For example, an anti-TIGIT antibody can comprise a CDR1 of a VH region having the sequence GYTF....TNYG (SEQ ID NO: [ ]), a CDR2 of a VH region having the sequence VDNN..NGNI (SEQ ID NO: [ ]), a CDR3 of a VH region having the sequence ARGLFSSRWYLWFDP (SEQ ID NO: [ ]), a CDR1 of a VL region having the sequence SSDVG...GYNY (SEQ ID NO: [ ]), a CDR2 of a VL region having the sequence EV.......T (SEQ ID NO: [ ]), and a CDR3 of a VL region having the sequence SSYTRSSTSYVV (SEQ ID NO: [ ]).

[0103] For example, an anti-TIGIT antibody can include a CDR1 of a VH region having the sequence GGTF....SSYA (SEQ ID NO: [ ]), a CDR2 of a VH region having the sequence ILPM..FGST (SEQ ID NO: [ ]), a CDR3 of a VH region having the sequence ARGRDIVAPSNSGFDV (SEQ ID NO: [ ]), a CDR1 of a VL region having the sequence SNNV....GNQG (SEQ ID NO: [ ]), a CDR2 of a VL region having the sequence RN.......D (SEQ ID NO: [ ]), and a CDR3 of a VL region having the sequence SAYDRSLNAWV (SEQ ID NO: [ ]).

[0104] In other embodiments, the armed CART can secrete an anti-PDL1 antibody or a fragment thereof. For example, the armed CART can secrete an anti-PDL1 antibody or a fragment thereof disclosed in Provisional Patent Application No. 62 / 624,455, the entire contents of which are incorporated herein by reference. Exemplary anti-PDL1 antibodies include antibodies having a VH amino acid sequence having SEQ ID NO: [ ] and / or a VL amino acid sequence having SEQ ID NO: [ ]. See, e.g., Figures 78 and 81.

[0105] In embodiments, the armed CART anti-PDL1 antibody comprises one or more of the anti-PDL1 antibody clones (or fragments thereof, e.g., FR1, FR2, FR3, FR4, CDR1, CDR2, CDR3, or any combination of framework and / or CDR regions therein) described in Figure 78 and / or Figure 81.

[0106] For example, the amino acid sequences of the complementarity determining regions of the heavy and light chains of the PDL-1 antibody are as follows: TIFF2025011142000010.tif129134

[0107] For example, an anti-PDL1 antibody has a heavy chain with three CDRs each comprising the amino acid sequences of SEQ ID NOs: [ ], [ ], and / or [ ], and a light chain with three CDRs each comprising the amino acid sequences of SEQ ID NOs: [ ], [ ], and / or [ ].

[0108] In other embodiments, the armed CART can secrete an anti-PD1 antibody or fragment thereof. Exemplary anti-PD1 antibodies include antibodies having a VH amino acid sequence having SEQ ID NO: [ ] and a VL amino acid sequence having SEQ ID NO: [ ]. See, e.g., Figure 78 and Figure 83.

[0109] In embodiments, the anti-PD1 antibody in the armed CART comprises one or more of the anti-PD1 antibody clones (or fragments thereof, e.g., FR1, FR2, FR3, FR4, CDR1, CDR2, CDR3, or any combination of framework and / or CDR regions therein) set forth in Figure 78 and Figure 83. For example, the anti-PD1 antibody has a heavy chain having three CDRs each comprising the amino acid sequences of SEQ ID NOs: [ ], [ ], and / or [ ], and a light chain having three CDRs each comprising the amino acid sequences of SEQ ID NOs: [ ], [ ], and / or [ ]. See, e.g., Figure 78 and Figure 83.

[0110] In embodiments, the amino acid sequences of the heavy and light chain complementarity determining regions of the PD-1 antibody are shown below. PD-1 antibody heavy chain (V H ) complementarity-determining regions (CDRs) TIFF2025011142000011.tif114134PD-1 antibody light chain (V L ) complementarity-determining regions (CDRs) TIFF2025011142000012.tif114134

[0111] In other embodiments, the armed CART can secrete an anti-CCR4 antibody or fragment thereof, for example, an anti-CCR4 antibody or fragment such as those described in WO2009 / 086514, WO2013 / 166500, PCT / US2015 / 054202, or PCT / US2016 / 026232.

[0112] For example, an armed CART anti-CCR4 protein antibody or fragment thereof can include an antibody having a VH CDR1 region having the amino acid sequence GYTFASYY (SEQ ID NO: [ ]), a VH CDR2 region having the amino acid sequence WINPGNVNTKYNEKFKG (SEQ ID NO: [ ]), a VH CDR3 region having the amino acid sequence STYYRPLDY (SEQ ID NO: [ ]), and / or a VL CDR1 region having the amino acid sequence KSSQSILYSSNQKNYLA (SEQ ID NO: [ ]), a VL CDR2 region having the amino acid sequence WASTRES (SEQ ID NO: [ ]), and / or a VL CDR3 region having the amino acid sequence HQYLSSYT (SEQ ID NO: [ ]).

[0113] For example, the armed CAR anti-CCR4 protein antibody or fragment thereof may have the VH amino acid sequence TIFF2025011142000013.tif22148 (SEQ ID NO: [ ]) and / or VL amino acid sequence The antibody may include the antibody having TIFF2025011142000014.tif13148 (SEQ ID NO: [ ]).

[0114] For example, an armed CART anti-CCR4 protein antibody or fragment thereof can include an antibody having:

[0115] VH chain of antibody 1-44 (SEQ ID NO: [ ]) TIFF2025011142000015.tif22149

[0116] VL chain of antibody 1-44 (SEQ ID NO: [ ]) TIFF2025011142000016.tif13148

[0117] VH chain of antibody 1-49 (SEQ ID NO: [ ]) TIFF2025011142000017.tif22150

[0118] VL chain of antibody 1-49 (SEQ ID NO: [ ]) TIFF2025011142000018.tif13145

[0119] VH chain of antibody 2-1 (SEQ ID NO: [ ]) TIFF2025011142000019.tif22149

[0120] VL chain of antibody 2-1 (SEQ ID NO: [ ]) TIFF2025011142000020.tif13148

[0121] VH chain of antibody 2-2 (SEQ ID NO: [ ]) TIFF2025011142000021.tif22148

[0122] VL chain of antibody 2-2 (SEQ ID NO: [ ]) TIFF2025011142000022.tif13148

[0123] VH chain of antibody 2-3 (SEQ ID NO: [ ]) TIFF2025011142000023.tif22149

[0124] VL chain of antibody 2-3 (SEQ ID NO: [ ]) TIFF2025011142000024.tif13148

[0125] The amino acid sequences of the complementarity determining regions of the heavy and light chains of the anti-CCR4 antibodies are shown in the table below. TIFF2025011142000025.tif135133

[0126] An armed CART can be constructed by including a nucleic acid encoding a polypeptide of interest after an intracellular signaling domain. In embodiments, an internal ribosome entry site (IRES) is located between the intracellular signaling domain and the polypeptide of interest. Those skilled in the art will appreciate that multiple IRES sequences can be used in tandem to express more than one polypeptide.

[0127] In one embodiment, the methods and compositions provided herein provide target-specific T cells, such as T cells with specificity for CAIX and / or CD70, capable of secreting polypeptides in the tumor microenvironment to combat T cell exhaustion, for example. For example, myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of early myeloid progenitors, immature granulocytes, macrophages, and dendritic cells at different stages of differentiation that comprise the tumor microenvironment. MDSCs are induced by proinflammatory cytokines and are found in large numbers in infectious and inflammatory conditions. Without wishing to be bound by theory, their presence in the tumor microenvironment suggests a causative role in promoting tumor-associated immunosuppression. Human MDSCs express Siglec-3 / CD33 (GENBANK accession numbers NM_001772.4 (nucleotide sequence) and NP_001763.3 (amino acid sequence)) and have heterogeneous expression of CD14 (GENBANK accession numbers NM_000591.4 (nucleotide sequence) and NP_000582.1 (amino acid sequence)) and CD15 (GENBANK accession numbers NM_002033.3 (nucleotide sequence) and NP_002024.1 (amino acid sequence)), indicating the existence of multiple subsets.Other MDSC markers useful for identifying MDSCs include B7-1 / CD8 (NM_001145873.1 (nucleotide sequence), NP_001139345.1 (amino acid sequence)), CCR2 (NM_001123041.2 (nucleotide sequence), NP_001116513.2 (amino acid sequence)), CD1d (NM_001766.3 (nucleotide sequence), NP_001757.1 (amino acid sequence)), CD2 (NM_001328609.2 (nucleotide sequence), NP_001315538.1 (amino acid sequence)), CD31 / PECAM-1 (NM_000442.5 (nucleotide sequence), NP_000433 .4 (amino acid sequence)), CD43 (NM_001030288.3 (nucleotide sequence), NP_001025459.1 (amino acid sequence)), CD44 (NM_000610.4 (nucleotide sequence), NP_000601.3 (amino acid sequence)), gp130 (NM_002184.4 (nucleotide sequence), NP_002175.2 (amino acid sequence)), PD-L1 (NM_014143.4 (nucleotide sequence), NP_054862.1 (amino acid sequence)), and CD162 (NM_001206609.2 (nucleotide sequence), NP_001193538.1 (amino acid sequence)).

[0128] In one embodiment, the methods and compositions provided herein provide target-specific T cells, such as T cells with specificity for CAIX and / or CD70, capable of secreting polypeptides that target MDSCs in the tumor microenvironment. For example, the secreted polypeptides can target one or more MDSC markers (e.g., CD33, CD14, and / or CD15, or other MDSC markers listed herein).

[0129] Transduction of constructs into CTLs The expression vector encoding the CAR can be introduced as one or more DNA molecules or constructs, where at least one marker may be present that allows for selection of host cells that contain the construct(s).

[0130] Constructs can be prepared using conventional methods, and the genes and regulatory regions can be isolated, ligated, cloned into a suitable cloning host, and analyzed by restriction or sequencing or other convenient means. In particular, PCR can be used to isolate individual fragments containing all or part of the functional unit, and one or more mutations can be introduced using primer repair, ligation, in vitro mutagenesis, or other suitable methods. Once the construct(s) are complete and have been demonstrated to have the correct sequence, they can then be introduced into CTLs by any convenient means. Constructs may be packaged and integrated into non-replicating, defective viral genomes, such as adenovirus, adeno-associated virus (AAV), or herpes simplex virus (HSV), or retroviral or lentiviral vectors, for cell infection and transduction. Constructs may also include viral sequences for transfection. Alternatively, constructs may be introduced by fusion, electroporation, biolistic methods, transfection, lipofection, or other methods. Host cells can be grown and expanded in culture before introducing the construct(s), after which the construct(s) can be introduced and treated appropriately to integrate the construct(s). The cells are then expanded and screened for the marker present in the construct. Various markers that have been successfully used include hprt, neomycin resistance, thymidine kinase, hygromycin resistance, etc.

[0131] In some cases, when the construct is integrated into a specific locus, it may have a target site for homologous recombination.For example, the endogenous gene can be knocked out and replaced with the gene encoded by the construct (at the same locus or elsewhere) using materials and methods known in the art for homologous recombination.For homologous recombination, either OMEGA or O-vector can be used.For example, see Thomas and Capecchi, Cell (1987) 51, 503-512; Mansour, et al., Nature (1988) 336, 348-352, and Joyner, et al., Nature (1989) 338, 153-156.

[0132] The construct can be introduced as a single DNA molecule that encodes at least CAR and optionally another gene, or as different DNA molecules that have one or more genes.Other genes include, for example, genes that encode therapeutic molecules or suicide genes.The constructs can be introduced simultaneously or sequentially, and each has the same or different markers.

[0133] Vectors containing useful elements such as bacterial or yeast replication origins, selectable and / or amplifiable markers, promoter / enhancer elements for expression in prokaryotes or eukaryotes, which can be used to prepare construct DNA stocks and perform transfections, are well known in the art and many are commercially available.

[0134] How to use Aspects of the present disclosure are directed to methods of treating a subject suffering from cancer.

[0135] For example, embodiments of the present disclosure are directed to methods of killing cancer cells, such as renal cancer cells. Referring to Figure 12, for example, B7-GGGGS-G36 CAR T cells had more killing activity against target renal cancer cells (CAIX+CD70+) than non-target cells (CAIX+CD70-, CAIX-CD70+, CAIX-CD70-). Furthermore, referring to Figure 57, for example, bispecific split CARs achieved superior killing when compared to mono- or bispecific CARs.

[0136] Aspects of the present disclosure are further directed to methods of halting or reducing the progression of, or promoting the regression of, cancer in a subject.

[0137] Still further, embodiments of the present disclosure are directed to methods of reducing cell proliferation of cancer cells in a subject. See, e.g., Figure 80.

[0138] "Cancer" and "cancerous" can refer to or describe, for example, a physiological condition in mammals characterized by unregulated cell growth. Examples of cancer include, but are not limited to, carcinoma, lymphoma, blastoma, sarcoma, and leukemia. More specific examples of such cancers include squamous cell carcinoma, small cell lung cancer, non-small cell lung cancer, lung adenocarcinoma, lung squamous cell carcinoma, peritoneal cancer, hepatocellular carcinoma, gastrointestinal cancer, pancreatic cancer, glioblastoma, cervical cancer, ovarian cancer, liver cancer, bladder cancer, hepatoma, breast cancer, colon cancer, colorectal cancer, endometrial or uterine cancer, salivary gland cancer, kidney cancer, liver cancer, prostate cancer, vulvar cancer, thyroid cancer, liver cancer, and various types of head and neck cancer. For example, the cancer is renal cell carcinoma, such as ccRCC.

[0139] In cancer, normal cell-cell interactions within tissues are disrupted, and the tumor microenvironment evolves to accommodate the growing tumor. The tumor microenvironment (TME) can refer to the cellular environment in which a tumor resides, including components such as surrounding blood vessels, immune cells, fibroblasts, bone marrow-derived inflammatory cells, lymphocytes, signaling molecules, and the extracellular matrix (ECM). The tumor microenvironment is complex and is strongly influenced by the immune system.

[0140] The present invention provides, inter alia, CAR-T cell therapies (such as those described herein) for renal cell carcinoma. Secretion of monospecific, bispecific, or trispecific minibodies, antibodies, or minibody / antibody fusion proteins by CAR-T cells at the tumor site may provide additional benefit by altering (i.e., modulating) the immunosuppressive tumor microenvironment.

[0141] In embodiments, the methods include administering to a subject suffering from cancer a therapeutically effective amount of the engineered cells described herein, which may depend on the severity and course of the cancer, previous treatments, the subject's health, weight, response to drugs, and the judgment of the treating physician.

[0142] The subject may be suffering from cancer, such as a liquid cancer (i.e., a blood cancer) and / or a solid cancer (i.e., a tumor). The cancer may be benign or malignant, and may be influenced by the immune system.

[0143] The embodiments described herein can modulate the immune system to treat a subject suffering from cancer. "Modulating" can refer to upregulation, induction, stimulation, enhancement, and / or release of inhibition, as well as inhibition, attenuation, and / or downregulation or suppression. In embodiments, the activity of the subject's immune system is modulated, or the microenvironment surrounding cancer cells and / or tumors is modulated, or both. For example, the embodiments described herein can alter the immunosuppressive tumor microenvironment, reduce microenvironment-dependent immunosuppression, and modulate (or enable) the immune system to kill tumor cells.

[0144] One embodiment is directed to the method for treating the subject suffering from renal cell carcinoma.Immunotherapy such as that described herein provides a great treatment option for RCC.For example, one embodiment includes engineering chimeric antigen receptor (CAR) T cells for RCC.

[0145] An "individual" or "subject" can be a mammal. Mammals include, but are not limited to, domestic animals (e.g., cows, sheep, cats, dogs, horses), primates (e.g., humans and non-human primates such as monkeys), rabbits, and rodents (e.g., mice and rats). In certain embodiments, the individual or subject is a human.

[0146] The cells according to the present disclosure can be used to treat cancer in a patient in need thereof, hi another embodiment, the isolated cells according to the present invention can be used in the manufacture of a medicament for the treatment of cancer, autoimmune disorders, viral infections in a patient in need thereof.

[0147] The present disclosure relies on a method for treating a patient in need of treatment for cancer, the method comprising at least one of the following steps: (a) providing chimeric antigen receptor cells according to the present invention, and (b) administering the cells to the patient.

[0148] The treatment can be ameliorative, curative or preventive.It can be part of autoimmune therapy or part of allogeneic immunotherapy treatment.Autologous means that the cells, cell lines or cell populations used in the treatment of patients are derived from the patient or from a human leukocyte antigen (HLA)-matched donor.Allogeneic means that the cells or cell populations used in the treatment of patients are derived from a donor, not from the patient.

[0149] The present invention is particularly suitable for allogeneic immunotherapy, insofar as it allows the transformation of T cells obtained from a donor into non-allo-reactive cells. This can be performed under standard protocols and replicated as many times as necessary. The resulting modified T cells can be pooled and administered to one or several patients, making them available as a "ready-to-use" therapeutic product.

[0150] The cells that can be used in the disclosed method are described in the previous section. The treatment can be used to treat patients diagnosed with cancer. Cancers that can be treated include tumors that are not vascularized or have not yet been substantially vascularized, as well as vascularized tumors. Cancers can include non-solid tumors (e.g., hematological tumors, such as leukemia and lymphoma) or solid tumors. The types of cancers that can be treated with the CAR of the present invention include, but are not limited to, carcinomas, blastomas, and sarcomas, as well as certain leukemias or lymphoid malignancies, benign and malignant tumors, and malignant tumors, such as sarcomas, carcinomas, and melanomas. Adult tumors / cancers and pediatric tumors / cancers are also included.

[0151] It may be a treatment in combination with one or more therapies for cancer selected from the group of antibody therapy, chemotherapy, cytokine therapy, dendritic cell therapy, gene therapy, hormone therapy, laser phototherapy and radiation therapy.

[0152] According to an embodiment of the present invention, the treatment can be administered to a patient undergoing immunosuppressive therapy. The present invention uses cells or cell populations that have been made resistant to at least one immunosuppressant due to inactivation of a gene encoding a receptor for the immunosuppressant. In this aspect, the immunosuppressive therapy should aid in the selection and expansion of T cells according to the present invention in the patient.

