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Germline alterations in human DNA mismatch repair genes are associated with hereditary nonpolyposis colorectal cancer (HNPCC). Mutation analysis of the genes reveals carriers with a high risk of colorectal cancer, who will benefit from surveillance. We wanted to find the best predictive parameter of a germline mutation in those genes among patients with familial colorectal cancer. Affected members from a total of 83 unrelated colorectal cancer families previously analyzed for mutations in MSH2 and MLH1 were used to evaluate different parameters' ability to predict a germline mutation. We studied various clinical criteria such as family structure, age of onset, and prevalence of endometrial cancer, as well as microsatellite instability in the tumors from the families. In total, 124 tumors from 59 of the families were tested for microsatellite instability (MSI) using PCR‐based mono‐ and dinucleotide markers to establish whether the families could be scored as MSI‐positive or ‐negative. The finding of MSI‐positive tumors in a family was the best predictor of a germline mutation, and was found in 73% of the MSI‐positive, but in less than 3% of the MSI‐negative families (P < 0.0001). In contrast, MSI in unselected colorectal cancer is not as useful, since most of these MSI‐positive tumors are sporadic. The finding of microsatellite instability in colorectal tumors seems efficient enough even to select those with germline mutations among families fulfilling HNPCC Amsterdam criteria, once used in identification of the DNA mismatch repair genes. Genes Chromosomes Cancer 27:17–25, 2000. © 2000 Wiley‐Liss, Inc.

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Summary Genomic instability has been proposed as a new mechanism of carcinogenesis involved in hereditary non-polyposis colorectal cancer (HNPCC) and in a large number of sporadic cancers like pancreatic and colon tumours. Mutations in human mismatch repair genes have been found in HNPCC patients, but their involvement in sporadic cancer has not been clarified yet. In this study we screened 21 pancreatic and 23 colorectal sporadic cancers for microsatellite instability by ten and six different microsatellite markers respectively. Microsatellite alterations were observed at one or more loci in 66.6% (14/21) of pancreatic cancers and in 26% (6/23) colon tumours, but all the pancreatic and half of the colon samples showed a low rate of microsatellite instability. All the unstable samples were further analysed for mutations in the hMLH1 and hMSH2 genes and for hypermethylation of the hMLH1 promoter region. Alterations in the hMLH1 gene were found only in colorectal tumours with a large presence of microsatellite instability. None of the pancreatic tumours showed any alteration in the two genes analysed. Our results demonstrate that microsatellite instability is unlikely to play a role in the tumorigenesis of sporadic pancreatic cancers and confirm the presence of mismatch repair gene alterations only in sporadic colon tumours with a highly unstable phenotype.

Germline alterations in one of five human DNA mismatch repair genes (hMSH2, hMLH1, hPMS1, hPMS2, and hMSH6) cause hereditary nonpolyposis colorectal cancer. Mutation analyses of these genes reveal gene carriers with a high risk for colorectal cancer, who benefit from surveillance to prevent disease. Equally important, presymptomatic testing allows nondisposed individuals to discontinue surveillance. We tested different mutation screening methods to optimize mutation detection in hMSH2 and hMLH1. Affected members from a total of 142 unrelated colorectal cancer families were analyzed. Denaturant gradient gel electrophoresis (DGGE), RT-PCR, and the protein truncation test (PTT) were used to screen for mutations on a DNA or RNA basis, respectively. In addition, a mutation-specific test on genomic DNA was used to find the Finnish mutation no. 1, a deletion of hMLH1 exon 16. DGGE identified most of the mutations in the mismatch repair genes hMLH1 and hMSH2. The RNA-based techniques were used to identify large deletions; however, these were rare in our materials. We describe our compiled results and experience from all our mutation screening studies, as well as unpublished data from our last DGGE screening of 58 patients and RT-PCR and PTT screening of 73 patients.