Ayyagari Vijayalakshmi N, Hsieh Tsung-Han Jeff, Diaz-Sylvester Paula L, Brard Laurent
Division of Gynecologic Oncology; Department of Obstetrics and Gynecology, Southern Illinois University School of Medicine, Springfield, IL, USA.
Center for Clinical Research, Southern Illinois University School of Medicine, Springfield, IL, USA.
BMC Cancer. 2017 Jan 13;17(1):49. doi: 10.1186/s12885-016-3034-2.
Combination drug therapy appears a promising approach to overcome drug resistance and reduce drug-related toxicities in ovarian cancer treatments. In this in vitro study, we evaluated the antitumor efficacy of cisplatin in combination with Bithionol (BT) against a panel of ovarian cancer cell lines with special focus on cisplatin-sensitive and cisplatin-resistant cell lines. The primary objectives of this study are to determine the nature of the interactions between BT and cisplatin and to understand the mechanism(s) of action of BT-cisplatin combination.
The cytotoxic effects of drugs either alone or in combination were evaluated using presto-blue assay. Cellular reactive oxygen species were measured by flow cytometry. Immunoblot analysis was carried out to investigate changes in levels of cleaved PARP, XIAP, bcl-2, bcl-xL, p21 and p27. Luminescent and colorimetric assays were used to test caspases 3/7 and ATX activity.
The efficacy of the BT-cisplatin combination depends upon the cell type and concentrations of cisplatin and BT. In cisplatin-sensitive cell lines, BT and cisplatin were mostly antagonistic except when used at low concentrations, where synergy was observed. In contrast, in cisplatin-resistant cells, BT-cisplatin combination treatment displayed synergistic effects at most of the drug ratios/concentrations. Our results further revealed that the synergistic interaction was linked to increased reactive oxygen species generation and apoptosis. Enhanced apoptosis was correlated with loss of pro-survival factors (XIAP, bcl-2, bcl-xL), expression of pro-apoptotic markers (caspases 3/7, PARP cleavage) and enhanced cell cycle regulators p21 and p27.
In cisplatin-resistant cell lines, BT potentiated cisplatin-induced cytotoxicity at most drug ratios via enhanced ROS generation and modulation of key regulators of apoptosis. Low doses of BT and cisplatin enhanced efficiency of cisplatin treatment in all the ovarian cancer cell lines tested. Our results suggest that novel combinations such as BT and cisplatin might be an attractive therapeutic approach to enhance ovarian cancer chemosensitivity. Combining low doses of cisplatin with subtherapeutic doses of BT can ultimately lead to the development of an innovative combination therapy to reduce/prevent the side effects normally occurring when high doses of cisplatin are administered.
联合药物疗法似乎是克服卵巢癌治疗中耐药性并降低药物相关毒性的一种有前景的方法。在这项体外研究中，我们评估了顺铂与硫双二氯酚（BT）联合对一组卵巢癌细胞系的抗肿瘤疗效，特别关注顺铂敏感和耐药细胞系。本研究的主要目的是确定BT与顺铂之间相互作用的性质，并了解BT-顺铂联合的作用机制。
使用碧云天试剂法评估单独或联合用药的细胞毒性作用。通过流式细胞术测量细胞活性氧。进行免疫印迹分析以研究裂解的PARP、XIAP、bcl-2、bcl-xL、p21和p27水平的变化。使用发光和比色测定法检测半胱天冬酶3/7和自噬素（ATX）活性。
BT-顺铂联合的疗效取决于细胞类型以及顺铂和BT的浓度。在顺铂敏感细胞系中，BT和顺铂大多表现为拮抗作用，低浓度使用时除外，此时观察到协同作用。相反，在顺铂耐药细胞中，BT-顺铂联合治疗在大多数药物比例/浓度下均显示出协同作用。我们的结果进一步表明，协同相互作用与活性氧生成增加和细胞凋亡有关。增强的细胞凋亡与促生存因子（XIAP、bcl-2、bcl-xL）的丧失、促凋亡标志物（半胱天冬酶3/7、PARP裂解）的表达以及细胞周期调节因子p21和p27的增强有关。
在顺铂耐药细胞系中，BT在大多数药物比例下通过增强活性氧生成和调节凋亡关键调节因子增强了顺铂诱导的细胞毒性。低剂量的BT和顺铂提高了所有测试卵巢癌细胞系中顺铂治疗的效率。我们的结果表明，如BT和顺铂这样的新型联合用药可能是增强卵巢癌化疗敏感性的一种有吸引力的治疗方法。将低剂量顺铂与低于治疗剂量的BT联合最终可能导致开发一种创新的联合疗法，以减少/预防高剂量顺铂给药时通常出现的副作用。
