UNDETECTABLE EXPRESSION OF HMLH1 PROTEIN IN SPORADIC COLORECTAL-CANCER WITH REPLICATION ERROR PHENOTYPE

PURPOSE: Four DNA mismatch repair genes have been identified as being susceptible genes for hereditary nonpolyposis colorectal cancer. Defic iency of one of the mis match repair genes causes the replication erro r phenotype in more than 80 percent of patients with hereditary nonpol yposis colorectal cancer and in 10 to 30 percent of patients with spor adic colorectal cancer. To determine which mismatch repair gene is lac king the function in patients with replication error-positive colorect al cancer, several approaches have been used at the nucleic acid and p rotein levels. We studied replication error in 40 samples of randomly selected colorectal cancers and expression of hMSH2 and hMLH1 proteins analyzed by immunoblot in the tumor and normal tissues of the replica tion error-positive and replication error-negative samples, MATERIALS AND METHODS: Frozen tumor and normal tissues were obtained from 40 Jap anese patients who had colorectal cancer. According to the Amsterdam c riteria, those patients were classified as having 39 sporadic and 1 un known colorectal cancers. Genomic DNA was extracted from tumor and nor mal tissues for determining replication error with eight microsatellit e markers. Expression of hMSH2 and hMLH1 proteins in cell lysates of t umor and normal tissues of 16 patients was analyzed by immunoblot. RES ULTS: The replication error phenotype was found in 6 (15 percent) of t he 39 sporadic cases. hMLH1 protein was not detected in two of the six replication error-positive tumor tissues and not in the normal tissue s, indicating that the tumor cells of the two patients had severe muta tions in both alleles of the hMLH1 gene. Another four replication erro r-positive and ten replication error-negative tumors and normal tissue s expressed hMLH1 protein. hMSH2 protein was detected in all samples. CONCLUSION: hMLH1 protein was undetectable in the two tumor tissues of the six replication error-positive samples of sporadic colorectal can cer. The detection procedure used here may have potential use for dete rmining a dysfunctional mismatch repair gene product.