Mesothelin vaccines and model systems

Abstract

Mesothelin can be used as an immunotherapeutic target. It induces a cytolytic T cell response. Portions of mesothelin which induce such responses are identified. Vaccines can be either polynucleotide- or polypeptide-based. Carriers for raising a cytolytic T cell response include bacteria and viruses. A mouse model for testing vaccines and other anti-tumor therapeutics and prophylactics comprises a strongly mesothelin-expressing, transformed peritoneal cell line.

1 . A method of inducing a T-cell response to a tumor that overexpresses mesothelin relative to normal tissue from which the tumor is derived, said method comprising: 
 administering to a patient who has said tumor or who has had said tumor removed, a vaccine comprising a polypeptide comprising an MHC Class I-binding epitope of mesothelin, wherein the epitope binds to an allelic form of MHC class I which is expressed by the patient, whereby a T-cell response to mesothelin is induced, wherein the vaccine does not comprise whole tumor cells.    
     
     
         2 . The method of  claim 1  wherein the tumor is selected from the group consisting of ovarian cancer, pancreatic cancer, mesothelioma, and squamous cell carcinoma.  
     
     
         3 . The method of  claim 1  wherein the tumor is a pancreatic cancer.  
     
     
         4 . The method of  claim 1  wherein the tumor is an ovarian cancer.  
     
     
         5 . The method of  claim 1  wherein epitope is selected from the group consisting of: SLLFLLFSL (SEQ ID NO: 1); VLPLTVAEV (SEQ ID NO: 2); ELAVALAQK (SEQ ID NO: 3); ALQGGGPPY (SEQ ID NO: 4); FYPGYLCSL (SEQ ID NO: 5); and LYPKARLAF (SEQ ID NO: 6).  
     
     
         6 . The method of  claim 1  wherein the polypeptide is mature mesothelin.  
     
     
         7 . The method of  claim 1  wherein the polypeptide is the primary translation product of mesothelin.  
     
     
         8 . The method of  claim 1  wherein a mixture of said polypeptides is administered.  
     
     
         9 . The method of  claim 8  wherein said polypeptides bind to a plurality of allelic forms of MHC Class I molecules.  
     
     
         10 . The method of  claim 8  wherein said polypeptides bind to a single allelic form of MHC Class I molecules.  
     
     
         11 . The method of  claim 1  wherein the polypeptide is selected as being an MHC class I-binding epitope using an algorithm.  
     
     
         12 . The method of  claim 1  wherein the polypeptide is selected as being an MHC class I-binding epitope using two algorithms.  
     
     
         13 . The method of  claim 1  wherein the T-cell response is induction of specific CD8 +  T cells.  
     
     
         14 . The method of  claim 1  wherein the vaccine is acellular.  
     
     
         15 . The method of  claim 1  wherein the vaccine comprises a bacterium selected from the group consisting of:  Shigella flexneri, E. coli, Listeria monocytogenes, Yersinia enterocolitica, Salmonella typhimurium, Salmonella typhi , and mycobacterium.  
     
     
         16 . The method of  claim 1  wherein the vaccine is administered in sufficient amount to induce tumor regression.  
     
     
         17 . The method of  claim 1  wherein the vaccine is administered in sufficient amount to keep the patient tumor-free after removal of the tumor.  
     
     
         18 . A method of inducing a T-cell response to a tumor that overexpresses mesothelin relative to normal tissue from which the tumor is derived, said method comprising: 
 administering to a patient who has said tumor or who has had said tumor removed, a vaccine comprising a polypeptide comprising an MHC Class II-binding epitope of mesothelin, wherein the epitope binds to an allelic form of MHC class II which is expressed by the patient, whereby a T-cell response to mesothelin is induced, wherein the vaccine does not comprise whole tumor cells.    
     
     
         19 . A method of inducing a T-cell response to tumor cells that overexpress mesothelin relative to normal cells from which the tumor cells are derived, said method comprising: 
 administering to a patient who is at risk of developing a tumor that overexrpresses mesothelin a vaccine comprising a polypeptide comprising an MHC class I-binding epitope of mesothelin or an MHC class II-binding epitope of mesothelin, wherein the epitope binds to an allelic form of MHC class I or class II which is expressed by the patient, whereby a T-cell response to mesothelin is induced, wherein the vaccine does not comprise whole tumor cells.    
     
     
         20 . The method of  claim 19  wherein the patient has been exposed to a carcinogen which is known to induce tumors which overexpress mesothelin relative to normal tissue from which the tumor is derived.  
     
