Protein Tyrosine Phosphatase Mutations in Cancers

Abstract

Tyrosine phosphorylation, regulated by protein tyrosine phosphatases (PTPs) and kinases (PTKs), is important in signaling pathways underlying tumorigenesis. A mutational analysis of the tyrosine phosphatase gene superfamily in human cancers identified 83 somatic mutations in six PTPs (PTPRF, PTPRG, PTPRT, PTPN3, PTPN13, PTPN14), affecting 26% of colorectal cancers and a smaller fraction of lung, breast and gastric cancers. Fifteen mutations were nonsense, frameshift or splice site alterations predicted to result in truncated proteins lacking phosphatase activity. Five missense mutations in the most commonly altered PTP (PTPRT) were biochemically examined and found to reduce phosphatase activity. Expression of wild-type but not a mutant PTPRT in human cancer cells inhibited cell growth. These observations suggest that the tyrosine phosphatase genes are tumor suppressor genes, regulating cellular pathways that may be amenable to therapeutic intervention.

We claim: 
     
         1 . A method for identifying mutations involved in cancer, comprising:
 determining nucleotide sequence differences in a human nucleotide sequence in matched pairs of cancer cells and normal cells, each pair being from a single individual, wherein the human nucleotide sequence encodes a protein tyrosine phosphatase selected from the group consisting of: PTPRF, PTPRG, PTPRT, PTPN3, PTPN13, and PTPN14.   
     
     
         2 . The method of  claim 1  wherein at least one of the nucleotide sequence differences is in a region encoding a catalytic domain. 
     
     
         3 . The method of  claim 1  wherein at least one of the nucleotide sequence differences is a point mutation. 
     
     
         4 . The method of  claim 1  wherein at least one of the nucleotide sequence differences is in a regulatory region. 
     
     
         5 . The method of  claim 1  further comprising determining whether a nucleotide sequence difference is synonymous or non-synonymous. 
     
     
         6 . The method of  claim 1  further comprising determining whether a nucleotide difference affects an evolutionarily conserved amino acid residue. 
     
     
         7 . The method of  claim 1  wherein the protein tyrosine phosphatase is PTPRT. 
     
     
         8 . The method of  claim 1  wherein the protein tyrosine phosphatase is PTPRG. 
     
     
         9 . The method of  claim 1  wherein the cancer cells are selected from the group consisting of bladder cancer, melanoma, breast cancer, non-Hodgkin's lymphoma, colon and rectal cancer, pancreatic cancer, endometrial cancer, prostate cancer, kidney cancer (renal cell), skin (non-melanoma), leukemia, thyroid cancer, and lung cancer. 
     
     
         10 . A method of screening test substances for use as anti-cancer agents, comprising:
 contacting a test substance with a wild-type form or a mutant form of a protein tyrosine phosphatase which is mutated in cancer cells, wherein the protein tyrosine phosphatase is selected from the group consisting of: PTPRF, PTPRG, PTPRT, PTPN3, PTPN13, and PTPN14;   testing activity of the form of the protein tyrosine phosphatase, wherein a test substance which increases the activity of the form of a protein tyrosine phosphatase is a potential anti-cancer agent.   
     
     
         11 . The method of  claim 10  wherein the form of a protein tyrosine phosphatase is in a cell. 
     
     
         12 . The method of  claim 10  wherein the form of a protein tyrosine phosphatase is isolated from a cell. 
     
     
         13 . The method of  claim 10  wherein the form of a protein tyrosine phosphatase is in a cell of a cancer cell line. 
     
     
         14 . The method of  claim 10  wherein the form of a protein tyrosine phosphatase is in a cell which has been modified to express the form of a protein tyrosine phosphatase. 
     
     
         15 . The method of  claim 10  wherein the form of a protein tyrosine phosphatase is PTPRF. 
     
     
         16 . The method of  claim 10  wherein the form of a protein tyrosine phosphatase is PTPRG. 
     
     
         17 . The method of  claim 10  wherein the form of a protein tyrosine phosphatase is PTPRT. 
     
     
         18 . The method of  claim 10  wherein the form of a protein tyrosine phosphatase is PTPN3. 
     
     
         19 . The method of  claim 10  wherein the form of a protein tyrosine phosphatase is wild-type. 
     
     
         20 . The method of  claim 10  wherein the form of a protein tyrosine phosphatase is PTPN13. 
     
     
         21 . The method of  claim 10  wherein the form of a protein tyrosine phosphatase is PTPN14. 
     
     
         22 . The method of  claim 15  wherein the PTPRF has a mutation selected from the group consisting of R218C, G645R, G1040V, R1333C, V1390I, 387-468 substitution, and A381V. 
     
