3-甲基腺嘌呤联合化疗对食管癌原位移植瘤治疗作用及机制

Antitumor effect and its mechanism analysis of chloroquine combination with chemotherapy for esophageal cancer in situ transplantation

目的探讨3-甲基腺嘌呤（3-MA）联合紫杉醇及顺铂（TP方案）治疗食管癌原位移植瘤的效果以及作用机制。方法60只食管癌原位移植瘤模型随机分为对照组、3-MA组、TP化疗组和联合治疗组。4组小鼠治疗30 d后，计算肿瘤体积；采用Western blot分析自噬底物p62和微管相关蛋白1轻链3（LC3）的变化，采用激酶反应分析VPS34激酶活性，采用原位缺口末端标记法（TUNEL）染色分析肿瘤细胞的凋亡，采用高效液相色谱法（HPLC）分析小鼠肿瘤细胞中化疗药物的浓度。结果与对照组比较，TP化疗组和联合治疗组肿瘤生长明显减缓（P＝0.002）。与TP化疗组比较，联合治疗组小鼠肿瘤生长速度减缓更明显（P＝0.010）。与对照组比较，3-MA和联合治疗组小鼠肿瘤组织中p62蛋白明显积累（P＝0.000）。与对照组LC3-Ⅱ/β-肌动蛋白（β-actin）比值比较，3-MA和联合治疗组小鼠肿瘤组织中LC3-Ⅱ/β-actin比值明显下降（P＝0.000）。与对照组比较，3-MA和联合治疗组小鼠肿瘤组织中VPS34激酶活性显著下调（P＝0.000）。与TP化疗组VPS34激酶活性比较，3-MA和联合治疗组小鼠肿瘤组织中VPS34激酶活性显著下调（P＝0.000）。与对照组比较，TP化疗组和联合治疗组小鼠肿瘤组织细胞凋亡水平显著增加（P＝0.000）。与TP化疗组比较，联合治疗组小鼠肿瘤细胞凋亡水平显著升高（P＝0.000）。结论3-MA可显著抑制肿瘤细胞自噬，与化疗药物联合可显著提高药物利用率，进而显著提高抗肿瘤效果。

ObjectiveTo explore antitumor effect and its mechanism analysis of Chloroquine combination with chemotherapy for esophageal cancer in situ transplantation.Methods60 esophageal orthotopic transplantation tumor models were randomly divided into control group, 3-methyladenine （3-MA）, paclitaxel and cisplatin combined therapy （TP） chemotherapy group and combination group. After 30 days treatment, tumor volume in four groups was analyzed. Autophagy substrate p62 and microtubule-associated protein 1 light chain 3 （LC3） were analyzed by Western blotting. VPS34 kinase activity was analyzed by kinase reaction. The apoptosis of tumor cells was determined by TdT-mediated dUTP nick end labeling （TUNEL） staining. The concentration of chemotherapeutic drugs in tumor cells of mice was analyzed by high performance liquid chromatography （HPLC） analysis.ResultsCompared with control group, tumor volume in TP chemotherapy group and combination group tumor growth significantly reduced （P=0.002）. Compared with TP chemotherapy group, tumor growth in combination group of mice significantly reduced （P=0.010）. Compared with the control group, p62 protein in tumor tissues in 3-MA and combination group significantly accumulated （P=0.000）. Compared with control group, LC3-Ⅱ/β-actin ratio in 3-MA and combination group obviously decreased （P=0.000）. Compared with the control group, VPS34 kinase activity in 3-MA and combination group obviously decreased （P=0.000）. Compared with control group, tumor cell apoptosis levels in TP chemotherapy group and combination group significantly increased （P=0.000）. Compared with TP chemotherapy group, the levels of tumor cell apoptosis in combination group significantly increased （P=0.000）.Conclusion3-MA can inhibit tumor cell autophagy, improve the utilization rate of drugs and enhance the anti-tumor effect.