FGFRs家族基因多态性和表达与乳腺癌相关性研究

The over-activation of the fibroblast growth factor receptors (FGFRs) family genes have been shown to be involved in the development and progression of cancer. The associations between the functional polymorphisms of FGFRs family genes and breast cancer risk, and the underlying mechanism still remain unknown. Our previous studies showed that FGFR2 polymorphism was associated with breast cancer risk in a Northern Chinese Han population. And high FGFR2 protein expressions correlated with poor prognosis in invasive ductal cancer patients. In this project, a case-control study is carried out. All the participants are interviewed to collect detailed information on breast cancer risk factors. Venous blood is collected and genotyping is analysed using the SNaPshot method. The association between FGFRs family gene polymorphisms with breast cancer is studied. The mRNA and protein expressions of FGFRs family genes are investigated by tissue microarray coupled with in situ hybridization and immune-histochemical methods respectively in the specimens of breast cancer tissues and adjacent normal tissues,. Then the associations between the mRNA and protein expressions and susceptive single nucleotide polymorphisms (SNPs) and clinical pathological datas and prognosis will be analysed. The biological functions of breast cancer susceptive SNPs are observed by electrophoretic mobility shift assays and luciferase reporter vector transfection experiment technology. This study may provide new insights to further elucidate the pathogenesis of sporadic breast cancer. This would also provide a scientific basis for screening and prevention of high-risk groups of breast cancer and improving patient outcomes.

FGFRs家族基因的异常持续活化被证实参与了肿瘤的发生和发展。目前， FGFRs家族基因功能性多态与乳腺癌易感性分析及功能研究鲜见报道。课题组的前期研究结果表明：FGFRs家族的FGFR2基因多态性与中国北方汉族妇女乳腺癌发病风险相关，其蛋白过表达影响预后。在此基础上，本课题拟采用病例对照研究的方法，调查乳腺癌相关危险因素并采集外周血，应用SNaPshot技术进行基因分型，分析FGFRs家族基因功能性多态位点与乳腺癌的关系。同时收集乳腺癌组织及癌旁正常组织生物样本，应用组织芯片、原位杂交和免疫组化方法测定FGFRs家族基因mRNA和蛋白的表达情况，分析其与乳腺癌相关多态位点、临床病理特征及预后的关系。运用凝胶迁移和荧光素酶报告载体转染实验技术检测FGFRs家族基因乳腺癌相关多态位点的生物学功能。通过上述研究，为阐明乳腺癌的发病机制、筛查乳腺癌高危人群及改善患者的预后提供科学依据。

本研究首先采用问卷调查，获得了调查人群流行病学资料及乳腺癌患者临床病理特征及随访资料，同时收集被调查者的血液样本，结合功能学意义和标签SNP(Tag SNP)策略综合筛选FGFRs家族基因潜在功能性多态位点，应用多重单碱基延伸单核苷酸多态性分型技术(SNaPshot)进行基因分型。从单位点分析、单倍型分析、位点-环境交互作用分析等几个方面综合判定FGFRs家族基因功能性多态位点与中国北方汉族妇女乳腺癌的关系，并进一步分析这些SNPs与乳腺癌临床病理特征及预后之间的相关性。其次，应用免疫组化方法，测定乳腺癌组织中FGFRs家族基因蛋白的表达情况，分析其与乳腺癌临床病理参数及预后的关系，并采用统计软件分析FGFRs家族基因潜在功能性多态位点与蛋白表达情况之间的关系。在此基础上，对FGFR4基因 rs351855多态位点进行了功能学研究。结果显示：病例组的BMI指数、哺乳时间显著高于对照组。FGFR1的 rs17182023 CC基因型能够降低乳腺癌的患病风险。FGFR2的rs1863744、rs2981582、rs1219648和rs3135718位点的变异基因型能够显著增加乳腺癌的发病风险。FGFR4基因rs1966265和 rs351855多态性与乳腺癌患病风险强相关，而且rs351855 与乳腺癌的淋巴结转移相关。FGFR3基因多态性rs2234909 和 rs3135848与该人群乳腺癌发病不相关。FGFR4蛋白在乳腺癌组织中高表达，而且FGFR4蛋白表达与腋窝淋巴结转移、ER及分子分型有关。FGFR4高表达、rs1966265及rs351855与乳腺癌患者的不良预后显著相关，仅腋窝淋巴结转移可作为独立的预后因素。FGFR1 基因多态位点rs17182023、FGFR4 基因多态位点rs351855及HER2基因多态位点rs1058808 和rs2517956可能对其相应的蛋白表达具有影响。FGFR4基因rs351855位点野生型和突变型过表达慢病毒载体转染MDA-MB-231细胞，采用Western Blot方法检测细胞蛋白，结果仅检测出微弱条带。综上，FGFRs家族基因多态性在中国北方汉族人群乳腺癌的发生发展中发挥重要作用，其可以作为该人群乳腺癌潜在的易感标志物。上述研究结果分别发表在Oncotarget和Cancer Epidemiology杂志上。