结直肠癌患者RAS、BRAF基因突变与MSI状态的相关性及其临床病理特征

Correlation between RAS,BRAF gene mutations and MSI status in colorectal cancer patients and their clinicopathological characteristics

目的:结直肠癌是人类最常见的恶行肿瘤之一,其发病率和病死率呈逐年上升趋势。本研究旨在分析结直肠癌患者大鼠肉瘤病毒癌基因(rat sarcoma viral oncogene,RAS)、鼠类肉瘤病毒癌基因同源物(V-Raf murine sarcoma viral oncogene homolog B,BRAF)突变与微卫星不稳定(microsatellite instability,MSI)状态相关性及其临床病理特征,为结直肠癌临床治疗及预后评估提供依据。方法:回顾性分析444例结直肠癌患者的临床病理资料,并提取肿瘤组织基因组DNA,采用荧光定量PCR法检测Kirsten大鼠肉瘤病毒癌基因同源物(Kirsten rat sarcoma viral oncogene homolog,KRAS)、Neuroblastoma大鼠肉瘤病毒癌基因同源物(neuroblastoma rat sarcoma viral oncogene homolog,NRAS)第2、3、4外显子及BRAF V600E突变;运用PCR-毛细血管电泳行MSI状态分析。结果:K(N)RAS基因突变184例(41.44%,184/444),其中KRAS突变173例(94.02%,173/184),NRAS突变11例(5.98%,11/184),K(N)RAS共突变3例。BRAF V600E突变21例(4.7%,21/444)。微卫星高度不稳定(microsatellite instability high,MSIH)48例(10.81%,48/444),微卫星低度不稳定(microsatellite instability low,MSI-L)和微卫星稳定(microsatellite stable,MSS)共396例(89.19%,396/444)。K(N)RAS基因突变右半结肠癌多于左半结肠癌,且黏液腺癌突变率高于单纯腺癌(均P<0.05)。K(N)RAS基因突变与患者性别、年龄、肿瘤大小、肉眼分型、组织学分化程度、淋巴结转移、病理分期均无关(均P>0.05)。女性BRAF V600E突变高于男性,低分化高于中分化(均P<0.05),BRAF V600E突变与患者发病年龄、肿瘤位置、大小、肉眼分型、组织学类型、淋巴结转移、病理分期均无关(均P>0.05)。发生MSI-H的患者中,右半结肠癌多于左半结肠癌,肿瘤直径≥4.5 cm病例多于直径更小病例,隆起型多于其他肉眼分型,黏液腺癌多于单纯腺癌,低分化多于高分化(均P<0.05),而与患者性别和年龄均无关(均P>0.05)。K(N)RAS突变与MSI-H发生无关(P>0.05),但RAS基因Gly位点突变与MSI-H发生相关(P<0.05)。携带RAS基因Gly位点突变的MSI-H结直肠癌患者男性多于女性,发病年龄无差异,右半结肠多于左半结肠,肉眼分型多为溃疡型,单纯腺癌比例高于黏液腺癌,多为中低分化,较多处于病理分期I~II期,常不伴淋巴结转移和BRAF V600E突变,Gly突变位点多为Gly12Ser/Asp突变。BRAF V600E突变与MSI-H发生有关(P<0.05)。与K(N)RAS突变型MSI-L/MSS结直肠癌比较,K(N)RAS突变型MSI-H结直肠癌较易发生于右半结肠,肿瘤直径≥4.5 cm的病例较多,组织学类型多为黏液腺癌,较少发生淋巴结转移,多处于病理分期I~II(均P<0.05);K(N)RAS突变型MSI-H结直肠癌与性别、年龄、肿瘤肉眼分型、组织学分化均无相关(均P>0.05)。与RAS野生型-MSI-L/MSS结直肠癌比较,RAS野生型-MSI-H结直肠癌多发生于右半结肠,肿瘤直径≥4.5 cm,多为溃疡型和隆起型,较多为黏液腺癌,且多为中低分化(均P<0.05),而RAS野生型-MSI-H结直肠癌与性别、年龄、淋巴结转移、病理分期均无关(均P>0.05)。结论:结直肠癌KRAS基因Gly位点突变、BRAF V600E突变与MSI-H发生相关;MSI-H-RAS基因Gly点突变结直肠癌患者有其独特的临床病理特征;RAS-MSI基因状态与临床病理特征相关;K(N)RAS、BRAF V600E、MSI检测有利于结直肠癌患者个体化治疗及预后评估。