     
         21 . The method of  claim 20  wherein the carcinogen is asbestos.  
     
     
         22 . A method of inducing a T-cell response to a tumor which overexpresses mesothelin relative to normal tissue from which it is derived, said method comprising: 
 administering to a patient who has said tumor or who has had said tumor removed, a vaccine comprising a polynucleotide encoding a polypeptide comprising an MHC Class I-binding epitope of mesothelin, wherein the epitope binds to an allelic form of MHC class I which is expressed by the patient, whereby a T-cell response to mesothelin is induced, wherein the vaccine does not comprise whole tumor cells.    
     
     
         23 . The method of  claim 22  wherein the tumor is selected from the group consisting of ovarian cancer, pancreatic cancer, mesothelioma, and squamous cell carcinoma.  
     
     
         24 . The method of  claim 22  wherein the tumor is a pancreatic cancer.  
     
     
         25 . The method of  claim 22  wherein the tumor is an ovarian cancer.  
     
     
         26 . The method of  claim 22  wherein epitope is selected from the group consisting of: SLLFLLFSL (SEQ ID NO: 1); VLPLTVAEV (SEQ ID NO: 2); ELAVALAQK (SEQ ID NO: 3); ALQGGGPPY (SEQ ID NO: 4); FYPGYLCSL (SEQ ID NO: 5); and LYPKARLAF (SEQ ID NO: 6).  
     
     
         27 . The method of  claim 22  wherein the polypeptide is mature mesothelin.  
     
     
         28 . The method of  claim 22  wherein the polypeptide is primary translation product of mesothelin.  
     
     
         29 . The method of  claim 22  wherein the vaccine comprises one or more polynucleotides encoding a mixture of said polypeptides.  
     
     
         30 . The method of  claim 29  wherein said polypeptides bind to a plurality of allelic forms of MHC Class I molecules.  
     
     
         31 . The method of  claim 29  wherein said polypeptides bind to a single allelic form of MHC Class I molecules.  
     
     
         32 . The method of  claim 22  wherein the polypeptide is selected as being an MHC class I-binding epitope using an algorithm.  
     
     
         33 . The method of  claim 22  wherein the polypeptide is selected as being an MHC class I-binding epitope using two algorithms.  
     
     
         34 . The method of  claim 22  wherein the T-cell response is induction of specific CD8 +  T cells.  
     
     
         35 . The method of  claim 22  wherein the vaccine is acellular.  
     
     
         36 . The method of  claim 22  wherein the vaccine comprises a bacterium selected from the group consisting of:  Shigella flexneri, E. coli, Listeria monocytogenes, Yersinia enterocolitica, Salmonella typhimurium, Salmonella typhi , and mycobacterium.  
     
     
         37 . The method of  claim 22  wherein the vaccine is administered in sufficient amount to induce tumor regression.  
     
     
         38 . The method of  claim 22  wherein the vaccine is administered in sufficient amount to keep the patient tumor-free after removal of the tumor.  
     
     
         39 . A method of inducing a T-cell response to a tumor that overexpresses mesothelin relative to normal tissue from which the tumor is derived, said method comprising: 
 administering to a patient who has said tumor or who has had said tumor removed, a vaccine comprising a polynucleotide encoding a polypeptide comprising an MHC Class II-binding epitope of mesothelin, wherein the epitope binds to an allelic form of MHC class II which is expressed by the patient, whereby a T-cell response to mesothelin is induced, wherein the vaccine does not comprise whole tumor cells.    
     
     
         40 . A method of inducing a T-cell response to tumor cells that overexpress mesothelin relative to normal cells from which the tumor cells are derived, said method comprising: 
 administering to a patient who is at risk of developing a tumor that overexrpresses mesothelin a vaccine comprising a polynucleotide encoding a polypeptide comprising an MHC class I-binding epitope of mesothelin or an MHC class II-binding epitope of mesothelin, wherein the epitope binds to an allelic form of MHC class I or class II which is expressed by the patient, whereby a T-cell response to mesothelin is induced, wherein the vaccine does not comprise whole tumor cells.    
     
     
         41 . The method of  claim 40  wherein the patient has been exposed to a carcinogen which is known to induce tumors which overexpress mesothelin relative to normal tissue from which the tumor is derived.  
     
     
         42 . The method of  claim 41  wherein the carcinogen is asbestos.  
     
