No abstract

The identification of germline mutations in families with HNPCC is hampered by genetic heterogeneity and clinical variability. In previous studies, MSH2 and MLH1 mutations were found in approximately two-thirds of the Amsterdam-criteria-positive families and in much lower percentages of the Amsterdam-criteria-negative families. Therefore, a considerable proportion of HNPCC seems not to be accounted for by the major mismatch repair (MMR) genes. Does the latter result from a lack of sensitivity of mutation detection techniques, or do additional genes underlie the remaining cases? In this study we address these questions by thoroughly investigating a cohort of clinically selected North American families with HNPCC. We analyzed 59 clinically well-defined U.S. families with HNPCC for MSH2, MLH1, and MSH6 mutations. To maximize mutation detection, different techniques were employed, including denaturing gradient gel electrophoresis, Southern analysis, microsatellite instability, immunohistochemistry, and monoallelic expression analysis. In 45 (92%) of the 49 Amsterdam-criteria-positive families and in 7 (70%) of the 10 Amsterdam-criteria-negative families, a mutation was detected in one of the three analyzed MMR genes. Forty-nine mutations were in MSH2 or MLH1, and only three were in MSH6. A considerable proportion (27%) of the mutations were genomic rearrangements (12 in MSH2 and 2 in MLH1). Notably, a deletion encompassing exons 1-6 of MSH2 was detected in seven apparently unrelated families (12% of the total cohort) and was subsequently proven to be a founder. Screening of a second U.S. cohort with HNPCC from Ohio allowed the identification of two additional kindreds with the identical founder deletion. In the present study, we show that optimal mutation detection in HNPCC is achieved by combining accurate and expert clinical selection with an extensive mutation detection strategy. Notably, we identified a common North American deletion in MSH2, accounting for approximately 10% of our cohort. Genealogical, molecular, and haplotype studies showed that this deletion represents a North American founder mutation that could be traced back to the 19th century.

Immunohistochemistry (IHC) of mismatch repair (MMR) proteins in colorectal tumors together with microsatellite analysis (MSI) can be helpful in identi-fying families eligible for mutation analysis. The aims were to determine sensitivity of IHC for MLH1, MSH2, and MSH6 and MSI analysis in tumors from known MMR gene mutation carriers; and to evaluate the use of tissue microarrays for IHC (IHC-TMA) of colon tumors in its ability to identify potential carriers of MMR gene mutations, and compare it with IHC on whole slides. IHC on whole slides was performed in colorectal tumors from 45 carriers of a germline mutation in one of the MMR genes. The TMA cohort consisted of 129 colon tumors from (suspected) hereditary nonpolyposis colorectal cancer (HNPCC) patients. Whole slide IHC analysis had a sensitivity of 89% in detecting MMR deficiency in carriers of a pathogenic MMR mutation. Sensitivity by MSI analysis was 93%. IHC can also be used to predict which gene is expected to harbor the mutation: for MLH1, MSH2, and MSH6, IHC on whole slides would have correctly predicted the mutation in 48%, 92%, and 75% of the cases, respectively. We propose a scheme for the diagnostic approach of families with (suspected) HNPCC. Comparison of the IHC results based on whole slides versus TMA, showed a concordance of 85%, 95%, and 75% for MLH, MSH2, and MSH6, respectively. This study therefore shows that IHC-TMA can be reliably used to simultaneously screen a large number of tumors from (suspected) HNPCC patients, at first in a research setting. Colorectal cancer (CRC) is the second most common cause of death because of malignancy in the Western world. The cause of CRC is multifactorial, involving hereditary and environmental factors and somatic genetic changes during tumor progression. 1 A family history of CRC is a clinically significant risk factor and may be found in up to 15% of all patients with CRC. 2 The most common hereditary CRC syndromes are familial adenomatous polyposis coli (FAP), accounting for Ͻ1% of CRC cases and HNPCC (hereditary nonpolyposis colorectal cancer), accounting for 1 to 6% of the cases. 3 HNPCC is an autosomal dominantly inherited disorder that is clinically defined by the Amsterdam Criteria. 4,5 In HNPCC, germline mutations have been identified in four DNA mismatch repair (MMR) genes, MSH2, 6 MLH1, 7 PMS2, 8 and MSH6. 9 -14 In 50 to 70% of the families fulfilling the Amsterdam criteria a germline mutation is detected in MLH1 or MSH2. 15,16 Germline mutations have been found in MSH6 in families with atypical HNPCC, ie, not entirely fulfilling the Amsterdam criteria. [11][12][13][14] Microsatellite instability (MSI) in colorectal tumors, first reported in 1993, [17][18][19] is caused by a failure of the DNA MMR machinery to repair errors occurring during DNA replication and leading to length alterations in simple, repetitive microsatellite sequences distributed throughout the genome. 20 According to international guidelines, a panel of five specific microsatellite markers has been recommended for MSI evalua...

Because of genetic heterogeneity, the identification of breast cancer-susceptibility genes has proven to be exceedingly difficult. Here, we define a new subset of families with breast cancer characterized by the presence of colorectal cancer cases. The 1100delC variant of the cell cycle checkpoint kinase CHEK2 gene was present in 18% of 55 families with hereditary breast and colorectal cancer (HBCC) as compared with 4% of 380 families with non-HBCC (P<.001), thus providing genetic evidence for the HBCC phenotype. The CHEK2 1100delC mutation was, however, not the major predisposing factor for the HBCC phenotype but appeared to act in synergy with another, as-yet-unknown susceptibility gene(s). The unequivocal definition of the HBCC phenotype opens new avenues to search for this putative HBCC-susceptibility gene.