Identification of DNA mismatch repair gene mutations in hereditary nonpolyposis colon cancer patients

Abstract

Hereditary nonpolyposis colorectal cancer (HNPCC) is a common autosomal dominant disease caused by germline mutations in DNA mismatch repair genes. The mutational spectrum in these genes appears to be diverse, in both the distribution and the nature of the mutations. However, most described mutations generate a premature stop codon and ultimately result in the synthesis of a truncated protein. We have employed an in vitro transcription/translation assay to identify germline mutations in DNA mismatch repair genes from patients suspected of belonging to HNPCC kindreds. Our results suggest that this approach will be highly effective in identifying mutations in these patients and may lead to a reliable diagnostic test for the pre-symptomatic identification of HNPCC.

Putative common origin of two MLH1 mutations in Italian‐Quebec hereditary non‐polyposis colorectal cancer families

“…We used the TNT® Quick‐Coupled Transcription/Translation kit from Promega (Madison, WI) for protein truncation test (PTT). The PCR‐amplified cDNA segments were transcribed and translated in vitro in a reaction mixture containing 35 S‐methionine, and the resultant polypeptides were separated on a 12% polyacrylamide SDS gel, dried, and subjected to autoradiography (12). Sequencing was carried out on PCR‐amplified DNA segments by cycle sequencing using Thermo Sequenase enzyme.…”

Use of SSCP analysis to identify germline mutations in HNPCC families fulfilling the Amsterdam criteria

“…The search for germline mutations in the human mismatch repair genes is time-consuming and expensive because of the involvement of multiple genes and the heterogeneity of the mutations found (Luce et al 1996). However, a knowledge of the gene carrier status in an individual from an HNPCC family enables targeted surveillance with the potential for curative surgical intervention (Lynch et al 1995;Cunningham and Dunlop 1996).…”

“…A number of different techniques have been used to search for mutations in the human mismatch repair genes. An in vitro transcription/translation (IVTT) assay, alternatively called the protein truncation test (PTT), has been successfully applied in identifying germline mutations in HNPCC patients (Liu et al 1996;Luce et al 1996). However, the IVTT/PTT assay has a number of limitations.…”

Ovarian Neoplasm and Endometrioid Carcinoma in a Patient with Turcot Syndrome

“…Affected members can develop other manifestations, including upper gastrointestinal tract adenomas, osseous and soft tissue tumors, and extracolonic neoplasms including carcinomas of the thyroid, bile duct, and duodenum (5). Also an autosomal dominant disorder, HNPCC is caused by germline mutation of one of the DNA‐mismatch repair genes, including hMSH2, hMLH1, hPMS1 and hPMS2 (human postmeiotic segregation 1 and 2) (6,7). Hereditary nonpolyposis colon cancer is characterized by absence of polyposis, proximal colon cancer, increased incidence of multiple primary colon cancers, and early age of onset (8).…”