     
         43 . A method of identifying immunogens useful as candidates for anti-tumor vaccines, comprising: 
 selecting a protein which is expressed by a tumor and which is minimally or not expressed by normal tissue from which the tumor is derived;    testing lymphocytes of humans who have been vaccinated with a vaccine which comprises said protein to determine if said lymphocytes comprise CD8+ T cells or CD4+ T cells which are specific for said protein, wherein the presence of said CD8+ T cells or CD4+ T cells indicates that the protein is a candidate for use as an anti-tumor vaccine.    
     
     
         44 . The method of  claim 43  wherein said humans have exhibited an anti-tumor immune response.  
     
     
         45 . The method of  claim 43  wherein the vaccine comprises whole tumor cells.  
     
     
         46 . The method of  claim 44  wherein the anti-tumor immune response results in prolonged disease-free survival post-surgical tumor removal relative to a similar population which has not been vaccinated.  
     
     
         47 . The method of  claim 44  wherein the anti-tumor immune response results in tumor regression.  
     
     
         48 . The method of  claim 44  wherein the anti-tumor immune response results in prolonged survival time.  
     
     
         49 . The method of  claim 44  wherein the anti-tumor immune response is delayed type hypersensitivity to autologous tumor cells.  
     
     
         50 . The method of  claim 43  wherein said lymphocytes are also tested to determine if they comprise CD8+ T cells or CD4+ T cells specific for an antigen not expressed by the vaccine.  
     
     
         51 . The method of  claim 43  wherein the humans are divided into two groups based on their response to the vaccine, wherein a first group comprises responders and a second group comprises non-responders, wherein if said CD8+ T cells or CD4+ T cells are found more frequently in responders than in non-responders then the protein is identified as more likely to be useful in an anti-tumor vaccine.  
     
     
         52 . The method of  claim 51  wherein responders display a DTH response to autologous tumor cells but non-responders do not display the response.  
     
     
         53 . The method of  claim 51  wherein responders have a longer period of disease free survival than non-responders.  
     
     
         54 . A method of predicting future response to a tumor vaccine comprising at least one T-cell epitope of mesothelin in a patient who has received the vaccine, comprising: 
 testing lymphocytes of the patient to determine if the lymphocytes comprise CD8+ T cells or CD4+ T cells which are specific for mesothelin, wherein the presence of said CD8+ T cells or CD4+ T cells predicts a longer survival time than the absence of said CD8 +  T cells.    
     
     
         55 . The method of  claim 54  wherein the vaccine comprises whole tumor cells.  
     
     
         56 . The method of  claim 54  wherein the vaccine comprises pancreatic tumor cells and the antigen is mesothelin.  
     
     
         57 . The method of  claim 54  wherein the vaccine comprises ovarian tumor cells and the antigen is mesothelin.  
     
     
         58 . The method of  claim 54  wherein the vaccine comprises mesothelioma cells and the antigen is mesothelin.  
     
     
         59 . A vaccine which induces a CD8 +  T cell or CD4 +  T cell response, comprising: 
 a polypeptide comprising an MHC Class I- or Class II-binding epitope of mesothelin, wherein the epitope binds to an allelic form of MHC class I of class II which is expressed by the patient, whereby a CD8 +  T cell or CD4 +  T-cell response to mesothelin is induced, wherein the vaccine does not comprise whole tumor cells; and    a carrier for stimulating a CD8 +  T cell or CD4 +  T cell immune response.    
     
     
         60 . The vaccine of  claim 59  wherein the polypeptide comprises an MHC Class I-binding epitope.  
     
     
         61 . The vaccine of  claim 59  wherein the polypeptide comprises between 6 and 20 amino acid residues.  
     
     
         62 . The vaccine of  claim 59  wherein the polypeptide comprises an epitope selected from the group consisting of SLLFLLFSL (SEQ ID NO: 1); VLPLTVAEV (SEQ ID NO: 2); ELAVALAQK (SEQ ID NO: 3); ALQGGGPPY (SEQ ID NO: 4); FYPGYLCSL (SEQ ID NO: 5); and LYPKARLAF (SEQ ID NO: 6).  
     
     
         63 . The vaccine of  claim 59  wherein the carrier is CD40/CD40 ligand.  
     
     
         64 . The vaccine of  claim 59  wherein the carrier is OX-40/OX-40 ligand.  
     
     
         65 . The vaccine of  claim 59  wherein the carrier is a CTLA-4 antagonist.  
     
     
         66 . The vaccine of  claim 59  wherein the carrier is GM-CSF.  
     
