中文摘要：

目的探讨应用焦磷酸测序技术检测血浆KRAS基因突变的实用性,了解晚期非小细胞肺癌（NSCLC）血浆KRAS基因突变率和突变特征。方法收集63例晚期NSCLC患者的外周血标本,分离血浆并提取DNA,采用巢式PCR结合焦磷酸测序分析KRAS基因12和13号密码子突变。结果在63例晚期NSCLC患者中,3例存在血浆KRAS基因突变（4.76%）,突变率较低,与亚裔组织KRAS突变率一致,低于西方人KRAS突变率,其突变类型均为单碱基替换突变。其中2例为12密码子突变,1例为13密码子突变。统计学分析未发现血浆KRAS突变与性别、吸烟史、年龄、病理类型、肿瘤分期、体力状况（PS）评分存在相关性。结论焦磷酸测序技术操作简便,可以快速、高通量地进行外周血循环KRAS基因突变检测,可广泛用于筛选对酪氨酸激酶抑制剂耐药的NSCLC患者,更有利于指导患者的个体化分子靶向治疗。

英文摘要：

Objective To evaluate the utility of pyrosequencing to detect KRAS mutations in plasma,and to investigate the frequency and characteristic of plasma KRAS mutations of patients with advanced NSCLC.Methods We collected 63 blood samples from advanced NSCLC patients,isolated plasma and extracted DNA.The mutations in codon 12 and 13 of KRAS were detected by pyrosequencing of the nested-PCR fragment.Results Somatic mutations in KRAS gene in plasma were identified from 3 cases of 63（4.76%） patients.The frequency of KRAS mutations in plasma of advanced NSCLC in our center coincided with that in tumor of Asian population,but was lower than that in Western population.The mutations were all one-base substitution,of which two cases in codon 12 and one case in codon 13.KRAS mutations did not relate with gender,smoking history,age,pathological type,tumor stage,and PS score.Conclusion Pyrosequencing is a fast method with high-throughput.It can be used for detecting KRAS mutations in circulating DNA,and is suitable for screening patients refractory with tyrosine kinase inhibitors.It is conductive to guide the patients to choose personalized therapy with molecular targeted agents.