[0153] In a further embodiment, the cell compositions of the present invention are administered to a patient in combination with (e.g., before, simultaneously with, or after) bone marrow transplantation, T cell ablative therapy using chemotherapy agents such as fludarabine, external beam radiation therapy (XRT), cyclophosphamide, or antibodies such as OKT3 or CAM PATH. In another embodiment, the cell compositions of the present invention are administered after B cell ablative therapy using an agent that reacts with CD20, e.g., Rituxan. For example, in one embodiment, a subject may undergo standard treatment with high-dose chemotherapy followed by a peripheral blood stem cell transplant. In certain embodiments, after transplantation, the subject receives an infusion of the expanded immune cells of the present invention. In a further embodiment, the expanded cells are administered before or after surgery. The modified cells obtained by any of the methods described herein can be used in certain embodiments of the invention to treat patients in need of treatment for host-versus-graft (HvG) rejection and graft-versus-host disease (GvHD), and therefore within the scope of the invention is a method of treating a patient in need of treatment for host-versus-graft (HvG) rejection and graft-versus-host disease (GvHD), comprising treating the patient by administering to the patient an effective amount of modified cells comprising an inactivated TCR alpha and / or TCR beta gene.

[0154] Cell administration The present disclosure is particularly suitable for allogeneic immunotherapy insofar as it allows the transformation of T cells obtained from a donor into non-allo-reactive cells. This can be performed under standard protocols and can be replicated as many times as necessary. The resulting modified T cells can be pooled and administered to one or several patients, making them available as an "off-the-shelf" therapeutic product.

[0155] Depending on the nature of the cells, cells can be introduced into a host organism, e.g., a mammal, in a variety of ways. In certain embodiments, cells can be introduced at the site of a tumor, while in alternative embodiments, the cells are trained or modified to train into cancer. The number of cells used depends on various circumstances, the purpose of the introduction, the lifespan of the cells, the protocol used, e.g., number of administrations, ability of the cells to proliferate, stability of the recombinant construct, etc. Cells can be applied, for example, as a suspension injected at or near the site of interest. Cells can be in a physiologically acceptable medium.

[0156] In some embodiments, the cells are encapsulated to inhibit immune recognition and are placed at the site of the tumor.

[0157] The cells can be administered as desired. A variety of protocols can be used depending on the desired response, the method of administration, the longevity of the cells, and the number of cells present. The number of administrations will depend, at least in part, on the factors described above.

[0158] Administration of the cells or cell populations of the present invention can be by any convenient method, including aerosol inhalation, injection, ingestion, transfusion, implantation, or transplantation. The compositions described herein can be administered to a patient subcutaneously, intradermally, intratumorally, intranodally, intramuscularly, intravenously, or intralymphatically, or intraperitoneally. In one embodiment, the cell compositions of the present invention are administered by intravenous injection.

[0159] Administration of the cells or cell population can consist of administering 104 to 109 cells per kg of body weight, e.g., 105 to 106 cells / kg of body weight, including all integer values ​​within these ranges. The cells or cell population can be administered in one or more doses. In another embodiment, the effective amount of cells is administered as a single dose. In another embodiment, the effective amount of cells is administered as two or more doses over a period of time. The timing of administration is within the discretion of the attending physician and depends on the clinical condition of the patient. The cells or cell population can be obtained from any source, such as a blood bank or donor. While individual needs vary, determining the optimal range of effective amounts of a given cell type for a particular disease or condition is within the skill of the art. An effective amount refers to an amount that provides a therapeutic or prophylactic benefit. The administered dosage will depend on the age, health, and weight of the recipient, the type of concurrent treatment, if any, the frequency of treatment, and the nature of the desired effect.

[0160] It should be understood that the system is influenced by many variables that may change over time and circumstances, such as cellular response to ligand, expression efficiency and, if appropriate, secretion levels, activity of the expressed product, the specific needs of the patient, the rate of cell loss or loss of cellular activity as a result of expression activity of individual cells, etc. Thus, with respect to individual patients, even if there are pluripotent cells that can be administered to an entire population, each patient will be monitored for appropriate individual dosages, and such practices of monitoring patients are routine in the art.

[0161] Nucleic Acid-Based Expression Systems The CAR of the present disclosure can be expressed from an expression vector. Recombinant techniques for producing such expression vectors are well known in the art.

[0162] The DNA constructs described herein, which may also be referred to as "DNA vectors," can be cloned into vectors used to transduce and generate chimeric antigen receptor T cells that secrete the polypeptides and / or fragments thereof. For example, the DNA constructs can be cloned into lentiviral vectors for the production of lentiviruses used to transduce and generate chimeric antigen receptor T cells that secrete monospecific, bispecific, or trispecific immunomodulatory antibodies / minibodies and / or antibody fusion proteins at tumor sites.

[0163] vector The term "vector" can refer to a carrier nucleic acid molecule into which a nucleic acid sequence can be inserted for introduction into a cell where it can be replicated. The nucleic acid sequence can be "exogenous," meaning that the sequence is foreign to the cell into which the vector is introduced, or that the sequence is homologous to a sequence in the cell, but is located in a location within the host cell nucleic acid where the sequence is not normally found. Vectors include plasmids, cosmids, viruses (bacteriophages, animal viruses, and plant viruses), artificial chromosomes (YACs, etc.). Those skilled in the art will be well equipped to construct vectors using standard recombinant techniques (see, for example, Maniatis et al., 1988 and Ausubel et al., 1994, both of which are incorporated herein by reference).

[0164] The term "expression vector" refers to any type of genetic construct containing a nucleic acid encoding an RNA that can be transcribed. In some cases, the RNA molecule is then translated into a protein, polypeptide, or peptide. In other cases, these sequences are not translated, for example, in the production of antisense molecules or ribozymes. Expression vectors can contain a variety of "control sequences," which refer to nucleic acid sequences necessary for the transcription and translation of an operably linked coding sequence in a particular host cell. In addition to control sequences that govern transcription and translation, vectors and expression vectors can contain nucleic acid sequences that perform other functions as well, as described below.

[0165] Promoters and enhancers A "promoter" is a regulatory sequence, which is a region of a nucleic acid sequence that controls the initiation and rate of transcription. It can contain genetic elements to which regulatory proteins and molecules, such as RNA polymerase and other transcription factors, bind to initiate specific transcription of a nucleic acid sequence. The terms "operably arranged," "operably linked," "under control," and "under transcriptional control" mean that the promoter is in the correct functional location and / or orientation relative to the nucleic acid sequence to control the initiation and / or expression of the transcription of that sequence.

[0166] Promoters contain sequences that function to position the start site for RNA synthesis. The most well-known example of this is the TATA box, but in some promoters lacking a TATA box, such as the promoters of the mammalian terminal deoxynucleotidyl transferase gene and the SV40 late gene promoters, separate elements surrounding the start site themselves help to anchor the start site. Additional promoter elements regulate the frequency of transcription initiation. These are located in the region 30-110 bp upstream of the start site, but some promoters have been shown to contain functional elements downstream of the start site as well. To place a coding sequence "under the control" of a promoter, the 5' end of the transcription start site of the transcriptional reading frame is positioned "downstream" (i.e., 3') of the selected promoter. The "upstream" promoter stimulates DNA transcription, promoting expression of the encoded RNA.

[0167] The spacing between promoter elements is often flexible, allowing elements to be inverted or moved relative to one another while still maintaining promoter function. In the tk promoter, the spacing between promoter elements can be increased by up to 50 bp before activity begins to decline. Depending on the promoter, individual elements may function cooperatively or independently to activate transcription. Promoters may or may not be used in conjunction with "enhancers," which refer to cis-acting regulatory sequences involved in the transcriptional activation of a nucleic acid sequence.

[0168] A promoter may be one naturally associated with a nucleic acid sequence, such as can be obtained by isolating 5-prime non-coding sequences located upstream of a coding segment and / or exon. Such a promoter may be referred to as "endogenous." Similarly, an enhancer may be one naturally associated with a nucleic acid sequence, located either downstream or upstream of that sequence. Alternatively, certain advantages may be obtained by placing a coding nucleic acid segment under the control of a recombinant or heterologous promoter, which refers to a promoter not normally associated with a nucleic acid sequence in its natural environment. A recombinant or heterologous enhancer also refers to an enhancer not normally associated with a nucleic acid sequence in its natural environment. Such promoters or enhancers may include promoters or enhancers of other genes, and promoters or enhancers isolated from other viruses, or from prokaryotic or eukaryotic cells, as well as promoters or enhancers that are "non-naturally occurring," i.e., promoters or enhancers containing different elements of different transcriptional regulatory regions and / or mutations that alter expression. For example, promoters most commonly used in recombinant DNA construction include the lactamase (penicillinase), lactose, and tryptophan (trp) promoter systems. In addition to synthetically producing promoter and enhancer nucleic acid sequences, the sequences may be produced using recombinant cloning and / or nucleic acid amplification techniques, including PCR™, in conjunction with the compositions disclosed herein (see U.S. Pat. Nos. 4,683,202 and 5,928,906, each of which is incorporated herein by reference). Additionally, control sequences that direct transcription and / or expression of sequences in non-nuclear organelles, such as mitochondria, chloroplasts, etc., may similarly be used.

[0169] Naturally, it will be important to use a promoter and / or enhancer that effectively directs expression of the DNA segment in the organelle, cell type, tissue, organ, or organism selected for expression. Those skilled in the art of molecular biology generally know the use of promoter, enhancer, and cell type combinations for protein expression (see, for example, Sambrook et al. 1989, incorporated herein by reference). The promoter used may be constitutive, tissue-specific, inducible, and / or useful under appropriate conditions to direct high-level expression of the introduced DNA segment, which is advantageous for large-scale production of recombinant proteins and / or peptides. The promoter may be heterologous or endogenous.

[0170] Additionally, any promoter / enhancer combination can be used to drive expression. The use of T3, T7, or SP6 cytoplasmic expression systems is another embodiment. Eukaryotic cells can support cytoplasmic transcription from certain bacterial promoters if the appropriate bacterial polymerase is provided as part of the delivery complex or as an additional gene expression construct.

[0171] The identity of tissue-specific promoters or elements, as well as assays to characterize their activity, are well known to those of skill in the art.

[0172] Specific initiation signals may also be required for efficient translation of the coding sequence. These signals include the ATG initiation codon or adjacent sequences. Exogenous translational control signals, including the ATG initiation codon, may need to be provided. One of ordinary skill in the art would readily be able to determine this and provide the necessary signals.

[0173] In certain embodiments of the present disclosure, the use of internal ribosome entry site (IRES) elements is used to create multigenic, or polycistronic, messages, which can be used in the present invention.

[0174] Vectors can contain a multiple cloning site (MCS), a nucleic acid region containing multiple restriction enzyme sites, any of which can be used to digest the vector in conjunction with standard recombinant techniques. "Restriction enzyme digestion" refers to the catalytic cleavage of a nucleic acid molecule by an enzyme that functions only at specific locations in the nucleic acid molecule. Many of these restriction enzymes are commercially available. The use of such enzymes is widely understood by those skilled in the art. Restriction enzymes that cut within the MCS are often used to linearize or fragment a vector so that an exogenous sequence can be ligated into the vector. "Ligation" refers to the process of forming phosphodiester bonds between two nucleic acid fragments, which may or may not be adjacent to each other. Techniques involving restriction enzymes and ligation reactions are well known to those skilled in the art of recombinant technology.

[0175] Splice sites, termination signals, origins of replication, and selectable markers may also be used.

[0176] Plasmid vector In certain embodiments, a plasmid vector can be used to transform a host cell. Plasmid vectors containing replicon and control sequences derived from species compatible with the host cell are used in connection with these hosts. The vector typically carries a replication site and marking sequences capable of providing phenotypic selection in transformed cells. In a non-limiting example, Escherichia coli (E. coli) is often transformed using derivatives of pBR322, a plasmid derived from the E. coli species. pBR322 contains genes for ampicillin and tetracycline resistance, allowing easy identification of transformed cells. The pBR plasmid, or other microbial plasmid or phage, must also contain, or be modified to contain, a promoter that can be used by the microorganism for expression of its own proteins, for example.

[0177] Additionally, phage vectors containing replicon and control sequences compatible with host microorganisms can be used as transforming vectors in connection with these hosts. For example, phage lambda GEM.TM.11 can be utilized to generate recombinant phage vectors that can be used to transform host cells such as E. coli LE392.

[0178] Further useful plasmid vectors include pIN vectors (Inouye et al., 1985), and pGEX vectors, which are used to generate glutathione S-transferase (GST) soluble fusion proteins for subsequent purification and isolation or cleavage. Other suitable fusion proteins are those with galactosidase, ubiquitin, etc.

[0179] Bacterial host cells, e.g., E. coli, containing the expression vector are grown in any of a number of suitable media, e.g., LB. As will be understood by those skilled in the art, expression of the recombinant protein in a particular vector can be induced by contacting the host cells with an agent specific for the particular promoter, e.g., by adding IPTG to the medium or by switching the incubation to a higher temperature. After culturing the bacteria for an additional period, e.g., 2-24 hours, the cells are harvested by centrifugation and washed to remove residual medium.

[0180] viral vectors The abilities of certain viruses to infect cells via receptor-mediated endocytosis, enter cells, integrate into the host cell genome, and stably and efficiently express viral genes make them attractive candidates for introducing foreign nucleic acids into cells (e.g., mammalian cells). A component of the present invention can be a viral vector encoding one or more CARs of the present invention. Non-limiting examples of viral vectors that can be used to deliver the nucleic acids of the present invention are described below.

[0181] Adenovirus vectors A specific method for nucleic acid delivery involves the use of adenoviral expression vectors. Adenoviral vectors are known to have a low ability to integrate into genomic DNA, but this feature is offset by the high gene transfer efficiency achieved by these vectors. The term "adenoviral expression vector" refers to a construct containing sufficient adenoviral sequences (a) to support packaging of the construct and (b) to ultimately express a tissue- or cell-specific construct cloned therein. Due to the genetic makeup or knowledge that adenovirus is a 36 kb, linear, double-stranded DNA virus, large segments of adenoviral DNA can be replaced with foreign sequences up to 7 kb in size (Grunhaus and Horwitz, 1992).

[0182] AAV vectors Adenovirus-assisted transfection can be used to introduce nucleic acids into cells. Increased transfection efficiency has been reported in cell lines using adenovirus coupled systems (Kelleher and Vos, 1994; Cotten et al., 1992; Curiel, 1994). Adeno-associated virus (AAV) is an attractive vector system for use in the cells of the present invention because of its high integration frequency and ability to infect non-dividing cells, and is therefore useful for gene delivery to mammalian cells, for example, in tissue culture (Muzyczka, 1992) or in vivo. AAV has a wide infectious host range (Tratschin et al., 1984; Laughlin et al., 1986; Lebkowski et al., 1988; McLaughlin et al., 1988). Details regarding the generation and use of rAAV vectors are described in U.S. Patent Nos. 5,139,941 and 4,797,368, each of which is incorporated herein by reference.

[0183] Retroviral vectors Retroviruses are useful as delivery vectors due to their ability to integrate their genes into the host genome, to introduce large amounts of foreign genetic material, to infect a wide range of species and cell types, and to be packaged in specialized cell lines (Miller, 1992).

[0184] To construct a retroviral vector, a nucleic acid (e.g., encoding a sequence of interest) is inserted into the viral genome in place of a specific viral sequence, producing a replication-deficient virus. To produce virions, a packaging cell line containing the gag, pol, and env genes but without the long-term repeat (LTR) and packaging components is constructed (Mann et al., 1983). When a recombinant plasmid containing a cDNA along with the retroviral long-term repeat (LTR) and packaging sequences is introduced into a specialized cell line (e.g., by calcium phosphate precipitation), the packaging sequences enable the RNA transcripts of the recombinant plasmid to be packaged into viral particles, which are then secreted into the culture medium (Nicolas and Rubenstein, 1988; Temin, 1986; Mann et al., 1983). The medium containing the recombinant retrovirus is then collected, optionally concentrated, and used for gene transfer. Retroviral vectors can infect a variety of cell types; however, integration and stable expression require host cell division (Paskind et al., 1975).

[0185] Lentiviruses are complex retroviruses that contain the common retroviral genes gag, pol, and env, as well as other genes with regulatory or structural functions. Lentiviral vectors are well known in the art (see, e.g., Naldini et al., 1996; Zufferey et al., 1997; Blomer et al., 1997; U.S. Patent Nos. 6,013,516 and 5,994,136). Some examples of lentiviruses include human immunodeficiency viruses (HIV-1, HIV-2) and simian immunodeficiency viruses (SIV). Lentiviral vectors are generated by multiple attenuation of HIV pathogenicity genes, e.g., deletion of genes env, vif, vpr, vpu, and nef, resulting in biologically safe vectors.

[0186] Recombinant lentiviral vectors can infect non-dividing cells and can be used for gene transfer and nucleic acid sequence expression both in vivo and ex vivo. For example, recombinant lentiviruses can infect non-dividing cells in which suitable host cells have been transfected with two or more vectors carrying packaging functions, namely gag, pol, and env, as well as rev and tat, as described in U.S. Patent No. 5,994,136, incorporated herein by reference. To target receptors on specific cell types, recombinant viruses can be targeted by binding the envelope protein to antibodies or specific ligands. For example, by inserting a sequence of interest (including regulatory regions) into a viral vector along with another gene encoding a ligand for a receptor on a specific target cell, the vector is now target-specific.

[0187] Other viral vectors Other viral vectors can be used as vaccine constructs in the present invention. Vectors derived from viruses such as vaccinia virus (Ridgeway, 1988; Baichwal and Sugden, 1986; Coupar et al., 1988), Sindbis virus, cytomegalovirus, and herpes simplex virus can be used. These offer several attractive features in various mammalian cells (Friedmann, 1989; Ridgeway, 1988; Baichwal and Sugden, 1986; Coupar et al., 1988; Horwich et al., 1990).