     
         23 . The method of  claim 16  wherein the PTPRG has a mutation selected from the group consisting of T361M, A462V, T514M, R593W, E955G, Y973C, R1312W, and I1326V. 
     
     
         24 . The method of  claim 17  wherein the PTPRT has a mutation selected from the group consisting of A209T, V1269M, A209T, F248S, Y280H, Y412F, N510K, T605M, V648G, R632X, A707T, D927G, A707V, R1021X, F74S, L708P, R975X, LOH, Q987K, A1118P, T1368M, N1128I, R1212W, M1259L, I395V, Y1351F, T1368M, R453C, K218T, R1346L, and R790I. 
     
     
         25 . The method of  claim 18  wherein the PTPN3 has a mutation selected from the group consisting of V154I, A239V, E610X, S300A, F307L, and R330Q. 
     
     
         26 . The method of  claim 20  wherein the PTPN13 has a mutation selected from the group consisting of H2Q, E952X, G1476C, R380X, M2443I, R402X, S443N, A529D, Q1691X, K2131N, D2154H, R2205W, R2338X, Y2279X, M2307T, I2458V, and E2474D. 
     
     
         27 . The method of  claim 21  wherein the PTPN14 has a mutation selected from the group consisting of L56M, R293Q, S314P, Q332R, R491Q, P525L, H633Y, P528L, T657M, E883D, and T1068M. 
     
     
         28 . An isolated, mutant form of protein tyrosine phosphatase selected from the group consisting of: PTPRF, PTPRG, PTPRT, PTPN3, PTPN13, and PTPN14, wherein enzymatic activity of the mutant form is reduced compared to wild-type. 
     
     
         29 . The isolated, mutant form of  claim 28  which is PTPRF. 
     
     
         30 . The isolated, mutant form of  claim 28  which is PTPRG. 
     
     
         31 . The isolated, mutant form of  claim 28  which is PTPRT. 
     
     
         32 . The isolated, mutant form of  claim 28  which is PTPN3. 
     
     
         33 . The isolated, mutant form of  claim 28  which is PTPN13. 
     
     
         34 . The isolated, mutant form of  claim 28  which is PTPN14. 
     
     
         35 . The isolated, mutant form of  claim 29  wherein the PTPRF has a mutation selected from the group consisting of R218C, G645R, G1040V, R1333C, V1390I, 387-468 substitution, and A381V. 
     
     
         36 . The isolated, mutant form of  claim 30  wherein the PTPRG has a mutation selected from the group consisting of T361M, A462V, T514M, R593W, E955G, Y973C, R1312W, and I1326V. 
     
     
         37 . The isolated, mutant form of  claim 31  wherein the PTPRT has a mutation selected from the group consisting of A209T, V1269M, A209T, F248S, Y280H, Y412F, N510K, T605M, V648G, R632X, A707T, D927G, A707V, R1021X, F74S, L708P, R975X, LOH, Q987K, A1118P, T1368M, N1128I, R1212W, M1259L, I395V, Y1351F, T1368M, R453C, K218T, R1346L, and R790I. 
     
     
         38 . The isolated, mutant form of  claim 32  wherein the PTPN3 has a mutation selected from the group consisting of V154I, A239V, E610X, S300A, F307L, and R330Q. 
     
     
         39 . The isolated, mutant form of  claim 33  wherein the mutant PTPN13 has a mutation selected from the group consisting of H2Q, E952X, G1476C, R380X, M2443I, R402X, S443N, A529D, Q1691X, K2131N, D2154H, R2205W, R2338X, Y2279X, M2307T, I2458V, and E2474D. 
     
     
         40 . The isolated, mutant form of  claim 34  wherein the mutant PTPN14 has a mutation selected from the group consisting of L56M, R293Q, S314P, Q332R, R491Q, P525L, H633Y, P528L, T657M, E883D, and T1068M. 
     
     
         41 . An isolated polynucleotide which encodes a mutant form of protein tyrosine phosphatase selected from the group consisting of: PTPRF, PTPRG, PTPRT, PTPN3, PTPN13, and PTPN14, wherein enzymatic activity of the mutant form is reduced compared to wild-type. 
     
     
         42 . A host cell comprising the polynucleotide of  claim 41 . 
     
     
         43 . A method of categorizing cancers, comprising:
 determining the coding sequence for or the amino acid sequence of one or more protein tyrosine phosphatase family members selected from the group consisting of PTPRF, PTPRG, PTPN3, PTPN13, and PTPN14 in a sample of a cancer tissue;   identifying a somatic mutation of said one or more protein tyrosine phosphatase family members in the cancer tissue;   assigning the cancer tissue to a group based on the presence or absence of the somatic mutation.   
     
