Construction of a Model Used for the Mismatch Repair Functional Analysis in vitro and Being Used in the Human Colorectal Cancer Cell Line and Tumor Tissue

Purpose A model for the analysis of mismatch repair (MMR) function has been constructed to measure the mismatch repair function of human colorectal cancer cell and tumor tissue.The results have been compared with the results of microsatellite stability (MSS) assay.Evaluate the values of these two measurements in the basic research and clinical diagnosis and treatment of colorectal cancer. Methods With the bacteriophage M13mp2 and its three derivates two heteroduplex DNA molecular has been constructed one with a single base mismatch,theother with two bases deletion.The heteroduplex DNA used as the template and TK6-a lymphoblastoid B-cell line with MMR competent ability make up of the model to analyze the MMR ability of all kinds of samples in vitro.Extract from Lovo-a human colorectal cancer cell line with hMSH2 homologous deletions in the exons 3 to 8 and tumor tissues of 1 case of HNPCC (hereditary non-polyposis colorectal cancer) and 1 cases of SRC (sporadic rectal cancer),which has been measurd by large T antigen dependent SV-40 DNA replication assay,has been incubated with the heteroduplex DNA.From the change of the percentage of the mixture plaques the functional status of MMR system has been evaluated.Five sites recommended by the international co-operation group of HNPCC have been measured to analyze the microsatellites in all the cell lines,tumor tissues and normal tissues of the patients. Results The repair efficiency of TK6 to del(2) was more than 60%,to G·G was more than 50%.Ther Lovo efficiency was to del(2) less than 10%,G·G is less than 20%.The tumor tissue of a case of NHPCC was lack of MMR ability and a case of spordic rectal cancer (SRC) still maintained MMR proficiency.Both tumor and normal tissue of the HNPCC patients and SRC patient had been measured by the microsatellits assay.The HNPCC was MSI-H and the SRC was MSS. Conclusions The whole MMR functional status of cell line and tissue can be measured by the model used for MMR functional analysis in vitro,which is sensitive and dependable.It's very useful to be used in the patients whose family history is not clear.But its operation is complex and need longer time to complete.The results of the functional analysis and microsatellite assay are concordance,which indicates the recommended sites used in the HNPCC are sensitive too.The microsatellite assay is stable and quick,which is suitable in the clinical diagnosis and treatment.It's meaningful to use these two measurements together in the mechanism research and classification of the colorectal cancer and in the clinical diagnosis and treatment of it.