Study Of The Regulation Of Mismatch Repair In Colorectal Cancer

Colorectal cancer (CRC) is one of the most common malignant tumors. Various factors and mechanisms are involved in the development of CRC. Lose of heterozygosity (LOH) and microsatellite instability (MSI) are considered to be important mechanisms.MSI was firstly reported in hereditary nonporyposis colorectal cancer (HNPCC). HNPCC is an inherited autosomal dominant syndrome with special clinicopathological features. Families who fit Amsterdam criteria are called ICG-HNPCC, while others who fit suspected HNPCC criteria are named as sHNPCC. It has been reported that ICG-HNPCC and sHNPCC families shared similar clinicopathological and molecular characteristics. HNPCC is generally associated with germ-line mutations in two mismatch repair (MMR) genes: hMLHl and hMSH2. In China, mere are still no systematic researches on HNPCC or sHNPCC and the deficiency of MMR genes in mem. Therefore, it is necessary and urgent to find out the characteristics HNPCC and the deficiency of MMR in Chinese populations.MSI was considered to be an indicator for MMR gene mutations. BAT26 梐 special 26 (A) repeated sequence, locating in the 5* intron of hMSH2 gene, was reported to be a good and specific marker. But some other authors thought it was not sensitive ^"PMg*1, and they suggested combining it with other markers. Therefore, we combined BAT26 with BAT25 to detect MSI in mis study. Furthermore, we also studied whether MSI can regulate the MMR function.Hypermethylation at promoter regions are considered to be another regulatory mechanism of MMR genes. Hypermemylation of hMLHl promoter were detected in 84% of MSI positive sporadic colorectal cancers. In China there was no report on hypermethylation of hMLHl promoter and its relationship withMSI. We do some studies in this text.Part 1: clinical phenotype of Chinese HNPCC and the expression of MMRAccording to follow-up records, 54 probands from HNPCC families (including 12 ICG-HNPCC families and 42 sHNPCC families) were screened out from 1502 patients with colorectal cancers operated upon in 2nd affiliated hospital of Zhejiang University from 1984 to 2001. Their climcopathological features were compared with those of sporadic CRCs. The tumor spectrums in these families were also analyzed. Expressions of hMLHl and hMSH2 genes were detected with immunohistochemical staining in 20 HNPCC and 16 sporadic colorectal cancers. Results: Compared with sporadic colorectal cancers, 54 probands from HNPCC families were obviously different at age, tumor site, differentiation and multiple tumors (PO.05). Besides of colorectal cancer, extracolonic tumors occurred in stomach, endometrium, hepatobiliary system, and so on, can also be seen.By immunohistochemical staining, absent expression of hMLHl and HMSH2 were detected 45% and 30% in HNPCC patients respectively, while 18.75% and 18.75% in sporadic colorectal cancers.Therefore, we made the conclusion that in China ICG-HNPCC and sHNPCC also have the similar hereditary background. Deficiency of hMLHl and KMSH2 are associated with 45% and 30% Chinese HNPCC respectively. The hMLHl is the main responsible gene.Part 2: Regulation of MMR gene1. MMR and MSISix of 60 sporadic colorectal cancers are mutated at the 5* exon of hMSH2 gene. BAT26 was used to detect their MSI by denatured polyacrylamide gel and Genescan. Only 50% of hMSH2 mutated tumors showed BAT26+. Therefore, it is better to combine BAT26 with other microsateUites to detect MSI+. BAT26 located in the 5* intronof hMSH2 gene, aD of three BAT26+ samples had mutations in the 5th exon of hMSH2, indicating that changes in BAT26 mightinduce the mutations. The results were confirmed by either denatured poryacrylamide gel method or Genescan. From this point, Genescan is a quick and sensitive way to detect MSI.2. Hypermethylation of MMR promoter and MSICombined with BAT2S and BAT26, MSI status of 41 pair colorectal cancers including 1 P-J syndrome and 1 HNPCC were detected by Genescan. Hypermethylation of hMLHl promoter were detected by MSP (methylation specific P...