内蒙古335例结直肠癌组织相关基因突变分析

Mutation analysis of related genes for 335 colorectal cancer patients in Inner Mongolia

目的随着结直肠癌分子靶向治疗的进展以及分子生物学技术的快速发展,结直肠癌患者有必要进行相应基因突变检测。本研究分析内蒙古地区结直肠癌患者组织中KRAS、NRAS、BRAF和PIK3CA基因突变类型及其与临床病理特征关系。方法选取2014-05-05-2017-05-30内蒙古自治区肿瘤医院(65例)和内蒙古自治区人民医院(270例)收治的结直肠癌石蜡包埋组织,提取DNA,采用二代测序法检测KRAS、NRAS、BRAF和PIK3CA基因突变状态,统计各个基因突变类型,分析各个基因突变频率与临床病理特征关系。结果在335例结直肠癌患者中,检测阳性突变共137例(40.90%),其中KRAS75例(22.39%),NRAS10例(2.99%),BRAF20例(5.97%),PIK3CA32例(9.55%),包括PIK3CA与KRAS、NRAS、BRAF基因发生双突变20例(14.60%)。KRAS主要突变类型包括G12V、G13D、V14I、G13_V14insG、G12C、G12S、G15S、A59E、T58IA11_G12insGA、G12A、Q61H、A146V、A130T和A59V,其中G12V、G13D突变频率较高。NRAS仅有6种突变,分别为G12D、G12C、G12S、G12D、G13R和Q61H,其中G12D为主要突变类型;BRAF主要突变类型为V600E、G596V、N581T和L584F;PIK3CA主要突变类型为H1047R、Q546L、E545K、E542K、E542V、V680I、T1025=、A1035V、M1043V和M1043I。KRAS基因突变与血管侵袭(χ~2=0.546,P=0.044)和肿瘤部位(χ~2=46.123,P<0.001)有关联,NRAS基因突变与性别(χ~2=5.113,P=0.033)有关联。结论结直肠癌中KRAS阳性突变率居高,PIK3CA次之,BRAF和NRAS突变率最低,且PIK3CA常与KRAS、NRAS和BRAF发生交叉突变。对结直肠癌患者行KRAS、NRAS、BRAF和PIK3CA等多基因联合检测,可根据患者基因突变情况,正确选择表皮生长因子受体靶向抗体药物,从而实现个体化精准检验、精准治疗和靶向治疗策略。

OBJECTIVE The experiment analyzed the gene mutation types of KRAS,NRAS,BRAF,PIK3CA in colorectal cancer tissues and the relationship between clinicopathologic indicators and gene mutation types.METHODS The DNAs were extracted from paraffin-embedded tissues of 335 cases of colorectal cancer and the gene mutations of KRAS,NRAS,BRAF,PIK3 CA were detected by next generation sequencing.The relationship between each gene mutation rate and the clinicopathologic indicators were analyzed together with the mutation types in each gene.RESULTS There were 137 cases(40.90%)harbouring mutations,including KRAS(75 cases,22.39%),NRAS(10 cases,2.99%),BRAF(20 cases,5.97%)and PIK3 CA(32 cases,9.55%),in which 20 cases(14.60%)had double mutation involving PIK3 CA with KRAS,NRASor BRAFgens.The main mutation types of KRASincluded G12 V,G13 D,V14 I,G13_V14 insG(InsertionIn frame),G12 C,G12 S,G15 S,A59 E,T58 IA11_G12 insGA(Insertion-In frame),G12 A,Q61 H,A146 V,A130 T and A59 V.NRASincluded G12 D,G12 C,G12 S,G12 D,G13 Rand Q61 H.BRAFincluded V600 E,G596 V,N581 Tand L584 F.PIK3 CAincluded H1047 R,Q546 L,E545 K,E542 K,E542 V,V680 I,T1025=,A1035 V,M1043 Vand M1043 I.Cross mutation occurred between PIK3 CAand KRAS,NRASor BRAF,but not among KRAS,NRASand BRAF.Analysis of the relationship between mutation frequency of each gene and clinicopathological characteristics showed that the incidence of KRAS mutation was related to vascular invasion(χ~2=0.546,P=0.044)and was also related to the location of colorectal tumors(χ~2=46.12,P<0.001).NRASgene mutation was associated with gender(χ~2=5.113,P=0.033).CONCLUSIONS In colorectal cancer,KRAS mutation rate is the highest,PIK3 CA the second and BRAF,NRASthe lowest.PIK3 CA gene mutation is always coupled with KRAS,NRASor BRAF.It is of great necessity to have the general gene mutation screen of KRAS,NRAS,BRAFand PIK3 CA for colorectal cancer patients,which can provide the right guidance to choose the anti-EGFR monoclonal antibody and realize the precision examine and precision medicine,and fulfill the individual target therapeutic strategics.