[0188] Delivery using modified viruses The nucleic acid to be delivered may be housed within an infectious virus engineered to express a specific binding ligand. Thus, the viral particle specifically binds to the cognate receptor on the target cell and delivers its contents into the cell. A new approach designed to enable specific targeting of retroviral vectors has been developed based on the chemical modification of retroviruses by adding lactose residues to the viral envelope. This modification allows specific infection of hepatocytes via sialoglycoprotein receptors.

[0189] Another approach to targeting recombinant retroviruses was designed, using biotinylated antibodies against retroviral envelope proteins and specific cellular receptors. The antibodies were conjugated via the biotin moiety using streptavidin (Roux et al., 1989). Using antibodies against major histocompatibility complex class I and class II antigens, they demonstrated in vitro infection of various human cells bearing these surface antigens by ecotropic viruses (Roux et al., 1989).

[0190] Vector delivery and cell transformation Suitable methods for nucleic acid delivery for cell transfection or transformation are known to those skilled in the art.Such methods include, but are not limited to, direct delivery of DNA by ex vivo transfection, injection, etc.By applying techniques known in the art, cells can be stably or transiently transformed.

[0191] Ex vivo transformation Methods for transfecting eukaryotic cells and tissues ex vivo are known to those skilled in the art. Thus, the nucleic acids of the present invention can be used to transfect cells or tissues ex vivo. In some aspects, the transplanted cells or tissues can be placed in an organism. In other embodiments, the nucleic acid is expressed in the transplanted cells.

[0192] Kits of the Invention Any of the compositions described herein can be included in a kit.In a non-limiting example, the kit can include one or more cells for use in cell therapy that contain a recombinant expression vector, and / or reagents for generating one or more cells for use in cell therapy.The components of the kit are provided in suitable container means.

[0193] Some components of the kit may be packaged in aqueous media or in lyophilized form. The container means of the kit may include at least one vial, test tube, flask, bottle, syringe, or other container means into which the components may be placed and suitably dispensed. If the kit has two or more components, the kit may also contain a second, third, or other additional container into which the additional components may be individually placed. However, various combinations of components may be contained in vials. The kits of the present invention may also include means for containing the components in close confinement for commercial sale. Such containers may include injection-molded or blow-molded plastic containers into which the desired vials are retained.

[0194] When the kit components are provided in one and / or more liquid solutions, the liquid solution is an aqueous solution, with a sterile aqueous solution being particularly useful. In some cases, the container means itself may be a syringe, pipette, and / or other similar device from which the formulation may be applied to an infected area of ​​the body, injected into an animal, and / or applied to and / or mixed with other components of the kit.

[0195] However, the components of the kit may also be provided as a dry powder(s). When reagents and / or components are provided as a dry powder, the powder may be reconstituted by the addition of a suitable solvent. For example, the solvent may also be provided in another container means. The kit may also comprise a second container means for containing a sterile pharmaceutically acceptable buffer and / or other diluent.

[0196] In certain embodiments of the present invention, the cells used for cell therapy are provided in a kit, and in some cases, the cells are essentially the only component of the kit. The kit may include reagents and materials for producing the desired cells. In certain embodiments, the reagents and materials include primers for amplifying the desired sequence, nucleotides, appropriate buffers or buffer reagents, salts, etc., and in some cases, the reagents include a vector and / or DNA encoding a CAR described herein and / or regulatory elements therefor.

[0197] In certain embodiments, there are one or more devices in the kit suitable for extracting one or more samples from an individual. The device may be a syringe, a scalpel, or the like.

[0198] In some cases of the present invention, in addition to the cell therapy embodiment, the kit also includes a second cancer therapy, such as, for example, chemotherapy, hormone therapy, and / or immunotherapy. The kit(s) can be tailored to the individual's particular cancer and can include the individual's respective second cancer therapy.

[0199] Combination therapy In certain embodiments of the present invention, the clinical aspects of the methods of the present invention are combined with other agents effective in treating hyperproliferative diseases, such as anti-cancer agents. "Anti-cancer" agents can adversely affect cancer in a subject by, for example, killing cancer cells, including apoptosis in cancer cells, reducing the rate of cancer cell proliferation, reducing the incidence or number of metastases, reducing tumor size, inhibiting tumor growth, reducing the blood supply to tumors or cancer cells, promoting an immune response to cancer cells or tumors, preventing or inhibiting cancer progression, or extending the lifespan of a subject with cancer. For example, these other compositions would be provided in combined amounts effective to kill or inhibit cell proliferation. This process can involve simultaneously contacting the cancer cells with the expression construct and the agent(s) or multiple factor(s). This can be accomplished by contacting the cells with a single composition or pharmacological formulation containing both agents, or by simultaneously contacting the cells with two different compositions or formulations (one composition containing the expression construct and the other composition containing the second agent(s)).

[0200] Resistance of tumor cells to chemotherapy and radiotherapy agents is a major problem in clinical oncology. One goal of current cancer research is to find ways to improve the effectiveness of chemotherapy and radiotherapy by combining them with other treatments. In one embodiment, cell therapy can be used in conjunction with chemotherapy, radiotherapy, or immunotherapy intervention, as well as with proapoptotic or cell cycle regulating agents.

[0201] Alternatively, the treatment of the present invention can precede or follow the treatment of the other agent by intervals ranging from minutes to weeks. In embodiments in which the other agent and the present invention are applied separately, it is generally necessary to ensure that the effective period between each delivery does not expire so that the agent and the treatment of the present invention can still exert their beneficial combined effect on the cells. In such cases, cells may be contacted with both modalities within about 12-24 hours of each other (e.g., within about 6-12 hours of each other). In some situations, it may be desirable to significantly extend the treatment period if several days (2, 3, 4, 5, 6, or 7) to several weeks (1, 2, 3, 4, 5, 6, 7, or 8) pass between each administration.

[0202] Treatment cycles would be repeated as necessary. For example, various standard therapies, as well as surgical interventions, may be applied in combination with the cell therapy of the present invention.

[0203] chemotherapy Cancer treatment also includes a variety of combination therapies using both chemical and radiation-based treatments. Combination chemotherapy includes, but is not limited to, e.g., Abraxane, altretamine, docetaxel, Herceptin, methotrexate, novantrone, zoladex, cisplatin (CDDP), carboplatin, procarbazine, mechlorethamine, cyclophosphamide, camptothecin, ifosfamide, melphalan, chlorambucil, busulfan, nitrosurea, dactinomycin, daunorubicin, doxorubicin, bleomycin, plicomycin, mitomycin, etoposide (VP16), tamoxifen, raloxifene, estrogen receptor binding agents, taxol, gemcitabine, navelbine, farnesyl protein tansferase inhibitors, transplatinum, 5-fluorouracil, vincristine, vinblastine, and methotrexate, or analog or derivative variants of any of the foregoing, and combinations thereof.

[0204] In certain embodiments, chemotherapy for an individual is employed in combination with the present invention, eg, before, during, and / or after administration of the present invention.

[0205] Radiation therapy Other agents that cause DNA damage and have been widely used include gamma rays, commonly known as X-rays, and / or the directed delivery of radioisotopes to tumor cells. Other forms of DNA damaging agents, such as microwave and ultraviolet radiation, are also useful. All of these agents most likely cause widespread damage to DNA, DNA precursors, DNA replication and repair, and chromosome assembly and maintenance. X-ray doses range from daily doses of 50–200 roentgens for prolonged periods (3–4 weeks) to single doses of 2000–6000 roentgens. Dose ranges for radioisotopes vary widely and depend on the half-life of the isotope, the strength and type of radiation emitted, and uptake by neoplastic cells.

[0206] As used herein, the terms "contact" and "exposure," as applied to cells, are used to describe the process by which a therapeutic construct and a chemotherapeutic or radiotherapeutic agent are delivered to or directly juxtaposed with a target cell. To achieve cell killing or stasis, both agents are delivered to the cell in a combined amount effective to kill the cell or prevent it from dividing.

[0207] immunotherapy Immunotherapy relies on the use of immune effector cells and molecules to target and destroy cancer cells. The immune effector can be, for example, an antibody specific to some marker on the surface of tumor cells. The antibody alone can function as the therapeutic effector or can recruit other cells to actually kill the cells. Antibodies can also be conjugated to drugs or toxins (such as chemotherapeutic agents, radionuclides, ricin A chain, cholera toxin, pertussis toxin, etc.) and simply function as targeting agents. Alternatively, the effector can be a lymphocyte carrying a surface molecule that interacts directly or indirectly with the tumor cell target. Various effector cells include cytotoxic T cells and NK cells.

[0208] Therefore, immunotherapies other than the therapies of the present invention described herein can be used in combination with the cell therapy as part of a combination therapy. Combination therapy approaches are described herein. For example, tumor cells must have some markers that are amenable to targeting, i.e., that are not present on the majority of other cells. Many tumor markers exist, any of which may be suitable for targeting in the context of the present invention. Common tumor markers include PD-1, PD-L1, CTLA4, carcinoembryonic antigen, prostate-specific antigen, urinary tumor-associated antigen, fetal antigen, tyrosinase (p97), gp68, TAG-72, HMFG, sialyl Lewis antigen, MucA, MucB, PLAP, estrogen receptor, laminin receptor, erb B, and p155.

[0209] gene In yet another embodiment, the secondary treatment is gene therapy in which a therapeutic polynucleotide is administered before, after, or simultaneously with the clinical embodiments of the present invention. A variety of expression products are encompassed by the present invention, including inducers of cell proliferation, inhibitors of cell proliferation, or regulators of programmed cell death.

[0210] surgery Approximately 60% of cancer patients will undergo some type of surgery, including preventative, diagnostic, staging, curative and palliative surgery. Curative surgery is a cancer treatment that can be used in combination with other therapies, such as the treatment of the present invention, chemotherapy, radiation therapy, hormone therapy, gene therapy, immunotherapy and / or alternative therapies.

[0211] Curative surgery includes resection, in which all or part of the cancerous tissue is physically removed, excised, and / or destroyed. Tumor resection refers to the physical removal of at least part of the tumor. In addition to tumor resection, surgical treatments include laser surgery, cryosurgery, electrosurgery, and microsurgery (Mohs surgery). For example, the present invention can be used in combination with the removal of superficial cancers, pre-cancers, or incidental amounts of normal tissue.

[0212] When all cancerous cells, tissues, or parts of tumors are removed, a cavity may be formed in the body. Treatment can be achieved by perfusion, direct injection, or local application of additional anti-cancer therapy to the site. Such treatment can be repeated, for example, every 1, 2, 3, 4, 5, 6, or 7 days, or every 1, 2, 3, 4, and 5 weeks, or every 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months. These treatments can also be at various dosages.

[0213] Other drugs In some embodiments, other agents can be used in combination with the present invention to improve the therapeutic efficacy of the treatment. These additional agents include immunomodulators, agents that affect the upregulation of cell surface receptors and GAP junctions, cytostatic and differentiation agents, cell adhesion inhibitors, or agents that enhance the sensitivity of hyperproliferative cells to apoptosis inducers. Immunomodulators include tumor necrosis factor, interferon alpha, beta, and gamma; IL-2 and other cytokines; F42K and other cytokine analogs; or MIP-1, MIP-1 beta, MCP-1, RANTES, and other chemokines. In some embodiments, upregulation of cell surface receptors or their ligands, such as Fas / Fas ligand, DR4, or DR5 / TRAIL, will enhance the apoptosis-inducing capabilities of the present invention by establishing autocrine or paracrine effects on hyperproliferative cells. Increasing intercellular signaling by increasing the number of GAP junctions will increase the anti-hyperproliferative effect on adjacent hyperproliferative cell populations. In other embodiments, cytostatic or differentiation agents can be used in combination with the present invention to improve the anti-hyperproliferative efficacy of the treatment. Cell adhesion inhibitors can also be used to improve the efficacy of the present invention. Examples of cell adhesion inhibitors are focal adhesion kinase (FAK) inhibitors and lovastatin. In some embodiments, other agents that increase the sensitivity of hyperproliferative cells to apoptosis, such as the antibody c225, can be used in combination with the present invention to improve therapeutic efficacy.

[0214] Methods for assessing the activity of engineered CAR T cells Aspects of the present disclosure are further directed to methods and kits for evaluating the killing capacity of engineered CAR T cells. Specifically, embodiments are directed to immunocomplex analysis methods and kits for determining CAR T cell activity during co-culture with cancer cells. First, various target cancer cells (e.g., HEK293T, MDA-MB-231, MDA-MB-468, HCC38, and skrc59) are stained with a dye (e.g., ViaStain™ Tracer Blue dye), seeded into a plate (e.g., a 96-well plate), and incubated for a period of time (e.g., 12 hours or overnight). Next, different T cell types (e.g., two different T cell types) are added to the wells (e.g., at an effector-to-target (E:T) ratio of 20:1) and co-cultured for a period of time (e.g., 24 hours). Finally, the plate is scanned and analyzed (e.g., using bright field and blue fluorescent channels). Immune complexes were analyzed by confluence measurement and compared to a negative control of untransduced T cells. The resulting data plots displayed CAR T cell activity for all target and effector cell combinations tested. The use of an image cytometry platform allows for visual confirmation of the interaction between effector and target cells, resulting in accurate and robust results. [Example]

[0215] Examples are provided below to facilitate a more complete understanding of the present invention. The following examples illustrate exemplary modes of making and practicing the present invention. However, the scope of the present invention is not limited to the specific embodiments disclosed in these examples, which are for illustrative purposes only, as alternative methods may be used to obtain similar results.

[0216] Example 1 Clear cell renal cell carcinoma (ccRCC) is the leading type of RCC and one of the 10 most common cancers in both men and women. Chimeric antigen receptor (CAR) T cells have proven to be a potent and clinically translatable immunotherapy for hematological malignancies. However, these results are not translatable to solid tumors due to inefficient CAR T cell homing, an inhibitory tumor microenvironment, and on-target off-tumor toxicity resulting from the sharing of CAR T-targeting epitopes on healthy tissue. To combat the inhibitory microenvironment, immune checkpoint blockade has shown enhanced efficacy on antitumor responses by restoring local antitumor immunity. A CAR T cell factory was engineered to empower CAR T cells by secreting human anti-immune checkpoint inhibitor monoclonal antibodies (mAbs) locally at the tumor site. Our results demonstrate that reversing the depletion of CAR T cells and tumor-infiltrating lymphocytes (TILs) dramatically improves CAR T-mediated killing of ccRCC in vivo and in vitro.

[0217] CAIX is an ideal target for ccRCC therapy and was used as the CAR target in initial clinical trials (see, e.g., Lamers, Sleijfer et al. 2006; Lamers, Willemsen et al. 2011). However, it caused serious side effects due to CAIX expression on the bile duct. Therefore, it is important to develop CARs with high efficacy and safety (e.g., limiting on-target off-tumor effects). To achieve this, second-generation CARs were developed by introducing a second targeting scFv (e.g., anti-CD70 scFv) into the CAR T cell factory together with the first targeting scFv (e.g., anti-CAIX scFv), allowing the CAR to simultaneously target two unique antigens. For example, see Figure 1. IHC staining of ccRCC patient samples revealed that CD70, highly expressed on ccRCC and coexpressed with CAIX, was an ideal target to use as a second target.

[0218] Our 27 billion-member human scFv-phage display library was panned against antigen-expressing skrc-59 ccRCC cells, subtracting antigen-free skrc-59 ccRCC cells, to identify novel scFvs. Their binding kinetics (K on / K off ) and affinity (K d ) were then measured via an OctetRed 96 instrument. ScFvs with desirable kinetics were evaluated for their ability to bind antigen-expressing cells. Candidates were cloned into vectors in which anti-CD70 and anti-CAIX scFvs were combined in different permutations by altering the order of the two targeting scFvs with various linkers connected to the costimulatory domains (CD28, 41BB) and activation domains (CD3). Using a fourth-generation lentiviral packaging system, we obtained CAR lentiviruses and transduced primary T cells isolated from PBMCs to express the dual CARs, which were then tested against different cell lines in vitro. For further evaluation in vivo, a humanized orthotopic ccRCC mouse model was established by injecting luciferized ccRCC cells under the kidney capsule of NSG-SGM3 mice with a reconstituted human immune system.

[0219] In summary, by utilizing the double CAR T discovery platform, a series of CARs with different scFv, linker and hinge are generated.Those skilled in the art will recognize that embodiments can include different combinations of scFv, linker and hinge.For example, different combinations of scFv, linker and hinge can be used to treat different cancers.

[0220] By combining the best dual CARs with payloads such as immune checkpoint inhibitor payloads, second-generation CAR Ts have been discovered. Without wishing to be bound by theory, second-generation CAR T cell factories can be used to treat cancers such as ccRCC, eliminating side effects on normal tissues.

[0221] References cited in this example Lamers, CH, S. Sleijfer, AGVulto, WHKruit, M. Kliffen, R. Debets, JW Gratama, G. Stoter and E. Oosterwijk (2006). “Treatment of metastatic renal cell carcinoma with autologous T-lymphocytes genetically retargeted against carbonic anhydrase IX:first clinical experience.” J Clin Oncol 24(13):e20-22. Lamers, CH, R. Willemsen, P. van Elzakker, S. van Steenbergen-Langeveld, M. Broertjes, J. Oosterwijk-Wakka, E. Oosterwijk, S. Sleijfer, R. Debets and JW Gratama (2011).”Immune responses to transgene and retroviral vector in patients treated with ex vivo-engineered T cells.”Blood 117(1):72-82.

[0222] Example 2 Chimeric antigen receptor (CAR) T cells have proven to be a potent immunotherapy for hematological malignancies, but these have not been translated to solid tumors. Our CAR T cell factory empowers CAR T cells by secreting antibodies, such as human anti-immune checkpoint inhibitor monoclonal antibodies (mAbs), locally at the tumor site, restoring the tumor microenvironment and achieving cure for solid tumors. With the introduction of one or more additional scFvs, second-generation CARs have shown improved efficacy and safety, showing great promise in the clinic.

[0223] We designed a bispecific tandem CAR to target tumor-associated antigens, such as CAIX and CD70 (see, e.g., Figures 1 and 2), which are highly expressed and co-expressed on primary ccRCC cells by IHC (Figure 3). By successfully panning a 27 billion-member phage display library, several anti-CD70 scFvs were identified (see, e.g., Figure 6). Our previously discovered anti-CAIX and anti-CD70 scFvs were cloned into pHAGE vectors with different linkers (see, e.g., Figure 9). Lentiviruses were then packaged and transduced into primary T cells. CAR T cells were obtained and evaluated in four different CRISPR-engineered cell lines (see, e.g., Figure 5). Killing activity was assessed by Celigo and Cr51 release assays (see, e.g., Figures 12 and 13). Selectivity was demonstrated as described herein.

