New strategy for cancer therapy using Hi-resolution microsatellite instability analysis and Real-time quantitative reverse transcriptase polymerase reaction

微小转移;采用实时定量逆转录酶聚合酶链反应（RT-PCR）检测骨髓和外周血中弥散性癌细胞。我们比较了20个细胞系的CEA、CK18、CK19和CK20的表达，以选择最合适的表达。通过对13名健康志愿者外周静脉血中CK 18 mRNA表达的分析，我们发现CK 18 mRNA的表达远低于癌细胞系。然而，所有样品中的表达水平也可以用常规RT-PCR检测到。因此，不仅定性，而且定量分析靶mRNA对检测微转移很重要。定量RT-PCR方法将比较目标基因表达水平可能存在的差异。对于临床应用，还需要进一步的研究。Microsalellite不稳定;我们采用高分辨率荧光微卫星分析（HFRMA）对800多例散发性乳腺癌、胃癌、结肠癌、肺癌和肝癌患者进行了研究。在该系统中，为了提高聚合酶链反应产物的重现性、电泳的迁移准确性和检测系统的特性，制备了几种装置。该系统观测到的微卫星变化可分为A型和B型，A型微卫星序列在有限的位点上变化相对较小，B型微卫星序列变化剧烈且分布广泛。前者被认为与DNA错配修复（MMR）中的异常活动有关。

Micrometastasis; Real time quantitative reverse transcriptase polymerase chain reaction (RT-PCR) was used to detect disseminated cancer cells in bone marrow and peripheral blood. We compared CEA, CK18, CK19 and CK20 expression using 20 of cell lines in order to select the most appropriate. In an analysis of CK 18 mRNA expression in peripheral venous blood in 13 healthy volunteers, we found that CK 18 mRNA was much less expressed than in cancer cell lines. However, the expression in all samples was at a level which was also detected using conventional RT-PCR. It would thus seem that not only qualitative, but also quantitative analysis, of the target mRNA is important to detect micrometastasis. Quantitative RT-PCR methods will make comparisons of the possible differences in expression levels of the target gene. For clinical applications, much further study is needed.Microsalellite Instability; Using high-resolution fluorescent microsatellite analysis (HFRMA), we studied over 800 patients with sporadic breast, gastric, colon, lung and liver cancer. In this system, several devices were prepared to improve reproducibility of polymerase chain reaction products, migration accuracy of electrophoresis, and characteristics of the detection system. All the microsatellite changes observed in this system can be classified into two types: type A with relatively small changes in microsatellite sequences observed in limited loci and type B with drastic and widely dispersed changes. The former was thought to be connected to abnormal activity in DNA mismatch repair (MMR).