食管鳞癌患者的基因变异及临床关联分析

Genomic alterations and clinical association analysis in patients with esophageal squamous cell carcinoma

目的:探讨中国食管鳞状细胞癌(esophageal squamous cell carcinoma,ESCC)患者的基因变异(genomic alterations,GAs),以明确ESCC的遗传背景;探讨ESCC患者临床病理特征与GAs及预后的相关性,以发现潜在的预后标志物及新的治疗靶点。方法:选取湖南省肿瘤医院2013年1月—2018年12月行手术切除的ESCC患者49例,收集患者的临床病理特征及随访信息,采用下一代测序技术评估ESCC组织样本中所有类别的GAs,包括胚系变异和体细胞变异(如点突变、短片段插入/缺失、融合/重排、基因拷贝数变异、长片段插入/缺失等)。绘制ESCC患者的GAs景观图,并对临床病理特征、预后与GAs进行关联性分析。结果:在49例ESCC患者中,总共检测到770个GAs,每个患者的GAs个数均在1个以上。其中,GAs个数<10个的病例数为13例,11~20个的病例数为23例,>20个的病例数为13例。GAs个数可能与患者年龄、肿瘤部位、T分期、淋巴结转移和TNM分期有关(P值均<0.05)。突变率在20%以上的基因包括肿瘤蛋白53(tumor protein 53,TP53)、细胞周期依赖性激酶抑制基因2A(cyclin-dependent kinase inhibitor 2A,CDKN2A)、NOTCH1、组蛋白-赖氨酸N-甲基转移酶2D(histone-lysine methyltransferase 2D,KMT2D)、脂肪非典型钙黏蛋白1(FAT atypical cadherin 1,FAT1)和磷酯酰肌醇3-激酶-催化亚单位α基因(phosphatidylinositol 3-hydroxy kinase,catalytic subunit alpha,PIK3CA),扩增率高的基因包括成纤维细胞生长因子(recombinant fibroblast growth factor,FGF)3、FGF4、细胞周期蛋白D1(cyclin D1,CCND1)和FGF19。在有随访数据的42例ESCC患者中,关联分析发现FGF3、FGF4、CCND1和FGF19基因扩增与淋巴结转移和TNM分期之间存在显著相关性(P值均<0.05),NOTCH1基因突变与TNM分期存在显著相关性(P<0.05)。FGF3/FGF4基因扩增组的无病生存率和总生存率均明显低于无扩增组(P值均<0.05),而CCND1/FGF19基因扩增组的无病生存率明显低于无扩增组(P<0.05)。肿瘤位于胸下段较胸上段及胸中段的预后更好(P值均<0.05),浸润深度深、分化程度差和TNM分期晚均提示预后不良(P值均<0.05)。结论:FGF3、FGF4、CCND1和FGF19基因扩增可能可以作为中国ESCC患者个体化分子治疗的生物标志物,为新型靶向治疗和精准医疗的发展奠定基础。

Objective:To investigate the genomic alterations(GAs)in Chinese patients with esophageal squamous cell carcinoma(ESCC)and clarify the genetic background of ESCC.To explore the correlation of clinicopathological characteristics with GAs and prognosis of ESCC patients,in order to find the potential prognostic markers and the new therapeutic targets for guiding targeted therapy of ESCC.Methods:A total of 49 patients with ESCC who underwent surgery at Hunan Cancer Hospital from January 2013 to December 2018 were enrolled in this study.The clinicopathological features and follow-up information of the patients were collected,and the tissue samples were used for next-generation sequencing to assess all types of GAs,including germline and somatic mutations(point mutations,short insertions and deletions,fusions and rearrangements,gene-copy number variation,long segment insertions and deletions,etc.).The landscape of GAs was drawn,and the correlation of clinicopathologic features with prognosis and GAs of the ESCC patients was analyzed.Results:A total of 770 GAs were detected in the 49 patients,and each patient had more than one GA.Among them,there were 13 cases with 0-10 GAs,23 cases with 11-20 GAs,and 13 cases with more than 20 GAs.The number of GAs was related to age,tumor location,T stage,lymph node metastasis,and TNM stage(all P<0.05).Genes with more than 20%mutation rate included tumor protein 53(TP53)(92%,45/49),cyclin-dependent kinase inhibitor 2A(CDKN2A)(43%,21/49),NOTCH1(29%,14/49),histone-lysine methyltransferase 2D(KMT2D)(29%,14/49),FAT atypical cadherin 1(FAT1)(22%,11/49)and phosphatidylinositol 3-hydroxy kinase,catalytic subunit alpha(PIK3CA)(22%,11/49),and the genes with high amplification frequency included recombinant fibroblast growth factor(FGF)3(45%,22/49),FGF4(45%,22/49),cyclin D1(CCND1)(43%,21/49),and FGF19(43%,21/49).In the 42 patients with available follow-up data,the amplification rates of FGF3,FGF4,CCND1 and FGF19 genes were significantly correlated with lymph node metastasis and TNM staging(all P<0.05),and there was a significant correlation between NOTCH1 mutation rate and TNM staging(P<0.05).The disease-free survival and overall survival rates of ESCC patients with FGF3/FGF4 amplification were significantly lower than those of the FGF3/FGF4 non-amplification group(both P<0.05),and the disease-free survival rate of ESCC patients with CCND1/FGF19 amplification was significantly lower than that of the CCND1/FGF19 non-amplification group(P<0.05).Tumors in the lower thoracic segment had better prognosis than those in the upper and middle thoracic segments(both P<0.05).The deeper the infiltration depth,the worse the degree of differentiation,and the later the TNM stage,the worse the prognosis was(all P<0.05).Conclusion:FGF3,FGF4,CCND1 and FGF19 amplification may serve as the biomarkers of individualized molecular therapy for ESCC patients in China,which lays a foundation for the development of novel targeted therapy and precision medicine.