[0224] This CAR T cell factory can be used to treat CAIX and / or CD70 overexpressing cancers or in combination with other therapies.

[0225] Example 3 Quantification of double IHC staining for CAIX and CD70 TIFF2025011142000026.tif149166

[0226] equivalent Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, numerous equivalents to the specific substances and procedures specifically described herein which equivalents are considered to be within the scope of this invention and covered by the appended claims.

[0227] Sequence information SEQUENCE LISTING <110> DANA-FARBER CANCER INSTITUTE, INC. <120> CHIMERIC ANTIGEN RECEPTOR FACTORIES AND METHODS OF USE THEREOF <150> US 62 / 826,462 <151> 2019-03-29 <150> US 62 / 773,885 <151> 2018-11-30 <160> 352 <170> PatentIn version 3.5 <210> 1 <400> 1 000 <210> 2 <400> 2 000 <210> 3 <400> 3 000 <210> 4 <400> 4 000 <210> 5 <400> 5 000 <210> 6 <400> 6 000 <210> 7 <400> 7 000 <210> 8 <400> 8 000 <210> 9 <400> 9 000 <210> 10 <400> 10 000 <210> 11 <400> 11 000 <210> 12 <400> 12 000 <210> 13 <400> 13 000 <210> 14 <400> 14 000 <210> 15 <400> 15 000 <210> 16 <400> 16 000 <210> 17 <400> 17 000 <210> 18 <400> 18 000 <210> 19 <400> 19 000 <210> 20 <400> 20 000 <210> 21 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 21 Gly Tyr Thr Phe Ala Ser Tyr Tyr 1 5 <210> 22 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 22 Gly Tyr Thr Phe Ala Ser Gln Trp 1 5 <210> 23 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 23 Gly Tyr Thr Phe Ala Ser Ser Trp 1 5 <210> 24 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 24 Gly Tyr Thr Phe Ala Ser Gln Tyr 1 5 <210> 25 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 25 Gly Tyr Thr Phe Ala Ser Ala Trp 1 5 <210> 26 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 26 Gln Ser Ile Leu Tyr Ser Ser Asn Gln Lys Asn Tyr 1 5 10 <210> 27 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 27 Ile Asn Pro Gly Asn Val Asn Thr 1 5 <210> 28 <211> 6 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 28 Trp Ala Ser Thr Arg Glu 1 5 <210> 29 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 29 Ser Thr Tyr Tyr Arg Pro Leu Asp Tyr 1 5 <210> 30 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 30 Ser Thr Trp Tyr Arg Pro Leu Asp Tyr 1 5 <210> 31 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 31 Ser Thr Trp Tyr Arg Pro Asn Asp Tyr 1 5 <210> 32 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 32 Thr Thr Arg Tyr Arg Pro Leu Asp Tyr 1 5 <210> 33 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 33 Leu Thr Tyr Tyr Arg Pro Pro Asp Tyr 1 5 <210> 34 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 34 His Gln Tyr Leu Ser Ser Tyr Thr 1 5 <210> 35 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 35 His Gln Tyr Ile Ser Ser Tyr Thr 1 5 <210> 36 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 36 His Gln Tyr Lys Ser Ser Tyr Thr 1 5 <210> 37 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 37 His Gln Tyr Arg Ser Ser Tyr Thr 1 5 <210> 38 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 38 His Gln Tyr Tyr Ser Ser Tyr Thr 1 5 <210> 39 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 39 His Gln Tyr Met Ser Ser Tyr Thr 1 5 <210> 40 <211> 4 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 40 Gly Gly Gly Ser 1 <210> 41 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 41 Gly Gly Gly Gly Ser Ser Ser 1 5 <210> 42 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 42 Gly Gly Gly Gly Ser Ser Ser Ser Ser 1 5 <210> 43 <211> 6 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 43 Glu Glu Asp Leu Pro Glu 1 5 <210> 44 <211> 5 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 44 Thr Tyr Ala Met Thr 1 5 <210> 45 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 45 Thr Gly Ser Arg Ser Asn Ile Gly Ala Asp Tyr Asp Val His 1 5 10 <210> 46 <211> 5 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 46 Ser Tyr Ala Met Ser 1 5 <210> 47 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 47 Thr Gly Ser Ser Ser Asn Ile Gly Arg Gly Tyr Asn Val His 1 5 10 <210> 48 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 48 Thr Gly Ser Ser Ser Asn Ile Gly Ala Gly Tyr Asp Val His 1 5 10 <210> 49 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 49 Gly Phe Thr Phe Ser Ser Tyr Ala 1 5 <210> 50 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 50 Ser Ser Asn Ile Gly Ala Gly Tyr Asp 1 5 <210> 51 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 51 Thr Gly Ser Ser Ser Asn Ile Gly Arg Gly Tyr Asn Val His 1 5 10 <210> 52 <211> 5 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 52 Asn Tyr Ala Met Thr 1 5 <210> 53 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 53 Gly Gly Asn Asn Ile Gly Ser Lys Ser Val Glu 1 5 10 <210> 54 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 54 Gly Phe Thr Phe Ser Asn Tyr Ala 1 5 <210> 55 <211> 6 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 55 Asn Ile Gly Ser Lys Ser 1 5 <210> 56 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 56 Thr Gly Ser Ser Ser Asn Ile Gly Ala Gly Phe Asp Val His 1 5 10 <210> 57 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 57 Thr Gly Thr Ser Ser Asn Ile Gly Ala Gly Tyr Asp Val His 1 5 10 <210> 58 <211> 5 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 58 Ser Tyr Gly Met His 1 5 <210> 59 <211> 13 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 59 Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn Tyr Val Tyr 1 5 10 <210> 60 <211> 5 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 60 Ile Tyr Ala Met Ser 1 5 <210> 61 <211> 5 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 61 Lys Tyr Ala Met Ser 1 5 <210> 62 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 62 Gln Gly Asn Ser Leu Arg Tyr Tyr Tyr Pro Ser 1 5 10 <210> 63 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 63 Gly Gly Asp Asn Ile Gly Arg Lys Ser Val His 1 5 10 <210> 64 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 64 Ala Val Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 65 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 65 Ala Asn Asn Asn Arg Pro Ser 1 5 <210> 66 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 66 Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 67 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 67 Gly Asn Thr Asn Arg Pro Ser 1 5 <210> 68 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 68 Ala Ile Ser Ala Asn Gly Gly Thr Thr Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 69 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 69 Gly Asn Ser Asn Arg Pro Ser 1 5 <210> 70 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 70 Ile Ser Gly Ser Gly Gly Ser Thr 1 5 <210> 71 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 71 Asp Asp Ile Asn Arg Pro Ser 1 5 <210> 72 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 72 Leu Ile Ser Tyr Asp Gly Ser Val Thr His Tyr Thr Asp Ser Val Lys 1 5 10 15 Gly <210> 73 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 73 Tyr Asp Ser Asp Arg Pro Ser 1 5 <210> 74 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 74 Ile Ser Tyr Asp Gly Ser Val Thr 1 5 <210> 75 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 75 Asp Asn Thr Asn Arg Pro Ser 1 5 <210> 76 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 76 Gly Asn Asn Asn Arg Pro Ser 1 5 <210> 77 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 77 Ala Ile Ser Gly Ser Gly Val Ser Thr Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 78 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 78 Val Ile Ser Tyr Asp Gly Ser Asn Lys Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 79 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 79 Arg Asn Asn Gln Arg Pro Ser 1 5 <210> 80 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 80 Ala Ile Ser Gly Ser Gly Gly Gly Thr Tyr His Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 81 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 81 Gly Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 82 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 82 Gly Lys Asn Asn Arg Pro Ser 1 5 <210> 83 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 83 Asp Asp Arg Asp Arg Pro Ser 1 5 <210> 84 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 84 Gly Pro Val Leu Arg Tyr Gly Phe Asp Ile 1 5 10 <210> 85 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 85 Gln Ser Tyr Asp Ser Ser Leu Arg Ala Trp Val 1 5 10 <210> 86 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 86 Ser His Ser Ser Gly Gly Phe Asp Tyr 1 5 <210> 87 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 87 Gln Ser Tyr Asp Ser Ser Leu Ser Ala Trp Val 1 5 10 <210> 88 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 88 Asn Gly Asn Tyr Arg Gly Ala Phe Asp Ile 1 5 10 <210> 89 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 89 Gln Ser Tyr Asp Arg Ser Leu Ser Trp Val 1 5 10 <210> 90 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 90 Ala Thr Tyr Gly Asp Tyr Gly Ser Leu Asp Tyr 1 5 10 <210> 91 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 91 Ala Ala Ala Gly Phe Asp Tyr 1 5 <210> 92 <211> 6 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 92 Gly Ser Gly Tyr Gln Glu 1 5 <210> 93 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 93 Gln Val Trp Asp Ser Ser Ser Asp His His Val Val 1 5 10 <210> 94 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 94 Ala Arg Gly Ser Gly Tyr Gln Glu His 1 5 <210> 95 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 95 Gln Ser Tyr Asp Ser Arg Leu Ser Ala Trp Val 1 5 10 <210> 96 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 96 Ile Gly Arg Tyr Ser Ser Ser Leu Gly Tyr 1 5 10 <210> 97 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 97 Gln Ser Tyr Asp Ser Gly Leu Arg Trp Val 1 5 10 <210> 98 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 98 Tyr Gly Asp Tyr Gly Ser Leu Asp Tyr 1 5 <210> 99 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 99 Gln Ser Tyr Asp Lys Ser Leu Ser Trp Val 1 5 10 <210> 100 <211> 13 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 100 Tyr Cys Ser Ser Thr Ser Cys Tyr Arg Gly Met Asp Val 1 5 10 <210> 101 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 101 Gln Ser Tyr Asp Lys Ser Leu Thr Trp Val 1 5 10 <210> 102 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 102 Gly Arg Ala Ala Arg Pro Pro Phe Asp Tyr 1 5 10 <210> 103 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 103 Ala Ala Trp Asp Asp Ser Leu Asn Gly Val Val 1 5 10 <210> 104 <211> 13 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 104 Glu Ala Pro Tyr Ser Ser Ser Leu Asp Ala Phe Asp Ile 1 5 10 <210> 105 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 105 His Ser Arg Asp Asn Asn Gly His His Ile 1 5 10 <210> 106 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 106 Phe Ser Ala Tyr Ser Gly Tyr Asp Leu 1 5 <210> 107 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 107 Gln Ser Tyr Asp Ser Thr Leu Arg Val Trp Met 1 5 10 <210> 108 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 108 Ser Ser Arg Ser Gly Tyr Phe Leu Pro Leu Asp Tyr 1 5 10 <210> 109 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 109 Ser Ser Arg Asp Asn Thr Asp Asn Arg Val Val 1 5 10 <210> 110 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 110 Ala Ala Val Thr Gly Gly Phe Asp Pro 1 5 <210> 111 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 111 Gln Val Trp Asp Ser Ser Ser Lys His Tyr Val 1 5 10 <210> 112 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 112 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Glu Phe Thr Phe Gly Thr Tyr 20 25 30 Ala Met Thr Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Val Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Arg Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gly Pro Val Leu Arg Tyr Gly Phe Asp Ile Trp Gly Gln Gly 100 105 110 Thr Met Val Ile Val Ser Ser 115 <210> 113 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 113 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Ile Thr Ile Ser Cys Thr Gly Ser Arg Ser Asn Ile Gly Ala Asp 20 25 30 Tyr Asp Val His Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu 35 40 45 Leu Ile Tyr Ala Asn Asn Asn Arg Pro Ser Gly Val Pro Gly Arg Phe 50 55 60 Ser Ala Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu 65 70 75 80 Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr Asp Ser Ser 85 90 95 Leu Arg Ala Trp Val Phe Gly Gly Gly Thr Lys Leu Ala Val Leu Gly 100 105 110 <210> 114 <211> 118 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 114 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Ser His Ser Ser Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr 100 105 110 Leu Val Thr Val Ser Ser 115 <210> 115 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 115 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Arg Gly 20 25 30 Tyr Asn Val His Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu 35 40 45 Leu Ile Tyr Gly Asn Thr Asn Arg Pro Ser Gly Val Pro Asp Arg Phe 50 55 60 Ser Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Thr Gly Leu 65 70 75 80 Gln Ala Glu Asp Glu Gly Asp Tyr Tyr Cys Gln Ser Tyr Asp Ser Ser 85 90 95 Leu Ser Ala Trp Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly 100 105 110 <210> 116 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 116 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Pro Phe Ser Ser Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Ala Asn Gly Gly Thr Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Asn Asn Gly Asn Tyr Arg Gly Ala Phe Asp Ile Trp Gly Gln Gly 100 105 110 Thr Met Val Thr Val Ser Ser 115 <210> 117 <211> 111 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 117 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Ala Gly 20 25 30 Tyr Asp Val His Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu 35 40 45 Leu Ile Tyr Gly Asn Ser Asn Arg Pro Ser Gly Val Pro Asp Arg Phe 50 55 60 Ser Gly Ser Lys Ser Gly Ser Ser Ala Ser Leu Ala Ile Thr Gly Leu 65 70 75 80 Gln Ala Glu Asp Glu Ala His Tyr Tyr Cys Gln Ser Tyr Asp Arg Ser 85 90 95 Leu Ser Trp Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly 100 105 110 <210> 118 <211> 107 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 118 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Thr Tyr Gly Asp Tyr Gly Ser Leu Asp Tyr 100 105 <210> 119 <211> 101 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 119 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Ala Gly 20 25 30 Tyr Asp Val His Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu 35 40 45 Leu Ile Tyr Ala Asn Asn Asn Arg Pro Ser Gly Val Pro Asp Arg Phe 50 55 60 Ser Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Thr Gly Leu 65 70 75 80 Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr Asp Ser Ser 85 90 95 Leu Arg Ala Trp Val 100 <210> 120 <211> 116 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 120 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Ala Ala Ala Gly Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val 100 105 110 Thr Val Ser Ser 115 <210> 121 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 121 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Arg Gly 20 25 30 Tyr Asn Val His Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu 35 40 45 Leu Ile Tyr Asp Asp Ile Asn Arg Pro Ser Gly Val Pro His Arg Phe 50 55 60 Ser Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Thr Gly Leu 65 70 75 80 Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr Asp Ser Ser 85 90 95 Leu Arg Ala Trp Val Phe Gly Gly Gly Thr Lys Leu Ala Val Leu Gly 100 105 110 <210> 122 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <220> <221> MOD_RES <222> (83)..