目的 研究钩枝藤总生物碱对非小细胞肺癌细胞系表达谱的影响，筛选出差异表达基因（differentially expressed genes，DEGs）并对功能基因进行分析和挖掘，以期为钩枝藤抗肺癌的分子机制提供理论依据。方法 将非小细胞肺癌NCI-H1975细胞分为对照组和钩枝藤总生物碱处理组，使用RNA-Seq技术开展全转录组测序，获取DEGs；对DEGs进行基因本体（gene ontology，GO）功能及京都基因与基因组百科全书（Kyoto encyclopedia of genes and genomes，KEGG）通路富集分析；筛选差异基因核心模块并分析其影响功能，进一步分析关键基因对非小细胞肺癌生存期的影响；通过分析钩枝藤总生物碱对非小细胞肺癌免疫细胞肿瘤浸润的情况，确定关键基因表达与免疫浸润水平的相关性。结果 对照组和药物组共检测到2976个DEGs，其中药物组664个DEGs上调，2312个DEGs下调。钩枝藤总生物碱对铁死亡、氧化磷酸化、线粒体、溶酶体、细胞间隙连接、核苷酸结合寡聚化结构域（nucleotide binding oligomerization domain，NOD）样受体等介导的信号通路产生重要影响。在DEGs核心模块中，白细胞介素11（interleukin 11，IL11）的高表达可显著影响非小细胞肺癌患者的生存期，是非小细胞肺癌患者不良预后的关键因素之一；角形四肽重复干扰素诱导蛋白3（interferon induced protein with tetratricopeptide repeats 3，IFIT3）、白细胞介素7受体（interleukin 7 receptor，IL7R）、2',5'-寡腺苷酸合成酶2（2',5'-oligoadenylate synthetase 2，OAS2）、S-腺苷甲硫氨酸基结构域蛋白2（radicalS-adenosyl methionine domain containing 2，RSAD2）、受体转运蛋白4（receptor transporter protein 4，RTP4）、不育α基序结构域9样（sterile alpha motif domain containing 9-like gene，SAMD9L）、X染色体连锁凋亡抑制蛋白相关因子1（X-linked inhibitor of apoptosis associated factor 1，XAF1）的表达水平影响了非小细胞肺癌组织的纯度和免疫细胞浸润的类型，可能改变非小细胞肺癌的免疫微环境。结论 钩枝藤总生物碱能够影响非小细胞肺癌细胞的转录谱，具有潜在的抗非小细胞肺癌活性作用。

Objective To evaluate the effect of total alkaloids of Ancistrocladus tectorius on expression profile of non-small cell lung cancer (NSCLC) cell lines, screen out differentially expressed genes (DEGs) for analysis and mining the functional genes, in order to provide a theoretical basis for exploring the molecular mechanism of total alkaloids of A. tectorius anti-lung cancer. Methods NCI-H1975 cells of NSCLC were treated with total alkaloids of A. tectorius. RNA-Seq technology was used to perform whole transcriptome sequencing to obtain DEGs. Gene ontology (GO) function and Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis for DEGs were performed. Key modules of differential genes were screened and their functions were analyzed, the impact of key genes on the survival of NSCLC were further analyzed. By analyzing of total alkaloids of A. tectorius on NSCLC immune infiltration, correlation of key gene expression and immune infiltration level were determined. Results Transcriptome sequencing showed that total 2976 DEGs were identified of which 664 were up-regulated and 2312 were down-regulated in NCI-H1975 cells treated with total alkaloids of A. tectorius. Total alkaloids of A. tectorius had an important impact on signaling pathways mediated by ferroptosis, oxidative phosphorylation, mitochondria, lysosomes, gap junctions, nucleotide binding oligomerization domain (NOD)-like receptors. In the hub module, high expression of interleukin 11 (IL11) could reduce the overall survival rate of NSCLC, and was one of the poor prognostic factors of NSCLC patients; Interferon induced protein with tetratricopeptide repeats 3 (IFIT3), interleukin 7 receptor (IL7R), 2',5'-oligoadenylate synthetase 2 (OAS2), radical S-adenosyl methionine domain containing 2 (RSAD2), receptor transporter protein 4 (RTP4), sterile alpha motif domain containing 9-like gene (SAMD9L) and X-linked inhibitor of apoptosis associated factor 1 (XAF1) expression levels could affect the purity of NSCLC tissue and type of immune cell infiltration, which may alter the immune microenvironment of NSCLC. Conclusion Total alkaloids of A. tectorius could affect the transcriptional profile of NSCLC cells and have potential anti-NSCLC activity.

R285.5

国家自然科学基金资助项目（82173913）；国家自然科学基金资助项目（82104532）；2020年国家民委中青年英才项目