目的体外评价新型微管靶向化合物WX-127-07的抗肿瘤活性,并探讨其作用机制。方法以3种微管靶向药物紫杉醇、长春新碱和秋水仙碱为阳性对照,化合物WX-127-07在0.3~300 nmol·L-1浓度下作用于5种细胞,72 h后用SRB法检测细胞增殖抑制活性,用流式细胞术检测细胞周期阻滞,用高内涵分析(HCA)技术分析该化合物的细胞毒性特征及其与肿瘤和炎症等相关的信号转导途径,用秋水仙碱/荧光-长春碱竞争实验和胰酶消化微管蛋白实验在分子水平测定该化合物的微管蛋白结合位点。结果WX-127-07对肝癌HepG2细胞、宫颈癌HeLa细胞、非小细胞肺癌A549细胞、人胚肺成纤维细胞及脐静脉血管内皮细胞增殖均具有明显的抑制活性,IC50分别为4.47±0.05,5.18±0.08,4.90±0.19,4.10±0.16和(5.04±0.08)nmol·L-1,低于秋水仙碱〔IC50分别为21.17±1.22,14.19±0.53,43.80±1.64,145.89±10.97和(27.67±1.79)nmol·L-1〕和长春新碱〔IC50分别为16.51±0.36,16.76±0.33,27.80±2.75,43.80±1.48和(9.15±0.78)nmol·L-1〕,低于或近似于紫杉醇〔IC50分别为10.68±0.61,12.86±0.25,4.81±0.61,102.07±15.17和(3.04±0.12)nmol·L-1〕。用HCA技术进行细胞多参数分析显示,与长春新碱和秋水仙碱相似,WX-127-07浓度依赖性地诱发A549细胞微管含量降低(P=0.0075),且作用浓度低于对照药物紫杉醇、长春新碱和秋水仙碱;WX-127-07、紫杉醇、长春新碱和秋水仙碱均能诱发A549细胞出现G2/M期阻滞,并使HepG2细胞核膜通透性增加,出现早期凋亡现象,但不影响肿瘤以及炎症相关的信号通路。微管蛋白位点竞争实验结果表明,WX-127-07浓度依赖性地抑制秋水仙碱与微管蛋白的结合(P=0.0259)。化合物与微管蛋白作用后用胰酶消化,其产物经SDS电泳,发现WX-127-07作用后的条带与秋水仙碱相似。结论WX-127-07具有较强的体外抗肿瘤细胞增殖活性,其抗肿瘤作用机制符合微管靶向药物的作用特点,选择性地作用于微管秋水仙碱位点,是新型高活性的微管解聚剂。 Objective To evaluate the antitumor activity of a novel microtubule targeting compound WX-127-07 in vitro and to explore its mechanism. Methods Three kinds of microtubule-targeted drugs, paclitaxel, vincristine and colchicine were used as positive control. Compound WX-127-07 was applied to five kinds of cells at a concentration of 0.3-300 nmol·L-1. After 72 h, Method was used to detect the cell proliferation inhibitory activity. Cell cycle arrest was detected by flow cytometry. Cytotoxicity of the compound and its signal transduction pathways related to tumor and inflammation were analyzed by high content analysis (HCA) Alkaline / Fluorescence-vinblastine competition experiments and pancreatin digestion of tubulin experiments The tubulin binding sites of the compounds were determined at the molecular level. Results WX-127-07 had significant inhibitory activities on the proliferation of HepG2 cells, HeLa cells, non-small cell lung cancer A549 cells, human embryonic lung fibroblasts and umbilical vein endothelial cells with IC50 of 4.47 ± 0.05, 5.18 ± 0.08, 4.90 ± 0.19, 4.10 ± 0.16 and (5.04 ± 0.08) nmol·L-1, respectively, which were lower than colchicine [IC50 was 21.17 ± 1.22, 14.19 ± 0.53, 43.80 ± 1.64, 145.89 ± 10.97 and (27.67 ± 1.79) nmol·L-1〕 and vincristine [IC50 were 16.51 ± 0.36,16.76 ± 0.33,27.80 ± 2.75,43.80 ± 1.48 and (9.15 ± 0.78) nmol·L-1, respectively], lower than or similar to Paclitaxel [IC50 was 10.68 ± 0.61, 12.86 ± 0.25, 4.81 ± 0.61, 102.07 ± 15.17 and (3.04 ± 0.12) nmol·L-1, respectively]. Multi-parameter analysis of HCA cells showed that, compared with vincristine and colchicine, WX-127-07 induced a concentration-dependent decrease of microtubules in A549 cells (P = 0.0075), and its concentration was lower than that of the control drug paclitaxel , Vincristine and colchicine. Both WX-127-07, paclitaxel, vincristine and colchicine could induce G2 / M arrest in A549 cells and increase the nuclear membrane permeability of HepG2 cells with early apoptosis Death phenomenon, but does not affect tumor and inflammation-related signaling pathways. Microtubule site competition experiments showed that WX-127-07 inhibited colchicine-tubulin binding in a concentration-dependent manner (P = 0.0259). Compounds were treated with trypsin and trypsin digestion, the product by SDS electrophoresis and found that the role of WX-127-07 after the band and colchicine similar. Conclusion WX-127-07 has strong anti-tumor cell proliferation activity in vitro. The anti-tumor mechanism of WX-127-07 is in line with the role of microtubule-targeting drugs. It selectively acts on the colchicine colchicine site and is a novel and highly active Microtubule depolymerization.