     
         44 . The method of  claim 43  wherein the mutation is one which truncates the protein tyrosine phosphatase family member. 
     
     
         45 . The method of  claim 43  wherein the mutation is non-synonymous. 
     
     
         46 . The method of  claim 43  wherein the mutation is somatic. 
     
     
         47 . The method of  claim 43  wherein the mutation is a point mutation. 
     
     
         48 . The method of  claim 43  wherein the protein kinase family member is PTPRF. 
     
     
         49 . The method of  claim 43  wherein the protein kinase family member is PTPRG. 
     
     
         50 . The method of  claim 43  wherein the protein kinase family member is PTPN3. 
     
     
         51 . The method of  claim 43  wherein the protein kinase family member is PTPN13. 
     
     
         52 . The method of  claim 43  wherein the protein kinase family member is PTPN14. 
     
     
         53 . The method of  claim 43  wherein the group is used to analyze or design clinical trials. 
     
     
         54 . The method of  claim 43  wherein the group is used to correlate with prognostic data. 
     
     
         55 . The method of  claim 43  wherein the group is used to correlate with recurrence data. 
     
     
         56 . The method of  claim 43  wherein the group is used to select an appropriate therapeutic agent. 
     
     
         57 . The method of  claim 43  wherein the group is used to identify cancer. 
     
     
         58 . The method of  claim 43  wherein the cancer tissue is colorectal. 
     
     
         59 . The method of  claim 43  wherein the cancer tissue is breast. 
     
     
         60 . The method of  claim 43  wherein the cancer tissue is lung. 
     
     
         61 . The method of  claim 43  wherein the cancer tissue is gastric. 
     
     
         62 . A method of inhibiting growth of cancer cells, comprising:
 administering to cancer cells a polynucleotide encoding a wild-type protein tyrosine phosphatase selected from the group consisting of PTPRF, PTPRG, PTPRT, PTPN3, PTPN13, and PTPN14, whereby growth of the cancer cells is inhibited.   
     
     
         63 . The method of  claim 62  wherein the cancer cells are colorectal. 
     
     
         64 . The method of  claim 62  wherein the cancer cells are breast. 
     
     
         65 . The method of  claim 62  wherein the cancer cells are lung. 
     
     
         66 . The method of  claim 62  wherein the cancer cells are gastric. 
     
     
         67 . The method of  claim 62  wherein the cancer cells are in culture. 
     
     
         68 . The method of  claim 62  wherein the cancer cells are in a human body. 
     
     
         69 . The method of  claim 62  wherein the polynucleotide is administered by intratumoral injection. 
     
     
         70 . The method of  claim 62  wherein the cancer cells comprise one or two mutant alleles of a protein tyrosine phosphatase selected from the group consisting of PTPRF, PTPRG, PTPRT, PTPN3, PTPN13, and PTPN14. 
     
     
         71 . A method of identifying cancer cells in a sample collected from a human, comprising:
 determining the coding sequence for or the amino acid sequence of one or more protein tyrosine phosphatase family members selected from the group consisting of PTPRF, PTPRG, PTPN3, PTPN13, and PTPN14 in a sample collected from the human, wherein the sample is selected from the group consisting of a suspected cancer tissue, blood, serum, plasma, and stool;   identifying a somatic mutation of said one or more protein tyrosine phosphatase family members in the cancer tissue;   identifying the sample as containing cancer cells if a somatic mutation is identified.   
     
     
         72 . The method of  claim 71  wherein the protein tyrosine phosphatase family member is PTPRF. 
     
     
         73 . The method of  claim 71  wherein the protein tyrosine phosphatase family member is PTPRG. 
     
     
         74 . The method of  claim 71  wherein the protein tyrosine phosphatase family member is PTPN3. 
     
     
         75 . The method of  claim 71  wherein the protein tyrosine phosphatase family member is PTPN13. 
     
     
         76 . The method of  claim 71  wherein the protein tyrosine phosphatase family member is PTPN14. 
     
     
         77 . The method of  claim 1  wherein the protein tyrosine phosphatase is PTPRF. 
     
     
         78 . The method of  claim 1  wherein the protein tyrosine phosphatase is PTPN3. 
     
     
         79 . The method of  claim 1  wherein the protein tyrosine phosphatase is PTPN13 
     
     
         80 . The method of  claim 1  wherein the protein tyrosine phosphatase is PTPN14.