     
         67 . A vaccine which induces a CD8 +  T cell or CD4 +  T cell response, comprising: 
 a polynucleotide encoding a polypeptide comprising an MHC Class I- or Class II-binding epitope of mesothelin, wherein the epitope binds to an allelic form of MHC class I or Class II which is expressed by the patient, whereby a CD8 +  T cell or CD4 +  T-cell response to mesothelin is induced, wherein the vaccine does not comprise whole tumor cells; and    a carrier for stimulating a CD8 +  T cell or CD4 +  T cell immune response.    
     
     
         68 . The vaccine of  claim 67  wherein the carrier is CD40/CD40 ligand.  
     
     
         69 . The vaccine of  claim 67  wherein the carrier is OX-40/OX-40 ligand.  
     
     
         70 . The vaccine of  claim 67  wherein the carrier is a CTLA-4 antagonist.  
     
     
         71 . The vaccine of  claim 67  wherein the carrier is GM-CSF.  
     
     
         72 . The vaccine of  claim 67  wherein the polypeptide comprises an epitope selected from the group consisting of SLLFLLFSL (SEQ ID NO: 1); VLPLTVAEV (SEQ ID NO: 2); ELAVALAQK (SEQ ID NO: 3); ALQGGGPPY (SEQ ID NO: 4); FYPGYLCSL (SEQ ID NO: 5); and LYPKARLAF (SEQ ID NO: 6).  
     
     
         73 . The vaccine of  claim 59  which comprises a bacterium.  
     
     
         74 . The vaccine of  claim 67  which comprises a bacterium.  
     
     
         75 . The vaccine of  claim 73  wherein the bacterium is selected from the group consisting of: 
   Shigella flexneri, E. coli, Listeria monocytogenes, Yersinia enterocolitica, Salmonella typhimurium, Salmonella typhi , and mycobacterium.    
     
     
         76 . The vaccine of  claim 74  wherein the bacterium is selected from the group consisting of: 
   Shigella flexneri, E. coli, Listeria monocytogenes, Yersinia enterocolitica, Salmonella typhimurium, Salmonella typhi , and mycobacterium.    
     
     
         77 . An isolated polypeptide of 9 to 25 amino acid residues comprising an epitope selected from the group consisting of SLLFLLFSL (SEQ ID NO: 1); VLPLTVAEV (SEQ ID NO: 2); ELAVALAQK (SEQ ID NO: 3); ALQGGGPPY (SEQ ID NO: 4); FYPGYLCSL (SEQ ID NO: 5); and LYPKARLAF (SEQ ID NO: 6).  
     
     
         78 . A fusion protein comprising a first and a second portion, wherein the first portion comprises a polypeptide of 9 to 25 amino acid residues comprising an epitope selected from the group consisting of SLLFLLFSL (SEQ ID NO: 1); VLPLTVAEV (SEQ ID NO: 2); ELAVALAQK (SEQ ID NO: 3); ALQGGGPPY (SEQ ID NO: 4); FYPGYLCSL (SEQ ID NO: 5); and LYPKARLAF (SEQ ID NO: 6), and the second portion comprises a segment of at least 6 amino acid residues, wherein the sequence of said second portion is not in mesothelin.  
     
     
         79 . An expression vector which encodes a polypeptide of 9 to 25 amino acid residues comprising an epitope selected from the group consisting of SLLFLLFSL (SEQ ID NO: 1); VLPLTVAEV (SEQ ID NO: 2); ELAVALAQK (SEQ ID NO: 3); ALQGGGPPY (SEQ ID NO: 4); FYPGYLCSL (SEQ ID NO: 5); and LYPKARLAF (SEQ ID NO: 6).  
     
     
         80 . A bacterium which comprises the expression vector of  claim 79 .  
     
     
         81 . The bacterium of  claim 80  which is selected from the group consisting of  Shigella flexneri, E. coli, Listeria monocytogenes, Yersinia enterocolitica, Salmonella typhimurium, Salmonella typhi , and mycobacterium.  
     
     
         82 . An expression vector which encodes the fusion protein of  claim 78 .  
     
     
         83 . A bacterium which comprises the expression vector of  claim 82 .  
     
     
         84 . The bacterium of  claim 83  which is selected from the group consisting of  Shigella flexneri, E. coli, Listeria monocytogenes, Yersinia enterocolitica, Salmonella typhimurium, Salmonella typhi , and mycobacterium.  
     
