Division of Pharmacology and Chemotherapy, Department of Internal Medicine, University of Pisa, 56126, Pisa, Italy.
Cancer Chemother Pharmacol. 2010 Aug;66(3):547-58. doi: 10.1007/s00280-009-1195-2. Epub 2009 Dec 18.
Oxaliplatin effect in the treatment of colorectal cancer is improved upon combination with thymidylate synthase (TS) inhibitors. Pemetrexed is polyglutamated by the folylpolyglutamate synthase (FPGS) and blocks folate metabolism and DNA synthesis by inhibiting TS, dihydrofolate reductase (DHFR) and glycinamide ribonucleotide formyltransferase (GARFT). The present study evaluates the pharmacological interaction between oxaliplatin and pemetrexed in colorectal cancer cells.
Human HT29, WiDr, SW620 and LS174T cells were treated with oxaliplatin and pemetrexed. Drug interaction was studied using the combination index method, while cell cycle was investigated with flow cytometry. The effects of drugs on Akt phosphorylation and apoptosis were studied with ELISA and fluorescence microscopy, respectively. RT-PCR analysis was performed to assess whether drugs modulated the expression of pemetrexed targets and of genes involved in DNA repair (ERCC1 and ERCC2). Finally, platinum-DNA adduct levels were detected by ultra-sensitive multi-collector inductively coupled plasma mass spectrometry (ICP-MS).
A dose-dependent inhibition of cell growth was observed after drug exposure, while a synergistic interaction was detected preferentially with sequential combinations. Oxaliplatin enhanced cellular population in the S-phase. Drug combinations increased apoptotic indices with respect to single agents, and both drugs inhibited Akt phosphorylation. RT-PCR analysis showed a correlation between the FPGS/(TS x DHFR x GARFT) ratio and pemetrexed sensitivity, as well as a downregulation of ERCC1, ERCC2, TS, DHFR and GARFT after drug exposure. In addition, pretreatment with pemetrexed resulted in an increase of oxaliplatin-DNA adducts.
These data demonstrate that oxaliplatin and pemetrexed synergistically interact against colon cancer cells, through modulation of cell cycle, inhibition of Akt phosphorylation, induction of apoptosis and modulation of gene expression.
奥沙利铂与胸苷酸合成酶（TS）抑制剂联合应用可提高结直肠癌的疗效。培美曲塞由叶酸多聚谷氨酸合成酶（FPGS）多聚谷氨酸化，通过抑制 TS、二氢叶酸还原酶（DHFR）和甘氨酰胺核苷酸甲酰转移酶（GARFT）来阻断叶酸代谢和 DNA 合成。本研究评估了奥沙利铂和培美曲塞在结直肠癌细胞中的药理相互作用。
用奥沙利铂和培美曲塞处理人 HT29、WiDr、SW620 和 LS174T 细胞。采用组合指数法研究药物相互作用，采用流式细胞术研究细胞周期。用酶联免疫吸附试验（ELISA）和荧光显微镜分别研究药物对 Akt 磷酸化和细胞凋亡的影响。采用 RT-PCR 分析评估药物是否调节培美曲塞靶点和参与 DNA 修复（ERCC1 和 ERCC2）的基因的表达。最后，采用超灵敏多收集电感耦合等离子体质谱法（ICP-MS）检测铂-DNA 加合物水平。
药物暴露后观察到细胞生长呈剂量依赖性抑制，而优先检测到序贯联合的协同作用。奥沙利铂增强了细胞群体在 S 期的分布。药物组合增加了与单药治疗相比的凋亡指数，并且两种药物均抑制 Akt 磷酸化。RT-PCR 分析显示，FPGS/（TS×DHFR×GARFT）比值与培美曲塞敏感性之间存在相关性，以及药物暴露后 ERCC1、ERCC2、TS、DHFR 和 GARFT 的下调。此外，培美曲塞预处理可增加奥沙利铂-DNA 加合物。
这些数据表明，奥沙利铂和培美曲塞通过调节细胞周期、抑制 Akt 磷酸化、诱导细胞凋亡和调节基因表达，协同作用于结肠癌细胞。