(83) <223> Any amino acid <400> 122 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Arg Gly 20 25 30 Tyr Asn Val His Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu 35 40 45 Leu Ile Tyr Gly Asn Thr Asn Arg Pro Ser Gly Val Pro Asp Arg Phe 50 55 60 Ser Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Thr Gly Leu 65 70 75 80 Gln Ala Xaa Asp Glu Gly Asp Tyr Tyr Cys Gln Ser Tyr Asp Ser Ser 85 90 95 Leu Ser Ala Trp Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly 100 105 110 <210> 123 <211> 116 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 123 Gln Val Thr Leu Lys Glu Ser Gly Gly Gly Val Val Gln Pro Gly Thr 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Ala Met Thr Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Gly Leu Ile Ser Tyr Asp Gly Ser Val Thr His Tyr Thr Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Thr Leu Arg Ala Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gly Ser Gly Tyr Gln Glu His Trp Gly Gln Gly Thr Leu Val 100 105 110 Thr Val Ser Ser 115 <210> 124 <211> 110 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 124 Leu Pro Val Leu Thr Gln Pro Pro Ser Val Ser Val Ala Pro Gly Gln 1 5 10 15 Thr Ala Arg Ile Thr Cys Gly Gly Asn Asn Ile Gly Ser Lys Ser Val 20 25 30 Glu Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val Ile Tyr 35 40 45 Tyr Asp Ser Asp Arg Pro Ser Gly Ile Pro Glu Arg Phe Ser Gly Ser 50 55 60 Asn Ser Gly Asn Thr Ala Thr Leu Thr Ile Ser Arg Val Glu Ala Gly 65 70 75 80 Asp Glu Ala Asp Tyr Tyr Cys Gln Val Trp Asp Ser Ser Ser Asp His 85 90 95 His Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly 100 105 110 <210> 125 <211> 105 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 125 Gln Val Thr Leu Lys Glu Ser Gly Gly Gly Val Val Gln Pro Gly Thr 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Ala Met Thr Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Gly Leu Ile Ser Tyr Asp Gly Ser Val Thr His Tyr Thr Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Thr Leu Arg Ala Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gly Ser Gly Tyr Gln Glu His 100 105 <210> 126 <211> 99 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 126 Leu Pro Val Leu Thr Gln Pro Pro Ser Val Ser Val Ala Pro Gly Gln 1 5 10 15 Thr Ala Arg Ile Thr Cys Gly Gly Asn Asn Ile Gly Ser Lys Ser Val 20 25 30 His Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val Ile Tyr 35 40 45 Tyr Asp Ser Asp Arg Pro Ser Gly Ile Pro Glu Arg Phe Ser Gly Ser 50 55 60 Asn Ser Gly Asn Thr Ala Thr Leu Thr Ile Ser Arg Val Glu Ala Gly 65 70 75 80 Asp Glu Ala Asp Tyr Tyr Cys Gln Val Trp Asp Ser Ser Ser Asp His 85 90 95 His Val Val <210> 127 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 127 Glu Val Gln Leu Val Gln Ser Gly Gly Gly Val Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Pro Phe Ser Ser Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Ala Asn Gly Gly Thr Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Asn Asn Gly Asn Tyr Arg Gly Ala Phe Asp Ile Trp Gly Gln Gly 100 105 110 Thr Met Val Thr Val Ser Ser 115 <210> 128 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 128 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Ala Gly 20 25 30 Phe Asp Val His Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu 35 40 45 Leu Ile Tyr Gly Asn Thr Asn Arg Pro Ser Gly Val Pro Asp Arg Phe 50 55 60 Ser Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Thr Gly Leu 65 70 75 80 Gln Ala Glu Asp Glu Thr Asp Tyr Tyr Cys Gln Ser Tyr Asp Ser Arg 85 90 95 Leu Ser Ala Trp Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly 100 105 110 <210> 129 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 129 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Val Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Pro Phe Ser Ser Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Ala Asn Gly Gly Thr Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Asn Asn Gly Asn Tyr Arg Gly Ala Phe Asp Ile Trp Gly Gln Gly 100 105 110 Thr Met Val Thr Val Ser Ser 115 <210> 130 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 130 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Ile Thr Ile Ser Cys Thr Gly Ser Arg Ser Asn Ile Gly Ala Asp 20 25 30 Tyr Asp Val His Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu 35 40 45 Leu Ile Tyr Ala Asn Asn Asn Arg Pro Ser Gly Val Pro Asp Arg Phe 50 55 60 Ser Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Thr Gly Leu 65 70 75 80 Gln Ala Glu Asp Glu Thr Asp Tyr Phe Cys Gln Ser Tyr Asp Ser Ser 85 90 95 Leu Ser Ala Trp Val Phe Gly Gly Gly Thr Lys Val Thr Val Leu Gly 100 105 110 <210> 131 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 131 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Ile Gly Arg Tyr Ser Ser Ser Leu Gly Tyr Trp Gly Gln Gly 100 105 110 Thr Leu Val Thr Val Ser Ser 115 <210> 132 <211> 111 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 132 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Arg Gly 20 25 30 Tyr Asn Val His Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu 35 40 45 Leu Ile Tyr Asp Asn Thr Asn Arg Pro Ser Gly Val Pro Ala Arg Phe 50 55 60 Ser Gly Ser Lys Ser Ala Thr Ser Ala Ser Leu Ala Ile Thr Gly Leu 65 70 75 80 Gln Ala Asp Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr Asp Ser Gly 85 90 95 Leu Arg Trp Val Phe Gly Gly Gly Thr Lys Leu Thr Leu Leu Arg 100 105 110 <210> 133 <211> 118 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 133 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Thr Tyr Gly Asp Tyr Gly Ser Leu Asp Tyr Trp Gly Gln Gly Thr 100 105 110 Leu Val Thr Val Ser Ser 115 <210> 134 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 134 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Ala Gly 20 25 30 Tyr Asp Val His Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu 35 40 45 Leu Ile Tyr Ala Asn Asn Asn Arg Pro Ser Gly Val Pro Asp Arg Phe 50 55 60 Ser Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Thr Gly Leu 65 70 75 80 Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr Asp Ser Ser 85 90 95 Leu Arg Ala Trp Val Phe Gly Gly Gly Thr Lys Leu Ala Val Leu Gly 100 105 110 <210> 135 <211> 111 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 135 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Ile Thr Ile Ser Cys Thr Gly Thr Ser Ser Asn Ile Gly Ala Gly 20 25 30 Tyr Asp Val His Trp Tyr Gln Gln Leu Pro Gly Ala Ala Pro Arg Val 35 40 45 Leu Ile Tyr Gly Asn Asn Asn Arg Pro Ser Gly Val Pro Asp Arg Phe 50 55 60 Ser Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Thr Gly Leu 65 70 75 80 Gln Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr Asp Lys Ser 85 90 95 Leu Ser Trp Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Arg 100 105 110 <210> 136 <211> 122 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 136 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Val Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Tyr Cys Ser Ser Thr Ser Cys Tyr Arg Gly Met Asp Val Trp 100 105 110 Gly Lys Gly Thr Leu Val Thr Val Ser Ser 115 120 <210> 137 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 137 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Arg Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Pro Phe Ser Ser Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Ala Asn Gly Gly Thr Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Asn Asn Gly Asn Tyr Arg Gly Ala Phe Asp Ile Trp Gly Gln Gly 100 105 110 Thr Thr Val Thr Val Ser Ser 115 <210> 138 <211> 111 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 138 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Ile Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Ala Gly 20 25 30 Tyr Asp Val His Trp Tyr Gln Gln Val Pro Gly Lys Ala Pro Lys Val 35 40 45 Val Ile Tyr Gly Asn Asn Asn Arg Pro Ser Gly Val Pro Asp Arg Phe 50 55 60 Ser Gly Ser Lys Ser Gly Ala Ser Ala Ser Leu Ala Ile Thr Gly Leu 65 70 75 80 Gln Thr Glu Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr Asp Lys Ser 85 90 95 Leu Thr Trp Val Phe Gly Gly Gly Thr Lys Val Thr Val Leu Gly 100 105 110 <210> 139 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 139 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Val Val Gln Pro Gly Arg 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Gly Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Val Ile Ser Tyr Asp Gly Ser Asn Lys Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gly Arg Ala Ala Arg Pro Pro Phe Asp Tyr Trp Gly Gln Gly 100 105 110 Thr Leu Val Thr Val Ser Ser 115 <210> 140 <211> 111 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 140 Gln Pro Val Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn 20 25 30 Tyr Val Tyr Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Pro 35 40 45 Ile Tyr Arg Asn Asn Gln Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Arg 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Ala Ala Trp Asp Asp Ser Leu 85 90 95 Asn Gly Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Arg 100 105 110 <210> 141 <211> 122 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 141 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Val Val Gln Pro Gly Arg 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Gly Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Val Ile Ser Tyr Asp Gly Ser Asn Lys Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Glu Ala Pro Tyr Ser Ser Ser Leu Asp Ala Phe Asp Ile Trp 100 105 110 Gly Gln Gly Thr Met Val Thr Val Ser Ser 115 120 <210> 142 <211> 111 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 142 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Arg Gly 20 25 30 Tyr Asn Val His Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu 35 40 45 Leu Ile Tyr Gly Asn Ser Asn Arg Pro Ser Gly Val Pro Asp Arg Phe 50 55 60 Ser Gly Ser Ser Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr Gly Ala 65 70 75 80 Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys His Ser Arg Asp Asn Asn 85 90 95 Gly His His Ile Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Ser 100 105 110 <210> 143 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 143 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Val Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Pro Phe Ser Ser Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Ala Asn Gly Gly Thr Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Asn Asn Gly Asn Tyr Arg Gly Ala Phe Asp Ile Trp Gly Gln Gly 100 105 110 Thr Met Val Thr Val Ser Ser 115 <210> 144 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 144 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Ala Gly 20 25 30 Tyr Asp Val His Trp Tyr Gln His Leu Pro Gly Thr Ala Pro Lys Leu 35 40 45 Leu Ile Tyr Gly Asn Ser Asn Arg Pro Ser Gly Val Pro Asp Arg Phe 50 55 60 Ser Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Thr Gly Leu 65 70 75 80 Gln Ala Glu Asp Glu Thr Asp Tyr Phe Cys Gln Ser Tyr Asp Ser Ser 85 90 95 Leu Ser Ala Trp Val Phe Gly Gly Gly Thr Lys Val Thr Val Leu Gly 100 105 110 <210> 145 <211> 118 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 145 Gln Val Gln Leu Gln Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ile Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Gly Gly Thr Tyr His Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Phe Ser Ala Tyr Ser Gly Tyr Asp Leu Trp Gly Gln Gly Thr 100 105 110 Leu Val Thr Val Ser Ser 115 <210> 146 <211> 111 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 146 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Arg Gly 20 25 30 Tyr Asn Val His Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu 35 40 45 Leu Ile Tyr Asp Asn Thr Asn Arg Pro Ser Gly Val Pro Ala Arg Phe 50 55 60 Ser Gly Ser Lys Ser Ala Thr Ser Ala Ser Leu Thr Ile Thr Gly Leu 65 70 75 80 Gln Ala Asp Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr Asp Ser Gly 85 90 95 Leu Arg Trp Val Phe Gly Gly Gly Thr Lys Leu Thr Leu Leu Gly 100 105 110 <210> 147 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 147 Glu Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Ala Asn Gly Gly Thr Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Asn Asn Gly Asn Tyr Arg Gly Ala Phe Asp Ile Trp Gly Gln Gly 100 105 110 Thr Thr Val Thr Val Ser Ser 115 <210> 148 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 148 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Ala Gly 20 25 30 Phe Asp Val His Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Arg Leu 35 40 45 Leu Ile Tyr Gly Asn Asn Asn Arg Pro Ser Gly Val Pro Asp Arg Phe 50 55 60 Ser Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Thr Gly Leu 65 70 75 80 Gln Ala Glu Asp Glu Thr Asp Tyr Phe Cys Gln Ser Tyr Asp Ser Ser 85 90 95 Leu Ser Ala Trp Val Phe Gly Gly Gly Thr Lys Val Thr Val Leu Arg 100 105 110 <210> 149 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 149 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Pro Phe Ser Ser Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Ala Asn Gly Gly Thr Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Asn Asn Gly Asn Tyr Arg Gly Ala Phe Asp Ile Trp Gly Gln Gly 100 105 110 Thr Met Val Ile Val Ser Ser 115 <210> 150 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 150 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Ala Gly 20 25 30 Tyr Asp Val His Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu 35 40 45 Leu Ile Tyr Gly Asn Thr Asn Arg Pro Ser Gly Val Pro Asp Arg Phe 50 55 60 Ser Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ile Gly Leu 65 70 75 80 Gln Ala Asp Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr Asp Ser Thr 85 90 95 Leu Arg Val Trp Met Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly 100 105 110 <210> 151 <211> 121 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 151 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Pro Glu Phe Thr Phe Ser Lys Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Gly Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Ser Ser Arg Ser Gly Tyr Phe Leu Pro Leu Asp Tyr Trp Gly 100 105 110 Gln Gly Thr Leu Val Thr Val Ser Ser 115 120 <210> 152 <211> 109 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 152 Ser Ser Glu Leu Thr Gln Asp Pro Ala Val Ser Val Ala Leu Gly Gln 1 5 10 15 Thr Val Arg Ile Thr Cys Gln Gly Asn Ser Leu Arg Tyr Tyr Tyr Pro 20 25 30 Ser Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val Ile Tyr 35 40 45 Gly Lys Asn Asn Arg Pro Ser Gly Ile Pro Asp Arg Phe Ser Gly Ser 50 55 60 Ser Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr Gly Thr Gln Ala Glu 65 70 75 80 Asp Glu Ala Asp Tyr Tyr Cys Ser Ser Arg Asp Asn Thr Asp Asn Arg 85 90 95 Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly 100 105 <210> 153 <211> 118 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 153 Glu Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Gly Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Ala Ala Val Thr Gly Gly Phe Asp Pro Trp Gly Gln Gly Thr 100 105 110 Leu Val Thr Val Ser Ser 115 <210> 154 <211> 109 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 154 Gln Pro Gly Leu Thr Gln Pro Pro Ser Val Ser Val Ala Pro Gly Gln 1 5 10 15 Thr Ala Arg Ile Thr Cys Gly Gly Asp Asn Ile Gly Arg Lys Ser Val 20 25 30 His Trp Tyr Gln Gln Arg Pro Gly Gln Ala Pro Ile Leu Val Ile Arg 35 40 45 Asp Asp Arg Asp Arg Pro Ser Gly Ile Pro Glu Arg Phe Ser Gly Ser 50 55 60 Ser Ser Val Asn Thr Ala Thr Leu Ile Ile Ser Arg Val Glu Ala Gly 65 70 75 80 Asp Glu Ala Asp Tyr Tyr Cys Gln Val Trp Asp Ser Ser Ser Lys His 85 90 95 Tyr Val Phe Gly Pro Gly Thr Lys Val Thr Ala Leu Gly 100 105 <210> 155 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 155 Ser Ser Asn Ile Gly Ser Asn Tyr 1 5 <210> 156 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 156 Gly Gly Thr Phe Ser Ser Gln Ala 1 5 <210> 157 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 157 Tyr Ser Val Phe His Ser Pro Asn Asn Lys Asn Tyr 1 5 10 <210> 158 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 158 Gly Phe Thr Val Ser Asn Tyr Ala 1 5 <210> 159 <211> 6 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 159 Ala Leu Pro Lys Lys Tyr 1 5 <210> 160 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 160 Ser Gly Ser Ile Ala Ser Asn Tyr 1 5 <210> 161 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 161 Gly Phe Thr Val Ser Thr Ser His 1 5 <210> 162 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 162 Ser Asn Asn Val Gly Asn Gln Gly 1 5 <210> 163 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 163 Gly Phe Ile Phe Ser Asp Tyr Tyr 1 5 <210> 164 <211> 6 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 164 Gln Asp Ile Gly Thr Asp 1 5 <210> 165 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 165 Ile Ser Gly Ser Gly Gly Ser Arg 1 5 <210> 166 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 166 Ile Ile Pro Phe Phe Gly Val Pro 1 5 <210> 167 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 167 Lys Ser Gly Ser Asp Gly Arg Thr 1 5 <210> 168 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 168 Lys Asp Ser Gly Gly Lys Thr 1 5 <210> 169 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 169 Ile Arg Ser Arg Arg Gly Glu Thr 1 5 <210> 170 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 170 Ala Arg Gly Arg Gly Gly His Gly Met Asp Val 1 5 10 <210> 171 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 171 Ala Ala Trp Asp Asp Ser Leu Asn Gly Leu Val 1 5 10 <210> 172 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 172 Ala Val Leu Lys Gly Arg Gly Asn Phe Asp Phe 1 5 10 <210> 173 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 173 Gln Gln Arg Ser Asn Trp Pro Leu Thr 1 5 <210> 174 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 174 Ala Lys Gly Ile Tyr Asp Val Thr Gly Ser Ser Phe Asp Ser 1 5 10 <210> 175 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 175 Tyr Ser Thr Asp Ser Ser Gly Asn His Lys Val 1 5 10 <210> 176 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 176 Gln Ser Tyr Asp Ser Gly Asn Arg Arg Val 1 5 10 <210> 177 <211> 19 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 177 Ala Arg Ala Arg Pro Ser Asp Pro Tyr Asp Gly Ser Gly Phe Asp Ala 1 5 10 15 Phe Asp Ile <210> 178 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 178 Ser Ala Trp Asp Ser Ser Leu Ser Ala Trp Val 1 5 10 <210> 179 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 179 Ala Arg His Arg Lys Ser Phe Thr Asp Leu Asp Ala Phe Asp Leu 1 5 10 15 <210> 180 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 180 Gln His Phe Asn Asn Tyr Pro Ala Thr 1 5 <210> 181 <211> 107 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 181 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Leu Ile Ser Gly Ser Gly Gly Ser Arg Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Asn Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gly Arg Gly Gly His Gly Met Asp Val 100 105 <210> 182 <211> 100 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 182 Gln Pro Gly Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn 20 25 30 Tyr Val Tyr Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Arg Asn Asn Gln Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Gln 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Ala Ala Trp Asp Asp Ser Leu 85 90 95 Asn Gly Leu Val 100 <210> 183 <211> 107 