基于转录组学探讨铁死亡相关基因在奥希替尼获得性耐药NSCLC细胞中的表达及意义

Study on expression and significance of ferroptosis related genes in osimertinib acquired-resistant NSCLC cells based on transcriptomics

目的基于转录组学探讨NSCLC细胞奥希替尼(osimertinib,Osi)获得性耐药的关键基因,并分析其在耐药细胞中的意义。方法体外培养人肺癌H1975细胞和Osi耐药H1975/OR细胞,CCK-8法检测Osi和铁死亡诱导剂对两种细胞存活率的影响;转录组学检测两种细胞的基因表达差异情况;Western blot检测人肺成纤维细胞HLF-1与两种肺癌细胞的铁死亡相关蛋白表达水平差异。结果不同浓度的Osi可抑制H1975细胞和H1975/OR细胞的增殖,且H1975细胞更敏感。通过转录组学分析,两种细胞在基因表达水平存在着较大差异,其中铁死亡通路差异较为显著。Western blot结果验证HLF-1、H1975和H1975/OR细胞在铁死亡相关蛋白的表达水平上存在着不同程度的差异。铁死亡诱导剂RSL3和Erastin处理后,耐药H1975/OR细胞对其更敏感,而H1975细胞无显著性差异。结论肺癌细胞H1975和Osi耐药细胞H1975/OR的基因表达水平存在较多差异,其中铁死亡通路对Osi的获得性耐药有着较为重要的作用。

Aim To explore the key genes of osimertinib acquired resistance in non-small cell lung cancer cells based on transcriptomics,and to analyze the significance in osimertinib resistant cells.Methods Human lung cancer H1975 cells and osimertinib resistant H1975/OR cells were cultured in vitro.The effect of osimertinib and ferroptosis inducers on cell viability was measured by CCK-8 assay.The difference in gene expression was measured by transcriptomics.The expression levels of ferroptosis-related proteins were measured by Western blot.Results Osimertinib could inhibit the proliferation of H1975 cells and H1975/OR cells in different concentrations,and H1975 cells were more sensitive.Transcriptomics analysis revealed remarkable differences in gene expression levels between H1975 and H1975/OR cells,among which the ferroptosis pathway was most significant.Western blot assay validated that different degrees of change existed in the expression levels of ferroptosis-related proteins in HLF-1,H1975 and H1975/OR cells.After the treatment of ferroptosis inducers RSL3 and Erastin,H1975/OR cells showed higher sensitivity,while little significance was observed on H1975 cells.Conclusions There were significant differences between H1975 lung cancer cells and osimertinib resistant H1975/OR cells on the gene expression levels,and the ferroptosis pathway plays an important role in acquired resistance to osimertinib.