Objective:Key genes were screened to analyze molecular mechanisms and their drug targets of endometriosis by applying a bioinformatics approach.Methods:Gene expression profiles of endometriosis and healthy controls we...Objective:Key genes were screened to analyze molecular mechanisms and their drug targets of endometriosis by applying a bioinformatics approach.Methods:Gene expression profiles of endometriosis and healthy controls were obtained from the Gene Expression Omnibus database.Significant differentially expressed genes were screened using the limma package.Correlation pathways were screened by Spearman correlation analysis on the echinoderm microtubule-associated protein-like 4(EML4)and enrichment in endometriosis pathways and estimated by the GSVA package.Immune characteristics were assessed by the“ESTIMATE”R package.Potential regulatory pathways were determined by enrichment analysis.The SWISS-MODE website was used in homology modeling with EML4 and EML4 protein activity was predicted.VarElect was employed in molecular docking for screening potential compound inhibitors targeting endometriosis.Results:Ten endometriosis and 10 normal samples were included.EML4 was significantly upregulated in endometriosis(p<0.05).Thirty significantly correlated pathways involving 18 positive and 12 negative correlations,including GLYCOSAMINOGLYCAN_BIOSYNTHESIS_HEPARAN_SULFATE and GLYCOSPHINGOLIPID_BIOSYNTHESIS_GANGLIO_SERIES were screened between EML4 and endometriosis.Immunocorrelation analysis showed a significant difference in immune-related pathways in endometriosis and normal samples(p<0.05).In endometriosis,EML4 was associated with T-cell CD4 resting memory,activated mast cells,plasma cells,activated NK cells,M2 macrophages,and follicular helper T cells(p<0.05).Molecular docking identified five potential inhibitors of EML4,and compound DB05104(asimadoline)bound well to EML4 protein to exert its physiological effects.Conclusion:Differential gene expression and immune correlation analyses revealed that EML4 may affect endometriosis through multiple targets and pathways,the mechanism of which involved immune cell activation and infiltration.Molecular docking and dynamics simulation verified DB05104 as a potential inhibitor of EML4 and a powerful target for endometriosis treatment.展开更多
背景与目的小分子靶向药物发生耐药的机制及寻找克服耐药的手段是目前提高临床疗效需要迫切解决的问题。本研究探讨采用不同方法建立对Crizotinib耐药的非小细胞肺癌NCI-H2228/Crizotinib细胞株的可行性及鉴定分析,为深入研究Crizotini...背景与目的小分子靶向药物发生耐药的机制及寻找克服耐药的手段是目前提高临床疗效需要迫切解决的问题。本研究探讨采用不同方法建立对Crizotinib耐药的非小细胞肺癌NCI-H2228/Crizotinib细胞株的可行性及鉴定分析,为深入研究Crizotinib耐药发生的机制并寻找克服耐药的手段提供实验基础和理论依据。方法采用逐步增加药物浓度和化学诱变剂处理NCI-H2228细胞,诱导细胞对Crizotinib耐药。MTT法检测亲本细胞和耐药细胞的50%抑制浓度(50%inhibitory concentration,IC50)和群体倍增时间。RT-PCR和Western blot实验检测棘皮动物微管相关蛋白样4-间变性淋巴瘤激酶(echinoderm microtubule-associated protein like 4-anaplastic lymphoma kinase,EML4-ALK)基因表达。对耐药细胞和亲本细胞的EML4-ALK基因全长测序并对比分析发生耐药的机制。结果逐步增加药物浓度的方法耗时过长,细胞恢复生长缓慢,不能有效诱导NCI-H2228细胞对Crizotinib耐药;化学诱变剂ENU可以在短时间内诱导NCI-H2228细胞对Crizotinib耐药[IC50=(3.810±1.100)μmol/L,P=0.002,9,vs亲本细胞]。耐药细胞EML4-ALK基因发生点突变的频率高于亲本细胞。结论化学诱变剂诱导细胞耐药操作简便,可有效缩短实验流程,为深入研究耐药发生机制,寻找克服靶向药物耐药的手段提供了前期技术方法和实验依据。展开更多
基金funded by the Role and Mechanism of EML4 in Regulating Oocyte Meiosis and Leading to the Infertility Project(SDFEYJGL2103).
