外周血循环肿瘤细胞在评估结直肠癌患者KRAS基因突变中的价值

The value of peripheral blood circulating tumor cells in KRAS mutation testing of colorectal cancer patients

目的 探索外周血标本在结直肠癌患者KRAS基因突变检测中的价值以及循环肿瘤细胞个数与KRAS基因突变检测的相关性。方法 收集江南大学附属医院2013年至2014年结直肠癌石蜡切片及对应的外周血标本各112例，另外收集10例肠炎外周血标本作为对照。运用扩增受阻突变体系法检测上述标本的KRAS基因突变状况，同时，运用免疫荧光原位杂交技术检测外周血样本及肠炎对照标本中循环肿瘤细胞的数目。结果 在112例结直肠癌中，石蜡包埋切片有46例发现了KRAS基因的突变（41.1%）；外周血中，有25例发现了KRAS基因的突变（22.3%），差异有统计学意义（χ2=40.12，P〈0.001）。但其中1例在石蜡包埋切片的检测中为野生型，而在外周血标本的检测中为突变型；另1例标本的KRAS基因突变位点在两种不同的样本中检测结果不同。当肿瘤细胞个数＞15时，其检测的敏感性为73.3%，而循环肿瘤细胞个数在5～15之间时，其检测敏感性为41.9%，循环肿瘤细胞个数为1~5时，检测敏感性仅为16.7%，差异有统计学意义（χ^2=23.70，P〈0.001）。10例肠炎标本均未检测到外周血循环肿瘤细胞，也未检测到KRAS基因的突变。结论 以外周血为标本进行KRAS基因检测具备良好的特异性，准确率欠佳。当患者不具备手术条件时，可作为结直肠患者KRAS基因检测的一种选择性方法。外周血中循环肿瘤细胞的个数越多，KRAS基因突变检测的敏感性越高。

ObjectiveTo explore the value of peripheral blood samples in KRAS mutation testing of colorectal cancer patients and the correlation between the number of circulating tumor cells and KRAS mutation testing. MethodsWe detected KRAS mutation using amplification refractory mutation system PCR method in paraffin embedded tissues and matched peripheral blood samples obtained from 112 colorectal cancer patients and 10 proctitic peripheral blood samples in Affiliated Hospital of Jiangnan University between 2013 and 2014. Meanwhile, immunofluorescence in situ hybridization method was used to count the circulating tumor cells in peripheral blood samples and proctitic control samples. ResultsAmong the 112 colorectal cancer samples tested, 25 cases of peripheral blood samples found KRAS mutation （41.1%） and which was 46 in formalin fixed paraffin embedded tissues testing （22.3%）, with a significant difference （χ2=40.12,P〈0.001）. One case with KRAS wild type in formalin fixed paraffin embedded tissues was mutation type in peripheral samples. In another case, mutation site was different in different kinds of samples. The sensibility of KRAS mutation testing was 73.3%, 41.9% and 16.7% when the number of circulating tumor cells was more than 15, 5 to 15, and 1 to 5, respectively, with significant differences （χ^2=23.70, P〈0.001）. No KRAS mutation and no circulating tumor cells were found in 10 proctitic control samples. ConclusionWe find high specificity in KRAS mutation testing of peripheral blood samples. but the accurate rate is not satisfying. KRAS mutation testing in peripheral blood samples may be an optional choice to test KRAS mutations for colorectal cancer patients who were not subjected to surgery. The sensibility of KRAS mutation testing in peripheral blood samples has a corretion with the number of circulating tumor cells.