     
         85 . An isolated antibody that binds to an epitope selected from the group consisting of SLLFLLFSL (SEQ ID NO: 1); VLPLTVAEV (SEQ ID NO: 2); ELAVALAQK (SEQ ID NO: 3); ALQGGGPPY (SEQ ID NO: 4); FYPGYLCSL (SEQ ID NO: 5); and LYPKARLAF (SEQ ID NO: 6).  
     
     
         86 . A T-cell line that binds to an epitope selected from the group consisting of SLLFLLFSL (SEQ ID NO: 1); VLPLTVAEV (SEQ ID NO: 2); ELAVALAQK (SEQ ID NO: 3); ALQGGGPPY (SEQ ID NO: 4); FYPGYLCSL (SEQ ID NO: 5); and LYPKARLAF (SEQ ID NO: 6).  
     
     
         87 . The polypeptide of  claim 77  which is bound to an MHC Class I molecule.  
     
     
         88 . The fusion protein of  claim 78  which is bound to an MHC Class I molecule.  
     
     
         89 . The vaccine of  claim 59  wherein the carrier is an MHC Class I molecule.  
     
     
         90 . The polypeptide of  claim 87  wherein the MHC Class I molecule is on a dendritic cell.  
     
     
         91 . The fusion protein of  claim 88  wherein the MHC Class I molecule is on a dendritic cell.  
     
     
         92 . The vaccine of  claim 89  wherein the MHC Class I molecule is on a dendritic cell.  
     
     
         93 . The polypeptide of  claim 87  wherein the MHC Class I molecule is on an antigen presenting cell.  
     
     
         94 . The polypeptide of  claim 88  wherein the MHC Class I molecule is on an antigen presenting cell.  
     
     
         95 . The vaccine of  claim 89  wherein the MHC Class I molecule is on an antigen presenting cell.  
     
     
         96 . A method of predicting future response to a tumor vaccine in a patient who has received the vaccine, comprising: 
 testing the patient to determine if the patient has a delayed type hypersensitivity (DTH) response to mesothelin, wherein the presence of said response predicts a longer survival time than the absence of said response.    
     
     
         97 . The method of  claim 96  wherein the vaccine comprises whole tumor cells.  
     
     
         98 . The method of  claim 96  wherein the vaccine comprises pancreatic tumor cells.  
     
     
         99 . The method of  claim 96  wherein the vaccine comprises ovarian tumor cells.  
     
     
         100 . The method of  claim 96  wherein the vaccine comprises mesothelioma cells.  
     
     
         101 . A recombinant mouse cell line which comprises peritoneal cells which have been transformed by HPV-16 E6 and E7 and an activated oncogene wherein the cell line is capable of forming ascites and tumors upon intraperitoneal injection into an immunocompetent mouse.  
     
     
         102 . The recombinant mouse cell line of  claim 101  wherein the activated oncogene is an activated c-Ha-ras.  
     
     
         103 . The recombinant mouse cell line of  claim 101  which expresses mesothelin.  
     
     
         104 . The recombinant mouse cell line of  claim 101  which is WF-3.  
     
     
         105 . A mouse model comprising: 
 a mouse which has been injected with the recombinant mouse cell line of  claim 101 .    
     
     
         106 . The mouse model of  claim 105  which is immunocompetent.  
     
     
         107 . A method of testing a substance to determine if it is a potential drug for treating a cancer selected from the group consisting of ovarian cancer, pancreatic cancer, mesothelioma, and squamous cell carcinoma, comprising: 
 contacting the mouse model of  claim 105  with a test substance; and    determining if the test substance causes regression of a tumor in the mouse model, diminution of ascites volume in the mouse model, or longer survival time in the mouse model.    
     
     
         108 . A method of testing a substance to determine if it is a potential drug for treating a cancer selected from the group consisting of ovarian cancer, pancreatic cancer, mesothelioma, and squamous cell carcinoma, comprising: 
 contacting a mouse with a test substance;    injecting the mouse with the recombinant cell line of  claim 101 , and determining if the test substance causes regression of a tumor in the mouse, diminution of ascites volume in the mouse, or longer survival time in the mouse.    
     
     
         109 . The vaccine of  claim 59 , wherein the polypeptide is mesothelin.  
     
     
         110 . The method of  claim 1 , wherein the polypeptide is mesothelin.  
     
     
         111 . The method of  claim 22 , wherein the polypeptide is mesothelin.  
     
     
         112 . The vaccine of  claim 67 , wherein the polypeptide is mesothelin.