Objective:To clarify the origin of defective mismatch repair (MMR) in sporadic endometrial cancers with microsatellite instability (MSI), a thorough mutation analysis was performed on the human mismatch repair gene MSH3.
Methods:Twenty-eight MSI-positive endometrial cancers were investigated for mutations in the human mismatch repair gene MSH-3 using single-strand conformation variant (SSCV) analysis of all 24 exons. All variants were sequenced. Loss of heterozygosity was investigated at all MSH3 polymorphisms discovered. A subset of tumors were investigated for methylation of the 5′ promoter region of MSH3 using Southern blot hybridization.
Results:An identical single-base deletion (ΔA) predicted to result in a truncated protein was discovered in six tumors (21.4%). This deletion occurs in a string of eight consecutive adenosine residues (A8). Because simple repeat sequences are unstable in cells with defective MMR, the observed mutation may be an effect, rather than a cause, of MSI. Evidence of inactivation of the second MSH3 allele in tumors with the ΔA mutation would strongly support a causal role for these MSH3 mutations. However, there was no evidence of a second mutation, loss of sequences, or methylation of the promoter region in any of the tumors with the ΔA mutation.
Conclusion: Although the ΔA mutation is a frequent event in sporadic MSI-positive endometrial cancers, it may not be causally associated with defective DNA MMR.
1. Rose PG. Endometrial carcinoma. N Engl J Med1996;335:640-649.
2.
2. Burks RT, Kessis TD, Cho KR, Hedrick L. Microsatellite instability in endometrial carcinoma. Oncogene1994;9:1163-1166.
3.
3. Peiffer SL, Herzog TJ, Tribune DJ, Mutch DG, Gersell DJ, Goodfellow PJ. Allelic loss of sequences from the long arm of chromosome 10 and replication errors in endometrial cancers. Cancer Res1995;55:1922-1926.
4.
4. Duggan BD, Felix JC, Muderspach LI, Turgeman D, Zheng J, Shibata DK. Microsatellite instability in sporadic endometrial carcinoma. J Natl Cancer Inst1994;86:1216-1221.
5.
5. Kobayashi K, Sagae S, Kudo R, Saito H, Koi S, Nakamura Y. Microsatellite instability in endometrial carcinomas: frequent replication errors in tumors of early onset and/or of poorly differentiated type. Genes Chromosom Cancer1995;14:128-132.
6.
6. Parsons R, Li G-M, Longley MJ, et al. Hypermutability and mismatch repair deficiency in MSI+ tumor cells. Cell1993;75:1227-1236.
7.
7. Nyström-Lahti M, Wu Y, Moisio A-L, et al. DNA mismatch repair gene mutations in 55 kindreds with verified or putative hereditary non-polyposis colorectal cancer. Hum Mol Genet1996;5(6):763-769.
8.
8. Buersteede J-M, Alday P, Torhorst J, Weber W, Müller H, Scott R. Detection of new mutations in 6 out of 10 Swiss HNPCC families by genomic sequencing of the hMSH2 and hMLH1 genes. J Med Genet1995;32:909-912.
9.
9. Bronner CE, Baker SM, Morrison PT, et al. Mutation in the DNA mismatch repair gene homologue hMLH1 is associated with hereditary non-polyposis colon cancer. Nature1994;368:258-261.
10.
10. Leach FS, Nicolaides NC, Papadopoulos N, et al. Mutations of a mutS homolog in hereditary nonpolyposis colorectal cancer. Cell1993;75:1215-1225.
11.
11. Liu B, Parsons RE, Hamilton SR, et al. hMSH2 mutations in hereditary nonpolyposis colorectal cancer kindreds. Cancer Res1994;54:4590-4594.
12.
12. Nicolaides NC, Papadopoulos N, Liu B, et al. Mutations of two PMS homologues in hereditary nonpolyposis colon cancer. Nature1994;371:75-80.
13.
13. Fishel R, Lescoe MK, Rao MRS, et al. The human mutator gene homolog MSH2 and its association with hereditary nonpolyposis colon cancer. Cell1993;75:1027-1038.
14.
14. Papadopoulos N, Nicolaides NC, Wei Y-F, et al. Mutations of a mutL homolog in hereditary colon cancer. Science1994;263:1625-1629.
15.
15. Aarnio M, Mecklin J-P, Aaltonen LA, Nyström-Lahti M, Järvinen HJ. Life-time risk of different cancers in hereditary nonpolyposis colorectal cancer (HNPCC) syndrome. Int J Cancer (Pred Oncol)1995;64:430-433.
16.
16. Kowalski LD, Mutch DG, Herzog TJ, Rader JS, Goodfellow PJ. Mutational analysis of MLH1 and MSH2 in 25 prospectively-acquired MSI+ endometrial cancers. Genes Chromosom Cancer1997;18:219-227.
17.
