CAJ | 학술논문

目的探讨K-RAS第12、13密码子及B-RAF V600E突变在结肠癌(CRC)演化不同阶段的分布情况及其与相关临床参数的关系.方法临床回顾性研究.收集2011年1月至2012年1月于上海市长宁区中心医院进行肠镜检查患者130例,其中结肠息肉21例、腺瘤44例与肠癌65例,抽提DNA.建立使用带有高、低温内标的高分辨率熔解分析体系,检测上述DNA样本中的K-RAS第12、13密码子及B-RAF V600E突变,并使用Sanger测序法对K-RAS突变进行明确分型,使用Fischer确切概率法统计各突变在疾病演化不同阶段的分布及与癌组织体积大小、Dukes分期及瘤体分化情况等临床参数间的关系.结果在息肉增生、腺瘤及癌组织中分别发现K-RAS突变2例、8例及26例,B-RAF突变1例、1例和2例.K-RAS突变率在息肉增生(9.5％)、腺瘤(18.2％)与癌组织(40％)中逐渐升高,且在腺瘤与癌组织中的差异具有统计学意义(P =0.013).K-RAS突变率在低分化癌中高于中、高分化癌(P=0.04),且第12密码子突变比率高于13密码子(P =0.028).所发现K-RAS突变主要类型为c.35G＞A、c.38G＞A、c.35G＞T与c.34G＞T.B-RAF突变率较低,在息肉、腺瘤与癌中分别为4％、2.3％与3.1％,且未发现同时存在K-RAS与B-RAF突变的病例.结论在CRC演化过程中,K-RAS突变主要出现于腺瘤向癌转化过程.对CRC患者而言,进行K-RAS突变检测并进行明确分型对于判断肿瘤分化情况具有一定的辅助意义.
목적 탐토K-RAS제12、13밀마자급B-RAF V600E돌변재결장암(CRC)연화불동계단적분포정황급기여상관림상삼수적관계.방법 림상회고성연구.수집2011년1월지2012년1월우상해시장저구중심의원진행장경검사환자130례,기중결장식육21례、선류44례여장암65례,추제DNA.건립사용대유고、저온내표적고분변솔용해분석체계,검측상술DNA양본중적K-RAS제12、13밀마자급B-RAF V600E돌변,병사용Sanger측서법대K-RAS돌변진행명학분형,사용Fischer학절개솔법통계각돌변재질병연화불동계단적분포급여암조직체적대소、Dukes분기급류체분화정황등림상삼수간적관계.결과 재식육증생、선류급암조직중분별발현K-RAS돌변2례、8례급26례,B-RAF돌변1례、1례화2례.K-RAS돌변솔재식육증생(9.5％)、선류(18.2％)여암조직(40％)중축점승고,차재선류여암조직중적차이구유통계학의의(P =0.013).K-RAS돌변솔재저분화암중고우중、고분화암(P=0.04),차제12밀마자돌변비솔고우13밀마자(P =0.028).소발현K-RAS돌변주요류형위c.35G＞A、c.38G＞A、c.35G＞T여c.34G＞T.B-RAF돌변솔교저,재식육、선류여암중분별위4％、2.3％여3.1％,차미발현동시존재K-RAS여B-RAF돌변적병례.결론 재CRC연화과정중,K-RAS돌변주요출현우선류향암전화과정.대CRC환자이언,진행K-RAS돌변검측병진행명학분형대우판단종류분화정황구유일정적보조의의.
Objective The aim of this study is to identify the K-RAS codon 12,13 and B-RAF V600E mutation patterns in different neoplastic progression stages of colorectal cancer (CRC) and analyze their relationship with the clinical parameters.Methods A clinical retrospective study of 130 patients with colonoscopy surgery from January 2011 to January 2012 in Shanghai Changning District Central Hospitalwas performed.DNA extractions were from frozen excision tissues of 21 normal mucosa,44 adenomas and 65 CRCs.High-resolution melting analysis (HRMA) systems with short amplicon primer and melting temperature calibrator were constructed to detect the K-RAS codon 12,13 and B-RAF V600E mutations from DNA extractions.Sanger sequencing was then performed to genotype the K-RAS mutations.The relationship between the mutation patterns and clinical parameters as neoplastic progression stage,size of lesion,dukes' stage and tumor differentiation was evaluated with Fisher exact probability test.Results K-RAS mutations were detected in 2 normal mucosa (9.5％),8 adenomas (18.2％) and 26 CRCs (40％).The mutation rates showed a rising trend with the disease progression,and had a significantly difference between the adenoma and CRC tissues (P =0.013).These K-RAS mutations were more frequent in the poorly differentiated CRCs than that in the moderately and highly differentiated CRCs (P =0.04).Main type of K-RAS mutations were identified as c.35G ＞ A,c.38G ＞ A,c.35G ＞T and c.34G ＞ T.Moreover,mutations in codon 12 had a higher frequency of occurrence compared with the mutations in codon 13 in the poorly differentiated CRC lesions (P =0.028).B-RAF mutation was detected in 1 mucosa (4％),1 adenoma (2.3％) and 2 CRCs (3.1％),while no concurrent K-RAS and B-RAF mutation was found in this study.Conclusion K-RAS mutations occurred mainly in the adenoma to cancer transformation process in the CRC neoplastic progression.Detection and genotyping of the K-RAS codon 12 and 13 mutation will help to determine the differentiation of CRC.