文摘Objective:Key genes were screened to analyze molecular mechanisms and their drug targets of endometriosis by applying a bioinformatics approach.Methods:Gene expression profiles of endometriosis and healthy controls were obtained from the Gene Expression Omnibus database.Significant differentially expressed genes were screened using the limma package.Correlation pathways were screened by Spearman correlation analysis on the echinoderm microtubule-associated protein-like 4(EML4)and enrichment in endometriosis pathways and estimated by the GSVA package.Immune characteristics were assessed by the“ESTIMATE”R package.Potential regulatory pathways were determined by enrichment analysis.The SWISS-MODE website was used in homology modeling with EML4 and EML4 protein activity was predicted.VarElect was employed in molecular docking for screening potential compound inhibitors targeting endometriosis.Results:Ten endometriosis and 10 normal samples were included.EML4 was significantly upregulated in endometriosis(p<0.05).Thirty significantly correlated pathways involving 18 positive and 12 negative correlations,including GLYCOSAMINOGLYCAN_BIOSYNTHESIS_HEPARAN_SULFATE and GLYCOSPHINGOLIPID_BIOSYNTHESIS_GANGLIO_SERIES were screened between EML4 and endometriosis.Immunocorrelation analysis showed a significant difference in immune-related pathways in endometriosis and normal samples(p<0.05).In endometriosis,EML4 was associated with T-cell CD4 resting memory,activated mast cells,plasma cells,activated NK cells,M2 macrophages,and follicular helper T cells(p<0.05).Molecular docking identified five potential inhibitors of EML4,and compound DB05104(asimadoline)bound well to EML4 protein to exert its physiological effects.Conclusion:Differential gene expression and immune correlation analyses revealed that EML4 may affect endometriosis through multiple targets and pathways,the mechanism of which involved immune cell activation and infiltration.Molecular docking and dynamics simulation verified DB05104 as a potential inhibitor of EML4 and a powerful target for endometriosis treatment.
文摘背景与目的小分子靶向药物发生耐药的机制及寻找克服耐药的手段是目前提高临床疗效需要迫切解决的问题。本研究探讨采用不同方法建立对Crizotinib耐药的非小细胞肺癌NCI-H2228/Crizotinib细胞株的可行性及鉴定分析,为深入研究Crizotinib耐药发生的机制并寻找克服耐药的手段提供实验基础和理论依据。方法采用逐步增加药物浓度和化学诱变剂处理NCI-H2228细胞,诱导细胞对Crizotinib耐药。MTT法检测亲本细胞和耐药细胞的50%抑制浓度(50%inhibitory concentration,IC50)和群体倍增时间。RT-PCR和Western blot实验检测棘皮动物微管相关蛋白样4-间变性淋巴瘤激酶(echinoderm microtubule-associated protein like 4-anaplastic lymphoma kinase,EML4-ALK)基因表达。对耐药细胞和亲本细胞的EML4-ALK基因全长测序并对比分析发生耐药的机制。结果逐步增加药物浓度的方法耗时过长,细胞恢复生长缓慢,不能有效诱导NCI-H2228细胞对Crizotinib耐药;化学诱变剂ENU可以在短时间内诱导NCI-H2228细胞对Crizotinib耐药[IC50=(3.810±1.100)μmol/L,P=0.002,9,vs亲本细胞]。耐药细胞EML4-ALK基因发生点突变的频率高于亲本细胞。结论化学诱变剂诱导细胞耐药操作简便,可有效缩短实验流程,为深入研究耐药发生机制,寻找克服靶向药物耐药的手段提供了前期技术方法和实验依据。