17. Kobayashi K, Matsushima M, Koi S, et al. Mutational analysis of mismatch repair genes, hMLH and hMSH2, in sporadic endometrial carcinomas with microsatellite instability. Jpn J Cancer Res1996;87:141-145.
18.
18. Katabuchi H, van Rees B, Lambers AR, et al. Mutations in DNA mismatch repair genes are not responsible for microsatellite instability in most sporadic endometrial carcinomas. Cancer Res1995;55:5556-5560.
19.
19. Gardiner-Garden M, Frommer M. CpG islands in vertebrate genomes. J Mol Biol1987;196:261-282.
20.
20. Craig JM, Bickmore WA. The distribution of CpG islands in mammalian chromosomes. Nat Genet1994;7:376-381.
21.
21. Laird PW, Jaenisch R. DNA methylation and cancer. Hum Mol Genet1994;3:1487-1495.
22.
22. Herman JG, Latif F, Weng Y, et al. Silencing of the VHL tumor-supressor gene by DNA methylation in renal carcinoma. Proc Natl Acad Sci U S A1994;91:9700-9704.
23.
23. Gonzalez-Zulueta M, Bender CM, Yang AS, et al. Methylation of the 5′ CpG island of the p16/CDKN2 tumor supressor gene in normal and transformed human tissues correlates with gene silencing. Cancer Res1995;55:4531-4535.
24.
24. Stirzaker C, Millar DS, Paul CL, et al. Extensive DNA methylation spanning the Rb promoter in retinoblastoma tumors. Cancer Res1997;57:2229-2237.
25.
25. Merlo A, Herman JG, Mao L, et al. 5′ CpG island methylation is associated with transcriptional silencing of the tumor supressor p16/CDKN2/MTS1 in human cancers. Nature Med1995;1:686-692.
26.
26. Herman JG, Jen J, Merlo A, Baylin SB. Hypermethylation-associated inactivation indicates a tumor supressor role for p15INK4B1. Cancer Res1996;56:722-727.
27.
27. Ahuja N, Mohan AL, Li Q, et al. Association between CpG island methylation and microsatellite instability in colorectal cancer. Cancer Res1997;57:3370-3374.
28.
28. Kane MF, Loda M, Gaida GM, et al. Methylation of the hMLH1 promoter correlates with lack of expression of hMLH1 in sporadic colon tumors and mismatch repair-defective human tumor cell lines. Cancer Res1997;57:808-811.
29.
29. Malkhosyan S, Rampino N, Yamamoto H, Perucho M. Frameshift mutator mutations. Nature1996;382:499-500.
30.
30. Akiyama Y, Tsubouchi N, Yuasa Y. Frequent somatic mutations of hMSH3 with reference to microsatellite instability in hereditary nonpolyposis colorectal cancers. Biochem Biophys Res Commun1997;236:248-252.
31.
31. Risinger JI, Umar A, Boyd J, Berchuck A, Kunkel TA, Barrett JC. Mutations of MSH3 in endometrial cancer and evidence for its functional role in heteroduplex repair. Nat Genet1996;14:102-105.
32.
32. Lahiri DK, Nurnberger JIA. A rapid non-enzymatic method for the preparation of HMW DNA from blood for RFLP studies. Nucleic Acids Res1991;19:5444-5444.
33.
33. Watanabe A, Ikejima M, Suzuki N, Shimada T. Genomic organization and expression of the human MSH3 gene. Genomics1996;31:311-318.
34.
34. Donis-Keller H, Dou S, Chi D, et al. Mutations in the RET proto-oncogene are associated with MEN-2A and FMTC. Hum Mol Genet1993;2:851-856.
35.
35. Goodfellow PJ, Darling SM, Thomas NS, Goodfellow PN. A pseudoautosomal gene in man. Science1986;234:740-743.
36.
36. Nakajima E, Orimo H, Ikejima M, Shimada T. Nine-bp repeat polymorphism in exon 1 of the hMSH3 gene. Jpn J Hum Genet1995;40:343-345.
37.
37. Liu B, Nicolaides NC, Markowitz S, et al. Mismatch repair gene defects in sporadic colorectal cancers with microsatellite instability. Nat Genet1995;9:48-55.
38.
38. Herfarth K-F, Kodner IJ, Whelan AJ, et al. Mutations in MLH1 are more frequent that in MSH2 in sporadic colorectal cancers with microsatellite instability. Genes Chromosom Cancer1997;18:42-49.
39.
39. Koi M, Umar A, Chauhan DP, et al. Human chromosome 3 corrects mismatch repair deficiency and microsatellite instability and reduced N -methyl-N ′-nitro-N -nitrosoguanidine tolerance in colon tumor cells with homozygous hMLH1 mutation. Cancer Res1994;54:4308-4312.
40.
40. Hemmunki A, Peltomäki P, Mecklin J-P, et al. Loss of the wild type MLH1 gene is a feature of hereditary nonpolyposis colorectal cancer. Nat Genet1994;8:405-410.
