基于基因表达谱筛选乳腺癌他莫昔芬耐药的生物标志物与治疗药物

Screening biomarkers and therapeutic agents for tamoxifen-resistant breast cancer based on gene expression profile

目的:利用基因表达谱和关联性图谱数据库筛选乳腺癌他莫昔芬耐药的潜在生物标志物和治疗药物。方法:在GEO(Gene Expression Omnibus)数据库中筛选得到他莫昔芬耐药的乳腺癌细胞基因表达谱数据集GSE67916和GSE26459,利用R语言和Bioconductor包筛选差异表达基因。Metascape对差异表达基因进行功能注释,Cytoscape构建差异表达基因编码蛋白互相作用网络图并确定核心基因。利用数据集GSE9893验证核心基因在他莫昔芬耐药患者中的表达及与临床预后的关系。最后将差异表达基因导入关联性图谱数据库筛选他莫昔芬耐药的候选治疗药物并进行细胞实验验证。结果:共获得差异表达基因462个,这些基因显著富集于雄激素响应、糖酵解和胆固醇平衡等通路。ATP-柠檬酸裂解酶(ATP-citrate lyase,ACLY)为乳腺癌他莫昔芬耐药的核心基因,ACLY在耐药患者中的表达水平显著高于敏感患者(P<0.05),ACLY低表达患者无复发生存率显著高于高表达患者(P<0.05)。最终筛选得到匹莫齐特、LY-294002等10种候选治疗药物,经证实LY-294002可有效抑制耐他莫昔芬乳腺癌细胞的增殖。结论:ACLY可作为他莫昔芬耐药的生物标志物。利用基于关联性图谱数据库可快速筛选出他莫昔芬耐药的潜在治疗药物。

Objective： To screen potential biomarkers and therapeutic agents for tamoxifen-resistant breast cancer based on gene expression profile and connectivity map. Methods： GSE67916 and GSE26459 which contain gene expression profiles of tamoxifen-resistant breast cancer cell were downloaded from the Gene Expression Omnibus（GEO） database. Differentially expressed genes（DEGs） were identified using the Bioconductor packages in the R language. Metascape was used to perform functional annotation for DEGs. Cytoscape was used to construct the protein-protein interaction network map of DEGs-coding proteins and then to identify the hub gene. GSE9893 was downloaded to validate the expression difference of hub gene between tamoxifen-sensitive patients and resistant patients and then to investigate the correlation between hub gene expression and relapse-free survival. Finally, potential therapeutic agents were screened by mapping the DEGs to connectivity map database and validated by cell proliferation assay. Results： A total of 462 genes were filtered as DEGs and they were mainly involved in androgen response, glycolysis, cholesterol homeostasis and so on. ATP-citrate lyase（ACLY） was identified as the hub gene related to tamoxifen resistance. The expression level of ACLY in tamoxifen-resistant patients was significantly higher than that in tamoxifen-sensitive patients（P0.05）. The relapse-free survival in patients with low expression levels of ACLY was significantly better than that in patients with high expression levels（P0.05）. Ten candidate therapeutic agents were screened for tamoxifen resistance, such as pimozide, LY-294002 and so on. LY-294002 was confirmed to inhibit the proliferation of tamoxifen-resistant breast cancer cells. Conclusion： ACLY may be used as a biomarker for tamoxifen resistant breast cancer. Connectivity map can be used to rapidly discover potential therapeutic agents and lay the foundation for further studies.