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 183 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ser 1 5 10 15 Ser Val Lys Val Ser Cys Arg Ser Ser Gly Gly Thr Phe Ser Ser Gln 20 25 30 Ala Phe Ser Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met 35 40 45 Gly Arg Ile Ile Pro Phe Phe Gly Val Pro Thr Tyr Ala Gln Arg Phe 50 55 60 Gln Gly Arg Val Thr Ile Thr Ala Asp Lys Ser Pro Thr Thr Ala Tyr 65 70 75 80 Met Glu Leu Thr Ser Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Val Leu Lys Gly Arg Gly Asn Phe Asp Phe 100 105 <210> 184 <211> 103 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 184 Asp Ile Val Met Thr Gln Ser Pro Asp Ser Leu Ala Val Ser Leu Gly 1 5 10 15 Glu Arg Ala Thr Ile Asn Cys Lys Ser Ser Tyr Ser Val Phe His Ser 20 25 30 Pro Asn Asn Lys Asn Tyr Leu Ala Trp Tyr Gln Gln Arg Pro Gly Gln 35 40 45 Pro Pro Lys Leu Leu Ile Tyr Trp Ala Ser Thr Arg Gly Ser Gly Val 50 55 60 Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr 65 70 75 80 Ile Ser Ser Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln 85 90 95 Arg Ser Asn Trp Pro Leu Thr 100 <210> 185 <211> 110 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 185 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Gln Pro Arg Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Val Ser Asn Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Thr Lys Ser Gly Ser Asp Gly Arg Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ala Arg Asp Asn Ser Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Ala Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Gly Ile Tyr Asp Val Thr Gly Ser Ser Phe Asp Ser 100 105 110 <210> 186 <211> 98 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 186 Ser Tyr Glu Leu Thr Gln Pro Pro Ser Val Ser Val Ser Pro Gly Gln 1 5 10 15 Thr Ala Arg Ile Thr Cys Ser Gly Asp Ala Leu Pro Lys Lys Tyr Ala 20 25 30 Tyr Trp Tyr Gln Gln Lys Ser Gly Gln Ala Pro Val Leu Val Met Phe 35 40 45 Glu Asp Ser Lys Arg Pro Ser Gly Ile Pro Glu Arg Phe Ser Gly Ser 50 55 60 Ser Ser Gly Thr Met Ala Thr Leu Thr Ile Ser Gly Ala Gln Val Glu 65 70 75 80 Asp Glu Ala Asp Tyr Tyr Cys Tyr Ser Thr Asp Ser Ser Gly Asn His 85 90 95 Lys Val <210> 187 <211> 101 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 187 Asn Phe Met Leu Thr Gln Pro His Ser Val Ser Glu Ser Pro Gly Lys 1 5 10 15 Thr Val Thr Ile Ser Cys Thr Arg Ser Ser Gly Ser Ile Ala Ser Asn 20 25 30 Tyr Val Gln Trp Tyr Gln Gln Arg Pro Gly Ser Ala Pro Thr Thr Val 35 40 45 Ile Tyr Glu Asp Asn Gln Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Ile Asp Ser Ser Ser Asn Ser Ala Ser Leu Thr Ile Ser Gly 65 70 75 80 Leu Lys Thr Glu Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr Asp Ser 85 90 95 Gly Asn Arg Arg Val 100 <210> 188 <211> 114 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 188 Glu Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Val Ser Thr Ser 20 25 30 His Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Leu 35 40 45 Ser Gly Lys Asp Ser Gly Gly Lys Thr Tyr Tyr Ala Asp Ser Val Arg 50 55 60 Gly Arg Phe Thr Ile Ala Arg Asp Asp Ser Leu Asn Thr Val Phe Leu 65 70 75 80 Gln Met Asn Asn Met Arg Asp Glu Asp Ser Gly Val Tyr Tyr Cys Ala 85 90 95 Arg Ala Arg Pro Ser Asp Pro Tyr Asp Gly Ser Gly Phe Asp Ala Phe 100 105 110 Asp Ile <210> 189 <211> 100 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 189 Ser Tyr Glu Leu Thr Gln Pro Pro Ser Val Ser Lys Gly Leu Arg Gln 1 5 10 15 Thr Ala Thr Leu Thr Cys Thr Gly Asn Ser Asn Asn Val Gly Asn Gln 20 25 30 Gly Ala Ala Trp Leu Gln Gln His Gln Gly His Pro Pro Lys Leu Leu 35 40 45 Ser Tyr Arg Asn Asn Asn Arg Pro Ser Gly Ile Ser Glu Arg Phe Ser 50 55 60 Ala Ser Arg Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr Gly Leu Gln 65 70 75 80 Pro Glu Asp Glu Ala Asp Tyr Tyr Cys Ser Ala Trp Asp Ser Ser Leu 85 90 95 Ser Ala Trp Val 100 <210> 190 <211> 111 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 190 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Lys Pro Arg Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Ile Phe Ser Asp Tyr 20 25 30 Tyr Met Ser Trp Ile Arg Gln Ala Pro Gly Lys Gly Leu Gln Trp Val 35 40 45 Ala Ser Ile Arg Ser Arg Arg Gly Glu Thr Asn Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ala Arg Asp Asn Ala Glu Lys Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Ala Ala Val Tyr Tyr Cys 85 90 95 Ala Arg His Arg Lys Ser Phe Thr Asp Leu Asp Ala Phe Asp Leu 100 105 110 <210> 191 <211> 97 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 191 Asp Ile Val Met Thr Gln Ser Pro Ser Thr Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Ile Gly Thr Asp 20 25 30 Leu Ser Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Lys Ala Ser Ser Leu Glu Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Asp Asp Phe Ala Thr Tyr Tyr Cys Gln His Phe Asn Asn Tyr Pro Ala 85 90 95 Thr <210> 192 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 192 Gly Tyr Thr Phe Thr Ser Tyr Gly 1 5 <210> 193 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 193 Ile Ser Ala Tyr Asn Gly Asn Thr 1 5 <210> 194 <211> 18 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 194 Ala Arg Asp Pro Gly Leu Trp Phe Gly Leu Thr His Asp Tyr Tyr Phe 1 5 10 15 Asp Tyr <210> 195 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 195 Ser Ser Asn Ile Gly Ser Asn Thr 1 5 <210> 196 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 196 Ala Ala Trp Asp Asp Ser Arg Ser Gly Pro Val 1 5 10 <210> 197 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 197 Gly Phe Thr Phe Ser Asp Tyr Ser 1 5 <210> 198 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 198 Ile Asn Ser Asp Gly Ser Arg Thr 1 5 <210> 199 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 199 Ala Arg Gly Pro Gly Phe Phe Gly Phe Asp Ile 1 5 10 <210> 200 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 200 Arg Ser Asn Ile Gly Arg Asn Ser 1 5 <210> 201 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 201 Ala Ala Trp Asp Ala Arg Leu Thr Gly Pro Leu 1 5 10 <210> 202 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 202 Gly Tyr Ser Phe Thr Asn Tyr Trp 1 5 <210> 203 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 203 Ile Asn Pro Val Asn Ser Arg Thr 1 5 <210> 204 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 204 Ala Arg Tyr Tyr Tyr Tyr Ala Met Glu Val 1 5 10 <210> 205 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 205 Glu Ala Trp Asp Asp Ser Leu Asn Gly Pro Val 1 5 10 <210> 206 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 206 Gly Tyr Thr Phe Thr Asn Tyr Gly 1 5 <210> 207 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 207 Val Asp Asn Asn Asn Gly Asn Ile 1 5 <210> 208 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 208 Ala Arg Gly Leu Phe Ser Ser Arg Trp Tyr Leu Trp Phe Asp Pro 1 5 10 15 <210> 209 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 209 Ser Ser Asp Val Gly Gly Tyr Asn Tyr 1 5 <210> 210 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 210 Ser Ser Tyr Thr Arg Ser Ser Thr Ser Tyr Val Val 1 5 10 <210> 211 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 211 Gly Gly Thr Phe Ser Ser Tyr Ala 1 5 <210> 212 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 212 Ile Leu Pro Met Phe Gly Ser Thr 1 5 <210> 213 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 213 Ala Arg Gly Arg Asp Ile Val Ala Pro Ser Asn Ser Gly Phe Asp Val 1 5 10 15 <210> 214 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 214 Ser Ala Tyr Asp Arg Ser Leu Asn Ala Trp Val 1 5 10 <210> 215 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 215 Ile Ile Pro Ile Phe Gly Thr Ala 1 5 <210> 216 <211> 13 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 216 Ala Arg Gly Arg Gln Met Phe Gly Ala Gly Ile Asp Phe 1 5 10 <210> 217 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 217 Gly Tyr Thr Leu Ser Ser His Gly 1 5 <210> 218 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 218 Ile Ser Ala His Asn Gly His Ala 1 5 <210> 219 <211> 13 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 219 Ala Arg Val His Ala Ala Leu Tyr Tyr Gly Met Asp Val 1 5 10 <210> 220 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 220 Ser Gly Ser Ile Asp Ser Asn Tyr 1 5 <210> 221 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 221 Gln Ser Tyr Asp Ser Asn Asn Arg His Val Ile 1 5 10 <210> 222 <211> 6 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 222 Asn Ile Gly Ser Lys Gly 1 5 <210> 223 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 223 Gln Val Trp Asp Ser Gly Ser Asp His Trp Val 1 5 10 <210> 224 <211> 6 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 224 Asn Ile Gly Asp Lys Gly 1 5 <210> 225 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 225 Gln Val Trp Asp Ser Ser Ser Asp His Trp Val 1 5 10 <210> 226 <211> 6 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 226 Asn Ile Gly Asn Lys Gly 1 5 <210> 227 <211> 6 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 227 Asn Ile Gly Gly Lys Gly 1 5 <210> 228 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 228 Gly Phe Thr Phe Asp Asp Tyr Ala 1 5 <210> 229 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 229 Ile Ser Trp Asn Ser Gly Ser Ile 1 5 <210> 230 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 230 Ala Ser Asp Tyr Gly Asp Lys Tyr Tyr Tyr Tyr Gly Met Asp Val 1 5 10 15 <210> 231 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 231 Gly Tyr Thr Phe Thr Thr Tyr Trp 1 5 <210> 232 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 232 Ile Tyr Pro Asp Asp Ser Asp Thr 1 5 <210> 233 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 233 Ala Phe Trp Gly Ala Ser Gly Ala Pro Val Asn Gly Phe Asp Ile 1 5 10 15 <210> 234 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 234 Gly Asp Ser Val Ser Ser Asp Asn Tyr Phe 1 5 10 <210> 235 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 235 Val Tyr Tyr Asn Gly Asn Thr 1 5 <210> 236 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 236 Ala Thr Glu Thr Pro Pro Thr Ser Tyr Phe Asn Ser Gly Pro Phe Asp 1 5 10 15 Ser <210> 237 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 237 Gly Tyr Thr Phe Asn Arg Phe Gly 1 5 <210> 238 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 238 Thr Asn Pro Tyr Asn Gly Asn Thr 1 5 <210> 239 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 239 Ala Arg Val Val Ala Val Asn Gly Met Asp Val 1 5 10 <210> 240 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 240 Ile Ser Tyr Asp Gly Ser Asn Lys 1 5 <210> 241 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 241 Ala Ser Gln Thr Val Ala Gly Ser Asp Tyr 1 5 10 <210> 242 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 242 Ala Ser Asp Tyr Gly Asp Lys Tyr Ser Tyr Tyr Gly Met Asp Val 1 5 10 15 <210> 243 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 243 Gly Phe Thr Phe Asp Asp Phe Ala 1 5 <210> 244 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 244 Ala Ala Trp Asp Gly Gly Leu Asn Gly Arg Gly Val 1 5 10 <210> 245 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 245 Ser Ser Asn Ile Gly Ala Gly Tyr Val 1 5 <210> 246 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 246 Ala Ala Trp Asp Asp Ser Leu Asn Ala Pro Val 1 5 10 <210> 247 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 247 Ser Asn Asn Val Gly Ala His Gly 1 5 <210> 248 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 248 Ser Ser Trp Asp Ser Ser Leu Ser Gly Tyr Val 1 5 10 <210> 249 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 249 Ser Gly Ser Ile Ala Ala Tyr Tyr 1 5 <210> 250 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 250 Gln Ser Tyr Asp Ser Ser Asn Leu Trp Val 1 5 10 <210> 251 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 251 Gln Val Trp His Ser Val Ser Asp Gln Gly Val 1 5 10 <210> 252 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 252 Trp Ile Asn Pro Gly Asn Val Asn Thr Lys Tyr Asn Glu Lys Phe Lys 1 5 10 15 Gly <210> 253 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 253 Lys Ser Ser Gln Ser Ile Leu Tyr Ser Ser Asn Gln Lys Asn Tyr Leu 1 5 10 15 Ala <210> 254 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 254 Trp Ala Ser Thr Arg Glu Ser 1 5 <210> 255 <211> 118 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 255 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Ala Ser Tyr 20 25 30 Tyr Met His Trp Met Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile 35 40 45 Gly Trp Ile Asn Pro Gly Asn Val Asn Thr Lys Tyr Asn Glu Lys Phe 50 55 60 Lys Gly Arg Ala Thr Leu Thr Val Asp Thr Ser Thr Asn Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Ser Thr Tyr Tyr Arg Pro Leu Asp Tyr Trp Gly Gln Gly Thr 100 105 110 Leu Val Thr Val Ser Ser 115 <210> 256 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 256 Asp Ile Val Met Thr Gln Ser Pro Asp Ser Leu Ala Val Ser Leu Gly 1 5 10 15 Glu Arg Ala Thr Ile Asn Cys Lys Ser Ser Gln Ser Ile Leu Tyr Ser 20 25 30 Ser Asn Gln Lys Asn Tyr Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln 35 40 45 Ser Pro Lys Leu Leu Ile Tyr Trp Ala Ser Thr Arg Glu Ser Gly Val 50 55 60 Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr 65 70 75 80 Ile Ser Ser Leu Gln Ala Glu Asp Val Ala Val Tyr Tyr Cys His Gln 85 90 95 Tyr Leu Ser Ser Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 <210> 257 <211> 118 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 257 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Ala Ser Gln 20 25 30 Trp Met His Trp Met Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile 35 40 45 Gly Trp Ile Asn Pro Gly Asn Val Asn Thr Lys Tyr Asn Glu Lys Phe 50 55 60 Lys Gly Arg Ala Thr Leu Thr Val Asp Thr Ser Thr Asn Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Ser Thr Trp Tyr Arg Pro Leu Asp Tyr Trp Gly Gln Gly Thr 100 105 110 Leu Val Thr Val Ser Ser 115 <210> 258 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 258 Asp Ile Val Met Thr Gln Ser Pro Asp Ser Leu Ala Val Ser Leu Gly 1 5 10 15 Glu Arg Ala Thr Ile Asn Cys Lys Ser Ser Gln Ser Ile Leu Tyr Ser 20 25 30 Ser Asn Gln Lys Asn Tyr Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln 35 40 45 Ser Pro Lys Leu Leu Ile Tyr Trp Ala Ser Thr Arg Glu Ser Gly Val 50 55 60 Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr 65 70 75 80 Ile Ser Ser Leu Gln Ala Glu Asp Val Ala Val Tyr Tyr Cys His Gln 85 90 95 Tyr Ile Ser Ser Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 <210> 259 <211> 118 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 259 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Ala Ser Ser 20 25 30 Trp Met His Trp Met Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile 35 40 45 Gly Trp Ile Asn Pro Gly Asn Val Asn Thr Lys Tyr Asn Glu Lys Phe 50 55 60 Lys Gly Arg Ala Thr Leu Thr Val Asp Thr Ser Thr Asn Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Ser Thr Trp Tyr Arg Pro Asn Asp Tyr Trp Gly Gln Gly Thr 100 105 110 Leu Val Thr Val Ser Ser 115 <210> 260 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 260 Asp Ile Val Met Thr Gln Ser Pro Asp Ser Leu Ala Val Ser Leu Gly 1 5 10 15 Glu Arg Ala Thr Ile Asn Cys Lys Ser Ser Gln Ser Ile Leu Tyr Ser 20 25 30 Ser Asn Gln Lys Asn Tyr Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln 35 40 45 Ser Pro Lys Leu Leu Ile Tyr Trp Ala Ser Thr Arg Glu Ser Gly Val 50 55 60 Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr 65 70 75 80 Ile Ser Ser Leu Gln Ala Glu Asp Val Ala Val Tyr Tyr Cys His Gln 85 90 95 Tyr Lys Ser Ser Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 <210> 261 <211> 118 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 261 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Ala Ser Ser 20 25 30 Trp Met His Trp Met Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile 35 40 45 Gly Trp Ile Asn Pro Gly Asn Val Asn Thr Lys Tyr Asn Glu Lys Phe 50 55 60 Lys Gly Arg Ala Thr Leu Thr Val Asp Thr Ser Thr Asn Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Thr Thr Arg Tyr Arg Pro Leu Asp Tyr Trp Gly Gln Gly Thr 100 105 110 Leu Val Thr Val Ser Ser 115 <210> 262 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 262 Asp Ile Val Met Thr Gln Ser Pro Asp Ser Leu Ala Val Ser Leu Gly 1 5 10 15 Glu Arg Ala Thr Ile Asn Cys Lys Ser Ser Gln Ser Ile Leu Tyr Ser 20 25 30 Ser Asn Gln Lys Asn Tyr Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln 35 40 45 Ser Pro Lys Leu Leu Ile Tyr Trp Ala Ser Thr Arg Glu Ser Gly Val 50 55 60 Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr 65 70 75 80 Ile Ser Ser Leu Gln Ala Glu Asp Val Ala Val Tyr Tyr Cys His Gln 85 90 95 Tyr Arg Ser Ser Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 <210> 263 <211> 118 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 263 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Ala Ser Gln 20 25 30 Tyr Met His Trp Met Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile 35 40 45 Gly Trp Ile Asn Pro Gly Asn Val Asn Thr Lys Tyr Asn Glu Lys Phe 50 55 60 Lys Gly Arg Ala Thr Leu Thr Val Asp Thr Ser Thr Asn Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Leu Thr Tyr Tyr Arg Pro Pro Asp Tyr Trp Gly Gln Gly Thr 100 105 110 Leu Val Thr Val Ser Ser 115 <210> 264 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 264 Asp Ile Val Met Thr Gln Ser Pro Asp Ser Leu Ala Val Ser Leu Gly 1 5 10 15 Glu Arg Ala Thr Ile Asn Cys Lys Ser Ser Gln Ser Ile Leu Tyr Ser 20 25 30 Ser Asn Gln Lys Asn Tyr Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln 35 40 45 Ser Pro Lys Leu Leu Ile Tyr Trp Ala Ser Thr Arg Glu Ser Gly