Objective:Colorectal cancer is one of the most common malignant tumors in humans,with incidence and mortality rates increasing annually.This study aims to analyze the correlation between rat sarcoma viral oncogene(RAS),V-Raf murine sarcoma viral oncogene homolog B(BRAF)mutations,microsatellite instability(MSI)status,and the clinicopathological characteristics of colorectal cancer patients to provide a basis for clinical treatment and prognosis evaluation.Methods:The clinicopathological data of 444 colorectal cancer patients were retrospectively analyzed.Genomic DNA was extracted from tumor tissues,and mutations in Kirsten rat sarcoma viral oncogene homolog(KRAS),neuroblastoma rat sarcoma viral oncogene homolog(NRAS)exon 2,3,and 4,as well as BRAF V600E,were detected using quantitative PCR.MSI status was analyzed using PCR-capillary electrophoresis.Results:Among the 444 patients,184 cases(41.44%)had K(N)RAS mutations,including 173 cases(94.02%)with KRAS mutations and 11 cases(5.98%)with NRAS mutations,with 3 cases having both KRAS and NRAS mutations.BRAF V600E mutations were found in 21 cases(4.7%),MSI high(MSI-H)was identified in 48 cases(10.81%),while MSI low(MSIL)and microsatellite stable(MSS)were found in 396 cases(89.19%).K(N)RAS mutations were more frequent in right-sided colorectal cancers and mucinous adenocarcinomas compared to simple adenocarcinomas(both P<0.05).K(N)RAS mutations were not associated with patient gender,age,tumor size,gross type,histological differentiation,lymph node metastasis,or pathological stage(all P>0.05).BRAF V600E mutations were more common in females and poorly differentiated tumors(both P<0.05),but were not related to age,tumor location,size,gross type,histological type,lymph node metastasis,or pathological stage(all P>0.05).MSI-H was more frequent in right-sided colorectal cancers,tumor diameters≥4.5 cm,elevated gross type,mucinous adenocarcinomas,and poorly differentiated tumors(all P<0.05),but was not associated with gender or age(both P>0.05).K(N)RAS mutation was not associated with MSI-H(P>0.05),but RAS mutation at the Gly position was correlated with MSI-H(P<0.05).Among MSI-H colorectal cancer patients with RAS Gly mutations,males were more common,and the tumors were predominantly right-sided,ulcerative,simple adenocarcinomas,moderately to poorly differentiated,and mostly at the pathological stage I-II,with no lymph node metastasis or BRAF V600E mutations.The most common RAS Gly mutations were Gly12Ser/Asp.BRAF V600E mutation was associated with MSI-H(P<0.05).Compared to K(N)RAS-mutated MSI-L/MSS colorectal cancers,K(N)RAS-mutated MSI-H colorectal cancers were more likely to occur in the right colon,with tumor diameters≥4.5 cm,mucinous adenocarcinoma histology,fewer lymph node metastasis,and mostly pathological stage I-II(all P<0.05).K(N)RAS-mutated MSI-H colorectal cancers were not associated with gender,age,gross type,or histological differentiation(all P>0.05).RAS wild-type MSI-H colorectal cancers were more common in the right colon,with tumor diameters≥4.5 cm,ulcerative and elevated gross types,mucinous adenocarcinoma histology,and moderate to poor differentiation(all P<0.05),but were not associated with gender,age,lymph node metastasis,or pathological stage(all P>0.05).Conclusion:KRAS Gly mutations and BRAF V600E mutations are associated with MSI-H in colorectal cancer.MSI-H colorectal cancer with RAS Gly mutations has unique clinicopathological characteristics.The RAS-MSI gene status is correlated with clinicopathologic features,and detecting K(N)RAS,BRAF V600E,and MSI can aid in personalized treatment and prognosis evaluation of colorectal cancer patients.