Val 50 55 60 Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr 65 70 75 80 Ile Ser Ser Leu Gln Ala Glu Asp Val Ala Val Tyr Tyr Cys His Gln 85 90 95 Tyr Tyr Ser Ser Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 <210> 265 <211> 118 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 265 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Ala Ser Ala 20 25 30 Trp Met His Trp Met Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile 35 40 45 Gly Trp Ile Asn Pro Gly Asn Val Asn Thr Lys Tyr Asn Glu Lys Phe 50 55 60 Lys Gly Arg Ala Thr Leu Thr Val Asp Thr Ser Thr Asn Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Ser Thr Tyr Tyr Arg Pro Leu Asp Tyr Trp Gly Gln Gly Thr 100 105 110 Leu Val Thr Val Ser Ser 115 <210> 266 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 266 Asp Ile Val Met Thr Gln Ser Pro Asp Ser Leu Ala Val Ser Leu Gly 1 5 10 15 Glu Arg Ala Thr Ile Asn Cys Lys Ser Ser Gln Ser Ile Leu Tyr Ser 20 25 30 Ser Asn Gln Lys Asn Tyr Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln 35 40 45 Ser Pro Lys Leu Leu Ile Tyr Trp Ala Ser Thr Arg Glu Ser Gly Val 50 55 60 Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr 65 70 75 80 Ile Ser Ser Leu Gln Ala Glu Asp Val Ala Val Tyr Tyr Cys His Gln 85 90 95 Tyr Met Ser Ser Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 <210> 267 <211> 5 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 267 Gly Gly Gly Gly Ser 1 5 <210> 268 <211> 121 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 268 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Asn Gly Asn Tyr Arg Gly Ser Leu Ala Phe Asp Ile Trp Gly 100 105 110 Gln Gly Thr Leu Val Thr Val Ser Ser 115 120 <210> 269 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 269 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Glu Phe Thr Phe Gly Thr Tyr 20 25 30 Ala Met Thr Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Val Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Arg Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gly Pro Val Leu Arg Tyr Gly Phe Asp Ile Trp Gly Gln Gly 100 105 110 Thr Met Val Thr Val Ser Ser 115 <210> 270 <211> 113 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 270 Gln Ser Val Leu Thr Leu Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Ala Gly 20 25 30 Tyr Asp Val His Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu 35 40 45 Leu Ile Tyr Gly Asn Asn Asn Arg Pro Ser Gly Val Pro Asp Arg Phe 50 55 60 Ser Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Thr Gly Leu 65 70 75 80 Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr Asp Ser Ser 85 90 95 Leu Ser Ala Trp Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu 100 105 110 Gly <210> 271 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 271 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Arg Gly 20 25 30 Tyr Asn Val His Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu 35 40 45 Leu Ile Tyr Asp Asp Thr Asn Arg Pro Ser Gly Val Pro His Arg Phe 50 55 60 Ser Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Thr Gly Leu 65 70 75 80 Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr Asp Ser Ser 85 90 95 Leu Arg Ala Trp Val Phe Gly Gly Gly Thr Lys Leu Ala Val Leu Gly 100 105 110 <210> 272 <211> 110 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 272 Leu Pro Val Leu Thr Gln Pro Pro Ser Val Ser Val Ala Pro Gly Gln 1 5 10 15 Thr Ala Arg Ile Thr Cys Gly Gly Asn Asn Ile Gly Ser Lys Ser Val 20 25 30 His Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val Ile Tyr 35 40 45 Tyr Asp Ser Asp Arg Pro Ser Gly Ile Pro Glu Arg Phe Ser Gly Ser 50 55 60 Asn Ser Gly Asn Thr Ala Thr Leu Thr Ile Ser Arg Val Glu Ala Gly 65 70 75 80 Asp Glu Ala Asp Tyr Tyr Cys Gln Val Trp Asp Ser Ser Ser Asp His 85 90 95 His Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly 100 105 110 <210> 273 <211> 459 <212> PRT <213> Homo sapiens <400> 273 Met Ala Pro Leu Cys Pro Ser Pro Trp Leu Pro Leu Leu Ile Pro Ala 1 5 10 15 Pro Ala Pro Gly Leu Thr Val Gln Leu Leu Leu Ser Leu Leu Leu Leu 20 25 30 Met Pro Val His Pro Gln Arg Leu Pro Arg Met Gln Glu Asp Ser Pro 35 40 45 Leu Gly Gly Gly Ser Ser Gly Glu Asp Asp Pro Leu Gly Glu Glu Asp 50 55 60 Leu Pro Ser Glu Glu Asp Ser Pro Arg Glu Glu Asp Pro Pro Gly Glu 65 70 75 80 Glu Asp Leu Pro Gly Glu Glu Asp Leu Pro Gly Glu Glu Asp Leu Pro 85 90 95 Glu Val Lys Pro Lys Ser Glu Glu Glu Gly Ser Leu Lys Leu Glu Asp 100 105 110 Leu Pro Thr Val Glu Ala Pro Gly Asp Pro Gln Glu Pro Gln Asn Asn 115 120 125 Ala His Arg Asp Lys Glu Gly Asp Asp Gln Ser His Trp Arg Tyr Gly 130 135 140 Gly Asp Pro Pro Trp Pro Arg Val Ser Pro Ala Cys Ala Gly Arg Phe 145 150 155 160 Gln Ser Pro Val Asp Ile Arg Pro Gln Leu Ala Ala Phe Cys Pro Ala 165 170 175 Leu Arg Pro Leu Glu Leu Leu Gly Phe Gln Leu Pro Pro Leu Pro Glu 180 185 190 Leu Arg Leu Arg Asn Asn Gly His Ser Val Gln Leu Thr Leu Pro Pro 195 200 205 Gly Leu Glu Met Ala Leu Gly Pro Gly Arg Glu Tyr Arg Ala Leu Gln 210 215 220 Leu His Leu His Trp Gly Ala Ala Gly Arg Pro Gly Ser Glu His Thr 225 230 235 240 Val Glu Gly His Arg Phe Pro Ala Glu Ile His Val Val His Leu Ser 245 250 255 Thr Ala Phe Ala Arg Val Asp Glu Ala Leu Gly Arg Pro Gly Gly Leu 260 265 270 Ala Val Leu Ala Ala Phe Leu Glu Glu Gly Pro Glu Glu Asn Ser Ala 275 280 285 Tyr Glu Gln Leu Leu Ser Arg Leu Glu Glu Ile Ala Glu Glu Gly Ser 290 295 300 Glu Thr Gln Val Pro Gly Leu Asp Ile Ser Ala Leu Leu Pro Ser Asp 305 310 315 320 Phe Ser Arg Tyr Phe Gln Tyr Glu Gly Ser Leu Thr Thr Pro Pro Cys 325 330 335 Ala Gln Gly Val Ile Trp Thr Val Phe Asn Gln Thr Val Met Leu Ser 340 345 350 Ala Lys Gln Leu His Thr Leu Ser Asp Thr Leu Trp Gly Pro Gly Asp 355 360 365 Ser Arg Leu Gln Leu Asn Phe Arg Ala Thr Gln Pro Leu Asn Gly Arg 370 375 380 Val Ile Glu Ala Ser Phe Pro Ala Gly Val Asp Ser Ser Pro Arg Ala 385 390 395 400 Ala Glu Pro Val Gln Leu Asn Ser Cys Leu Ala Ala Gly Asp Ile Leu 405 410 415 Ala Leu Val Phe Gly Leu Leu Phe Ala Val Thr Ser Val Ala Phe Leu 420 425 430 Val Gln Met Arg Arg Gln His Arg Arg Gly Thr Lys Gly Gly Val Ser 435 440 445 Tyr Arg Pro Ala Glu Val Ala Glu Thr Gly Ala 450 455 <210> 274 <211> 437 <212> PRT <213> Mus sp. <400> 274 Met Ala Ser Leu Gly Pro Ser Pro Trp Ala Pro Leu Ser Thr Pro Ala 1 5 10 15 Pro Thr Ala Gln Leu Leu Leu Phe Leu Leu Leu Gln Val Ser Ala Gln 20 25 30 Pro Gln Gly Leu Ser Gly Met Gln Gly Glu Pro Ser Leu Gly Asp Ser 35 40 45 Ser Ser Gly Glu Asp Glu Leu Gly Val Asp Val Leu Pro Ser Glu Glu 50 55 60 Asp Ala Pro Glu Glu Ala Asp Pro Pro Asp Gly Glu Asp Pro Pro Glu 65 70 75 80 Val Asn Ser Glu Asp Arg Met Glu Glu Ser Leu Gly Leu Glu Asp Leu 85 90 95 Ser Thr Pro Glu Ala Pro Glu His Ser Gln Gly Ser His Gly Asp Glu 100 105 110 Lys Gly Gly Gly His Ser His Trp Ser Tyr Gly Gly Thr Leu Leu Trp 115 120 125 Pro Gln Val Ser Pro Ala Cys Ala Gly Arg Phe Gln Ser Pro Val Asp 130 135 140 Ile Arg Leu Glu Arg Thr Ala Phe Cys Arg Thr Leu Gln Pro Leu Glu 145 150 155 160 Leu Leu Gly Tyr Glu Leu Gln Pro Leu Pro Glu Leu Ser Leu Ser Asn 165 170 175 Asn Gly His Thr Val Gln Leu Thr Leu Pro Pro Gly Leu Lys Met Ala 180 185 190 Leu Gly Pro Gly Gln Glu Tyr Arg Ala Leu Gln Leu His Leu His Trp 195 200 205 Gly Thr Ser Asp His Pro Gly Ser Glu His Thr Val Asn Gly His Arg 210 215 220 Phe Pro Ala Glu Ile His Val Val His Leu Ser Thr Ala Phe Ser Glu 225 230 235 240 Leu His Glu Ala Leu Gly Arg Pro Gly Gly Leu Ala Val Leu Ala Ala 245 250 255 Phe Leu Gln Glu Ser Pro Glu Glu Asn Ser Ala Tyr Glu Gln Leu Leu 260 265 270 Ser His Leu Glu Glu Ile Ser Glu Glu Gly Ser Lys Ile Glu Ile Pro 275 280 285 Gly Leu Asp Val Ser Ala Leu Leu Pro Ser Asp Phe Ser Arg Tyr Tyr 290 295 300 Arg Tyr Glu Gly Ser Leu Thr Thr Pro Pro Cys Ser Gln Gly Val Ile 305 310 315 320 Trp Thr Val Phe Asn Glu Thr Val Lys Leu Ser Ala Lys Gln Leu His 325 330 335 Thr Leu Ser Val Ser Leu Trp Gly Pro Arg Asp Ser Arg Leu Gln Leu 340 345 350 Asn Phe Arg Ala Thr Gln Pro Leu Asn Gly Arg Thr Ile Glu Ala Ser 355 360 365 Phe Pro Ala Ala Glu Asp Ser Ser Pro Glu Pro Val His Val Asn Ser 370 375 380 Cys Phe Thr Ala Gly Asp Ile Leu Ala Leu Val Phe Gly Leu Leu Phe 385 390 395 400 Ala Val Thr Ser Ile Ala Phe Leu Leu Gln Leu Arg Arg Gln His Arg 405 410 415 His Arg Ser Gly Thr Lys Asp Arg Val Ser Tyr Ser Pro Ala Glu Met 420 425 430 Thr Glu Thr Gly Ala 435 <210> 275 <211> 98 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 275 Gln Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Lys Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Tyr 20 25 30 Tyr Met Ser Trp Ile Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Tyr Ile Ser Ser Ser Ser Ser Tyr Thr Asn Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg <210> 276 <211> 95 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 276 Asp Ile Gln Met Thr Gln Ser Pro Ser Thr Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Ser Ile Ser Ser Trp 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Lys Ala Ser Ser Leu Glu Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Glu Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Asp Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Asn Ser Tyr Ser 85 90 95 <210> 277 <211> 98 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 277 Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys <210> 278 <211> 98 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 278 Asn Phe Met Leu Thr Gln Pro His Ser Val Ser Glu Ser Pro Gly Lys 1 5 10 15 Thr Val Thr Ile Ser Cys Thr Arg Ser Ser Gly Ser Ile Ala Ser Asn 20 25 30 Tyr Val Gln Trp Tyr Gln Gln Arg Pro Gly Ser Ser Pro Thr Thr Val 35 40 45 Ile Tyr Glu Asp Asn Gln Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Ile Asp Ser Ser Ser Asn Ser Ala Ser Leu Thr Ile Ser Gly 65 70 75 80 Leu Lys Thr Glu Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr Asp Ser 85 90 95 Ser Asn <210> 279 <211> 98 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 279 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ser 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Gly Thr Phe Ser Ser Tyr 20 25 30 Ala Ile Ser Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met 35 40 45 Gly Gly Ile Ile Pro Ile Phe Gly Thr Ala Asn Tyr Ala Gln Lys Phe 50 55 60 Gln Gly Arg Val Thr Ile Thr Ala Asp Lys Ser Thr Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg <210> 280 <211> 101 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 280 Asp Ile Val Met Thr Gln Ser Pro Asp Ser Leu Ala Val Ser Leu Gly 1 5 10 15 Glu Arg Ala Thr Ile Asn Cys Lys Ser Ser Gln Ser Val Leu Tyr Ser 20 25 30 Ser Asn Asn Lys Asn Tyr Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln 35 40 45 Pro Pro Lys Leu Leu Ile Tyr Trp Ala Ser Thr Arg Glu Ser Gly Val 50 55 60 Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr 65 70 75 80 Ile Ser Ser Leu Gln Ala Glu Asp Val Ala Val Tyr Tyr Cys Gln Gln 85 90 95 Tyr Tyr Ser Thr Pro 100 <210> 281 <211> 98 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 281 Gln Ser Val Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn 20 25 30 Tyr Val Tyr Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Arg Asn Asn Gln Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Arg 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Ala Ala Trp Asp Asp Ser Leu 85 90 95 Ser Gly <210> 282 <211> 96 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 282 Ser Tyr Glu Leu Thr Gln Pro Pro Ser Val Ser Val Ser Pro Gly Gln 1 5 10 15 Thr Ala Arg Ile Thr Cys Ser Gly Asp Ala Leu Pro Lys Lys Tyr Ala 20 25 30 Tyr Trp Tyr Gln Gln Lys Ser Gly Gln Ala Pro Val Leu Val Ile Tyr 35 40 45 Glu Asp Ser Lys Arg Pro Ser Gly Ile Pro Glu Arg Phe Ser Gly Ser 50 55 60 Ser Ser Gly Thr Met Ala Thr Leu Thr Ile Ser Gly Ala Gln Val Glu 65 70 75 80 Asp Glu Ala Asp Tyr Tyr Cys Tyr Ser Thr Asp Ser Ser Gly Asn His 85 90 95 <210> 283 <211> 97 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 283 Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Val Ser Ser Asn 20 25 30 Tyr Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Val Ile Tyr Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys 50 55 60 Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu 65 70 75 80 Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala 85 90 95 Arg <210> 284 <211> 98 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 284 Gln Ala Gly Leu Thr Gln Pro Pro Ser Val Ser Lys Gly Leu Arg Gln 1 5 10 15 Thr Ala Thr Leu Thr Cys Thr Gly Asn Ser Asn Asn Val Gly Asn Gln 20 25 30 Gly Ala Ala Trp Leu Gln Gln His Gln Gly His Pro Pro Lys Leu Leu 35 40 45 Ser Tyr Arg Asn Asn Asn Arg Pro Ser Gly Ile Ser Glu Arg Leu Ser 50 55 60 Ala Ser Arg Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr Gly Leu Gln 65 70 75 80 Pro Glu Asp Glu Ala Asp Tyr Tyr Cys Ser Ala Trp Asp Ser Ser Leu 85 90 95 Ser Ala <210> 285 <211> 109 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 285 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ser 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Gly Thr Phe Ser Ser Tyr 20 25 30 Ala Ile Ser Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met 35 40 45 Gly Gly Ile Ile Pro Ile Phe Gly Thr Ala Asn Tyr Ala Gln Lys Phe 50 55 60 Gln Gly Arg Val Thr Ile Thr Ala Asp Lys Ser Thr Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gly Arg Gln Met Phe Gly Ala Gly Ile Asp Phe 100 105 <210> 286 <211> 98 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 286 Asn Phe Met Leu Thr Gln Pro His Ser Val Ser Glu Ser Pro Gly Lys 1 5 10 15 Thr Val Thr Ile Ser Cys Thr Gly Ser Ser Gly Ser Ile Ala Ser Asn 20 25 30 Tyr Val Gln Trp Tyr Gln Gln Arg Pro Gly Ser Ala Pro Thr Thr Val 35 40 45 Ile Tyr Glu Asp Asn Gln Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Ile Asp Ser Ser Ser Asn Ser Ala Ser Leu Thr Ile Ser Gly 65 70 75 80 Leu Lys Thr Glu Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr Asp Ser 85 90 95 Ser Asn <210> 287 <211> 102 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 287 Asn Phe Met Leu Thr Gln Pro His Ser Val Ser Glu Ser Pro Gly Lys 1 5 10 15 Thr Val Thr Ile Ser Cys Thr Arg Ser Ser Gly Ser Ile Asp Ser Asn 20 25 30 Tyr Val Gln Trp Tyr Gln Gln Arg Pro Gly Ser Ala Pro Thr Thr Val 35 40 45 Ile Tyr Glu Asp Asn Gln Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Ile Asp Ser Ser Ser Asn Ser Ala Ser Leu Thr Ile Ser Gly 65 70 75 80 Leu Lys Thr Glu Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr Asp Ser 85 90 95 Asn Asn Arg His Val Ile 100 <210> 288 <211> 99 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 288 Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Arg 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Asp Asp Tyr 20 25 30 Ala Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Gly Ile Ser Trp Asn Ser Gly Ser Ile Gly Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Leu Tyr Tyr Cys 85 90 95 Ala Lys Asp <210> 289 <211> 111 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 289 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Gln Pro Gly Arg 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Asp Asp Phe 20 25 30 Ala Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Gly Ile Ser Trp Asn Ser Gly Ser Ile Gly Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Val Ser Arg Asp Asn Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Ser Asp Tyr Gly Asp Lys Tyr Ser Tyr Tyr Gly Met Asp Val 100 105 110 <210> 290 <211> 98 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 290 Gln Ser Val Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn 20 25 30 Thr Val Asn Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Ser Asn Asn Gln Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Gln 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Ala Ala Trp Asp Asp Ser Leu 85 90 95 Asn Gly <210> 291 <211> 101 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 291 Gln Pro Gly Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn 20 25 30 Thr Val Asn Trp Tyr Gln Gln Phe Pro Gly Lys Ala Pro Lys Leu Leu 35 40 45 Ile Phe Asp Asp Asn Gln Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Ala Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Gln 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Ala Ala Trp Asp Gly Gly Leu 85 90 95 Asn Gly Arg Gly Val 100 <210> 292 <211> 99 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 292 Gln Ser Val Leu Thr Gln Pro Pro Ser Val Ser Gly Ala Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Ala Gly 20 25 30 Tyr Asp Val His Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu 35 40 45 Leu Ile Tyr Gly Asn Ser Asn Arg Pro Ser Gly Val Pro Asp Arg Phe 50 55 60 Ser Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Thr Gly Leu 65 70 75 80 Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr Asp Ser Ser 85 90 95 Leu Ser Gly <210> 293 <211> 98 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 293 Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Ala Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Val Ile Ser Tyr Asp Gly Ser Asn Lys Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg <210> 294 <211> 96 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 294 Ser Tyr Val Leu Thr Gln Pro Pro Ser Val Ser Val Ala Pro Gly Gln 1 5 10 15 Thr Ala Arg Ile Thr Cys Gly Gly Asn Asn Ile Gly Ser Lys Ser Val 20 25 30 His Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val Val Tyr 35 40 45 Asp Asp Ser Asp Arg Pro Ser Gly Ile Pro Glu Arg Phe Ser Gly Ser 50 55 60 Asn Ser Gly Asn Thr Ala Thr Leu Thr Ile Ser Arg Val Glu Ala Gly 65 70 75 80 Asp Glu Ala Asp Tyr Tyr Cys Gln Val Trp Asp Ser Ser Ser Asp His 85 90 95 <210> 295 <211> 98 <212> PRT <213> Homo sapiens <400> 295 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr 20 25 30 Gly Ile Ser Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met 35 40 45 Gly Trp Ile Ser Ala Tyr Asn Gly Asn Thr Asn Tyr Ala Gln Lys Leu 50 55 60 Gln Gly Arg Val Thr Met Thr Thr Asp Thr Ser Thr Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Arg Ser Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg <210> 296 <211> 125 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 296 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr 20 25 30 Gly Ile Ser Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met 35 40 45 Gly Trp Ile Ser Ala Tyr Asn Gly Asn Thr Asn Tyr Ala Gln Lys Leu 50 55 60 Gln Gly Arg Val Thr Met Thr Thr Asp Thr Ser Thr Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Arg Ser Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Asp Pro Gly Leu Trp Phe Gly Leu Thr His Asp Tyr Tyr Phe 100 105 110 Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser 115 120 125 <210> 297 <211> 110 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 297 Ser Tyr Glu Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn 20 25 30 Thr Val Ser Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Arg Asn Asn Gln Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Asn Gly Leu Gln 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Ala Ala Trp Asp Asp Ser Arg 85 90 95 Ser Gly Pro Val Phe Gly Gly Gly Thr Arg Leu Thr Val Leu 100 105 110 <210> 298 <211> 98 <212> PRT <213> Homo sapiens <400> 298 Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Trp Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Val Trp Val 35 40 45 Ser Arg Ile Asn Ser Asp Gly Ser Ser Thr Ser Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg <210> 299 <211> 118 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 299 Glu Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Lys Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Glu Ala Ser Gly Phe Thr Phe Ser Asp Tyr 20 25 30 Ser Met Ser Trp Ile Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Arg Ile Asn Ser Asp Gly Ser Arg Thr Asn Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Ala Arg Gly Pro Gly Phe Phe Gly Phe Asp Ile Trp Gly Gln Gly Thr 100 105 110 Leu Val Thr Val Ser Ser 115 <210> 300 <211> 110 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 300 Ser Tyr Glu Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Arg Ser Asn Ile Gly Arg Asn 20 25 30 Ser Val Asn Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Ser Asn Asn Gln Arg Pro Ser Gly Val Pro Gly Arg Phe Ser 50 55 60 Gly Ser Arg Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Gln 65 70 75 80 Ser Glu Asp Glu Thr Asp Tyr Tyr Cys Ala Ala Trp Asp Ala Arg Leu 85 90 95 Thr Gly Pro Leu Phe Gly Gly Gly Thr Lys Leu Ser Val Leu 100 105 110 <210> 301 <211> 98 <212> PRT <213> Homo sapiens <400> 301 Glu Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Glu 1 5 10 15 Ser Leu Lys Ile Ser Cys Lys Gly Ser Gly Tyr Ser Phe Thr Ser Tyr 20 25 30 Trp Ile Gly Trp Val Arg Gln Met Pro Gly Lys Gly Leu Glu Trp Met 35 40 45 Gly Ile Ile Tyr Pro Gly Asp Ser Asp Thr Arg Tyr Ser Pro Ser Phe 50 55 60 Gln Gly Gln Val Thr Ile Ser Ala Asp Lys Ser Ile Ser Thr Ala Tyr 65 70 75 80 Leu Gln Trp Ser Ser Leu Lys Ala Ser Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Ala Arg <210> 302 <211> 117 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 302 Glu Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Glu 1 5 10 15 Ser Leu Lys Ile Ser Cys Lys Gly Ser Gly Tyr Ser Phe Thr Asn Tyr 20 25 30 Trp Ile Gly Trp Val Arg Gln Met Pro Gly Lys Gly Leu Glu Trp Met 35 40 45 Gly Ile Ile Asn Pro Val Asn Ser Arg Thr Ile Tyr Ser Pro Ser Phe 50 55 60 Gln Gly Gln Val Thr Ile Ser Val Asp Lys Ser Val Thr Thr Ala Tyr 65 70 75 80 Leu Gln Trp Ser Ser Leu Lys Ala Ser Asp Thr Ala Met Tyr Tyr Cys 85 90 95 Ala Arg Tyr Tyr Tyr Tyr Ala Met Glu Val Trp Gly Arg Gly Thr Leu 100 105 110 Val Thr Val Ser Ser 115 <210> 303 <211> 110 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 303 Leu Pro Val Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn 20 25 30 Thr Val Asn Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Arg Asn Asn Gln Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Thr Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Gln 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Glu Ala Trp Asp Asp Ser Leu 85 90 95 Asn Gly Pro Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu 100 105 110 <210> 304 <211> 122 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 304 Glu Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Asn Tyr 20 25 30 Gly Ile Ser Trp Val Arg Gln Ala Pro Gly Gly Gly Leu Glu Trp Met 35 40 45 Gly Trp Val Asp Asn Asn Asn Gly Asn Ile Asn Tyr Ala Gln Lys Phe 50 55 60 Leu Gly Arg Val Thr Met Thr Thr Asp Thr Ser Thr Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Arg Ser Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gly Leu Phe Ser Ser Arg Trp Tyr Leu Trp Phe Asp Pro Trp 100 105 110 Gly Gln Gly Thr Leu Val Thr Val Ser Ser 115 120 <210> 305 <211> 99 <212> PRT <213> Homo sapiens <400> 305 Gln Ser Ala Leu Thr Gln Pro Ala Ser Val Ser Gly Ser Pro Gly Gln 1 5 10 15 Ser Ile Thr Ile Ser Cys Thr Gly Thr Ser Ser Asp Val Gly Gly Tyr 20 25 30 Asn Tyr Val Ser Trp Tyr Gln Gln His Pro Gly Lys Ala Pro Lys Leu 35 40 45 Met Ile Tyr Glu Val Ser Asn Arg Pro Ser Gly Val Ser Asn Arg Phe 50 55 60 Ser Gly Ser Lys Ser Gly Asn Thr Ala Ser Leu Thr Ile Ser Gly Leu 65 70 75 80 Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Ser Ser Tyr Thr Ser Ser 85 90 95 Ser Thr Leu <210> 306 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 306 Gln Ser Ala Leu Thr Gln Pro Ala Ser Val Ser Gly Ser Pro Gly Gln 1 5 10 15 Ser Ile Thr Ile Ser Cys Thr Gly Thr Ser Ser Asp Val Gly Gly Tyr 20 25 30 Asn Tyr Val Ser Trp Tyr Gln Gln His Pro Gly Lys Ala Pro Lys Leu 35 40 45 Met Ile Tyr Glu Val Thr Glu Arg Pro Ser Gly Val Ser Asn Arg Phe 50 55 60 Ser Gly Ser Lys Ser Gly Asn Thr Ala Ser Leu Thr Ile Ser Gly Leu 65 70 75 80 Gln Ala Glu Asp Glu Gly Asp Tyr Tyr Cys Ser Ser Tyr Thr Arg Ser 85 90 95 Ser Thr Ser Tyr Val Val Phe Gly Gly Gly Thr Lys Val Thr Val Leu 100 105 110 <210> 307 <211> 98 <212> PRT <213> Homo sapiens <400> 307 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ser 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Gly Thr Phe Ser Ser Tyr 20 25 30 Ala Ile Ser Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met 35 40 45 Gly Gly Ile Ile Pro Ile Phe Gly Thr Ala Asn Tyr Ala Gln Lys Phe 50 55 60 Gln Gly Arg Val Thr Ile Thr Ala Asp Glu Ser Thr Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg <210> 308 <211> 123 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 308 Gln Val Gln Leu Gln Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ser 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Gly Thr Phe Ser Ser Tyr 20 25 30 Ala Ile Ser Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met 35 40 45 Gly Gly Ile Leu Pro Met Phe Gly Ser Thr Asn Tyr Ala Gln Lys Phe 50 55 60 Gln Gly Arg Leu Thr Leu Ile Ala Asp Glu Ser Thr Arg Thr Val Tyr 65 70 75 80 Leu Glu Leu Asn Ser Leu Thr Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gly Arg Asp Ile Val Ala Pro Ser Asn Ser Gly Phe Asp Val 100 105 110 Trp Gly Gln Gly Thr Thr Val Thr Val Ser Ser 115 120 <210> 309 <211> 110 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 309 Ser Tyr Glu Leu Thr Gln Pro Pro Ser Val Ser Lys Gly Leu Arg Gln 1 5 10 15 Thr Ala Thr Leu Thr Cys Thr Gly Asn Ser Asn Asn Val Gly Asn Gln 20 25 30 Gly Ala Ala Trp Leu Gln Gln His Gln Gly His Pro Pro Lys Leu Leu 35 40 45 Ser Tyr Arg Asn Asp Asn Arg Pro Ser Gly Ile Ser Glu Arg Phe Ser 50 55 60 Ala Ser Arg Ser Gly Asn Thr Ala Ser Leu Thr Ile Ser Gly Leu Gln 65 70 75 80 Pro Glu Asp Glu Ala Asp Tyr Tyr Cys Ser Ala Tyr Asp Arg Ser Leu 85 90 95 Asn Ala Trp Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu 100 105 110 <210> 310 <211> 120 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 310 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ser 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Gly Thr Phe Ser Ser Tyr 20 25 30 Ala Ile Ser Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met 35 40 45 Gly Gly Ile Ile Pro Ile Phe Gly Thr Ala Asn Tyr Ala Gln Lys Phe 50 55 60 Gln Gly Arg Val Thr Ile Thr Ala Asp Lys Ser Thr Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gly Arg Gln Met Phe Gly Ala Gly Ile Asp Phe Trp Gly Pro 100 105 110 Gly Thr Leu Val Thr Val Ser Ser 115 120 <210> 311 <211> 120 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 311 Gln Val Gln Leu Val Gln Ser Gly Gly Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Leu Ser Ser His 20 25 30 Gly Ile Thr Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met 35 40 45 Gly Trp Ile Ser Ala His Asn Gly His Ala Ser Asn Ala Gln Lys Val 50 55 60 Glu Asp Arg Val Thr Met Thr Thr Asp Thr Ser Thr Asn Thr Ala Tyr 65 70 75 80 Met Glu Leu Arg Ser Leu Thr Ala Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Val His Ala Ala Leu Tyr Tyr Gly Met Asp Val Trp Gly Gln 100 105 110 Gly Thr Leu Val Thr Val Ser Ser 115 120 <210> 312 <211> 112 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 312 Asn Phe Met Leu Thr Gln Pro His Ser Val Ser Glu Ser Pro Gly Lys 1 5 10 15 Thr Val Thr Ile Ser Cys Thr Arg Ser Ser Gly Ser Ile Asp Ser Asn 20 25 30 Tyr Val Gln Trp Tyr Gln Gln Arg Pro Gly Ser Ala Pro Thr Thr Val 35 40 45 Ile Tyr Glu Asp Asn Gln Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Ile Asp Ser Ser Ser Asn Ser Ala Ser Leu Thr Ile Ser Gly 65 70 75 80 Leu Lys Thr Glu Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr Asp Ser 85 90 95 Asn Asn Arg His Val Ile Phe Gly Gly Gly Thr Lys Leu Thr Val Leu 100 105 110 <210> 313 <211> 108 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 313 Ser Tyr Glu Leu Thr Gln Pro Pro Ser Val Ser Leu Ala Pro Gly Gln 1 5 10 15 Ser Ala Arg Ile Ser Cys Gly Gly Asp Asn Ile Gly Ser Lys Gly Val 20 25 30 His Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Val Val Val Tyr 35 40 45 Asp Asp Arg Asp Arg Pro Ser Gly Ile Pro Glu Arg Phe Ser Gly Ser 50 55 60 Asn Ser Gly Asn Thr Ala Thr Leu Thr Ile Ser Arg Val Glu Ala Gly 65 70 75 80 Asp Glu Ala Asp Tyr Tyr Cys Gln Val Trp Asp Ser Gly Ser Asp His 85 90 95 Trp Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu 100 105 <210> 314 <211> 108 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 314 Leu Pro Val Leu Thr Gln Pro Pro Ser Val Ser Ala Ala Pro Gly Gln 1 5 10 15 Thr Ala Arg Ile Ser Cys Gly Gly Ser Asn Ile Gly Asp Lys Gly Val 20 25 30 His Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val Ile Tyr 35 40 45 Asp Asp Ser Asp Arg Pro Ser Gly Ile Pro Glu Arg Phe Ser Gly Ser 50 55 60 Asn Ser Gly Asn Thr Ala Thr Leu Thr Ile Ser Arg Val Glu Ala Gly 65 70 75 80 Asp Glu Ala Asp Tyr Tyr Cys Gln Val Trp Asp Ser Ser Ser Asp His 85 90 95 Trp Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu 100 105 <210> 315 <211> 108 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 315 Ser Tyr Glu Leu Thr Gln Pro Pro Ser Val Ser Val Ala Pro Gly Gln 1 5 10 15 Thr Ala Arg Ile Thr Cys Gly Gly Asn Asn Ile Gly Asn Lys Gly Val 20 25 30 His Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val Val Tyr 35 40 45 Asp Asp Ser Asp Arg Pro Ser Gly Ile Pro Glu Arg Phe Ser Gly Ser 50 55 60 Asn Ser Gly Asn Thr Ala Thr Leu Thr Ile Ser Arg Val Glu Ala Gly 65 70 75 80 Asp Glu Ala Asp Tyr Tyr Cys Gln Val Trp Asp Ser Ser Ser Asp His 85 90 95 Trp Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu 100 105 <210> 316 <211> 108 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 316 Leu Pro Val Leu Thr Gln Pro Pro Ser Val Ser Val Ala Leu Gly Gln 1 5 10 15 Thr Ala Arg Ile Thr Cys Arg Gly Asn Asn Ile Gly Gly Lys Gly Val 20 25 30 His Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val Val Tyr 35 40 45 Asp Asp Tyr Ser Arg Arg Ser Gly Ile Pro Glu Arg Phe Ser Gly Ser 50 55 60 His Ser Gly Ser Ala Ala Thr Leu Thr Ile Ser Arg Val Glu Ala Gly 65 70 75 80 Asp Glu Ala Asp Tyr Tyr Cys Gln Val Trp Asp Ser Ser Ser Asp His 85 90 95 Trp Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu 100 105 <210> 317 <211> 366 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 317 caggtgcagc tggtgcagtc tgggggaggc ttggtacagc ctggcaggtc cctgagactc 60 tcctgtgcag cctctggatt cacctttgat gattatgcca tgcactgggt ccggcaagct 120 ccagggaagg gcctggagtg ggtctcaggt attagttgga atagtggtag cataggctat 180 gcggactctg tgaagggccg attcaccgtc tccagagaca acgccaagaa ctcactgtat 240 ctgcaaatga acagcctgag agccgaggac acggctgtgt attactgtgc gagtgactac 300 ggtgacaaat actactacta cggtatggac gtctggggca aagggaccac ggtcaccgtc 360 tcctca 366 <210> 318 <211> 333 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 318 cagcctgggc tgactcagcc accctcagcg tctgggaccc ccgggcagag ggtcaccatc 60 tcttgttctg gaagcagctc caacatcgga agtaatactg tcaactggta tcagcaattc 120 cccggaaagg cccccaaact cctcatcttt aatgataatc agcggccctc aggggtccct 180 gaccgcttct ctgcttccaa gtctggcacc tcagcctccc tggccattag tggcctccag 240 tctgaggatg aggctgacta ttactgtgcg gcatgggatg gcggtctgaa tggtcgaggg 300 gtgttcggcg gagggaccaa actgaccgtc cta 333 <210> 319 <211> 122 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 319 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Gln Pro Gly Arg 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Asp Asp Tyr 20 25 30 Ala Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Gly Ile Ser Trp Asn Ser Gly Ser Ile Gly Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Val Ser Arg Asp Asn Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Ser Asp Tyr Gly Asp Lys Tyr Tyr Tyr Tyr Gly Met Asp Val Trp 100 105 110 Gly Lys Gly Thr Thr Val Thr Val Ser Ser 115 120 <210> 320 <211> 111 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 320 Gln Pro Gly Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn 20 25 30 Thr Val Asn Trp Tyr Gln Gln Phe Pro Gly Lys Ala Pro Lys Leu Leu 35 40 45 Ile Phe Asn Asp Asn Gln Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Ala Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Gln 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Ala Ala Trp Asp Gly Gly Leu 85 90 95 Asn Gly Arg Gly Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu 100 105 110 <210> 321 <211> 285 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 321 accaagggcc catcggtctt ccccctggca ccctcctcca agagcacctc tgggggcaca 60 gcggccctgg gctgcctggt caaggactac ttccccgaac cggtgacggt gtcgtggaac 120 tcaggcgccc tgaccagcgg cgtgcacacc ttcccggctg tcctacagtc ctcaggactc 180 tactccctca gcagcgtggt gaccgtgccc tccagcagct tgggcaccca gacctacatc 240 tgcaacgtga atcacaagcc cagcaacacc aaggtggaca agaaa 285 <210> 322 <211> 48 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 322 gcagagccca aatcttgtga caaaactcac acatgcccac cgtgccca 48 <210> 323 <211> 330 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 323 gcacctgaac tcctgggggg accgtcagtc ttcctcttcc ccccaaaacc caaggacacc 60 ctcatgatct cccggacccc tgaggtcaca tgcgtggtgg tggacgtgag ccacgaagac 120 cctgaggtca agttcaactg gtacgtggac ggcgtggagg tgcataatgc caagacaaag 180 ccgcgggagg agcagtacaa cagcacgtac cgtgtggtca gcgtcctcac cgtcctgcac 240 caggactggc tgaatggcaa ggagtacaag tgcaaggtct ccaacaaagc cctcccagcc 300 cccatcgaga aaaccatctc caaagccaaa 330 <210> 324 <211> 324 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 324 gggcagcccc gagaaccaca ggtgtacacc ctgcccccat cccgggatga gctgaccaag 60 aaccaggtca gcctgacctg cctggtcaaa ggcttctatc ccagcgacat cgccgtggag 120 tgggagagca atgggcagcc ggagaacaac tacaagacca cgcctcccgt gctggactcc 180 gacggctcct tcttcctcta cagcaagctc accgtggaca agagcaggtg gcagcagggg 240 aacgtcttct catgctccgt gatgcatgag gctctgcaca accactacac gcagaagagc 300 ctctccctgt ctccgggtaa atga 324 <210> 325 <211> 321 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 325 ggtcagccca aggctgcccc ctcggtcact ctgttcccgc cctcctctga ggagcttcaa 60 gccaacaagg ccacactggt gtgtctcata agtgacttct acccgggagc cgtgacagtg 120 gcctggaagg cagatggcag ccccgtcaag gcgggagtgg agaccaccac accctccaaa 180 caaagcaaca acaagtacgc ggccagcagc tatctgagcc tgacgcctga gcagtggaag 240 tcccacagaa gctacagctg ccaggtcacg catgaaggga gcaccgtgga gaagacagtg 300 gcccctacag aatgttcatg a 321 <210> 326 <211> 97 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 326 Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 1 5 10 15 Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30 Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45 Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser 50 55 60 Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr 65 70 75 80 Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95 Lys <210> 327 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 327 Ala Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro 1 5 10 15 <210> 328 <211> 110 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 328 Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys 1 5 10 15 Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val 20 25 30 Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr 35 40 45 Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu 50 55 60 Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His 65 70 75 80 Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys 85 90 95 Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys 100 105 110 <210> 329 <211> 107 ...

Claims

[Claim 1] The invention described in the specification of this application.

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