The Group of Biomaterials and Nanotechnology for Improved Medicines (BIONIMED), Department of Pharmaceutical Technology, Faculty of Pharmacy and Biochemistry, University of Buenos Aires, 956 Junín St, Sixth Floor, Buenos Aires CP1113, Argentina.
Mol Pharm. 2011 Aug 1;8(4):1152-64. doi: 10.1021/mp2000132. Epub 2011 Jun 7.
Primary hepatocellular carcinoma is the third most common fatal cancer worldwide with more than 500,000 annual deaths. Approximately 40% of the patients with HCC showed tumoral overexpression of transmembrane proteins belonging to the ATP-binding cassette protein superfamily (ABC) which pump drugs out of cells. The overexpression of these efflux transporters confers on the cells a multiple drug resistance phenotype, which is considered a crucial cause of treatment refractoriness in patients with cancer. The aim of this study was to investigate the inhibitory effect of different concentrations of pH- and temperature-responsive X-shaped poly(ethylene oxide)-poly(propylene oxide) block copolymers (poloxamines, Tetronic, PEO-PPO) showing a wide range of molecular weights and EO/PO ratios on the functional activity of three different ABC proteins, namely P-glycoprotein (P-gp or MDR1), breast cancer resistance protein (BCRP), and multidrug resistance-associated protein MRP1, in two human hepatocarcinoma cell lines, HepG2 and Huh7. First, the cytotoxicity of the different copolymers (at different concentrations) on both liver carcinoma cell lines was thoroughly evaluated by means of apoptosis analysis using annexin V and propidium iodide (PI). Thus, viable cells (AV-/PI-), early apoptotic cells (AV+/PI-) and late apoptotic cells (V-FITC+/PI+) were identified. Results pointed out copolymers of intermediate to high hydrophobicity and intermediate molecular weight (e.g., T904) as the most cytotoxic. Then, DiOC2, rhodamine 123 and vinblastine were used as differential substrates of these pumps. HeLa, an epithelial cell line of human cervical cancer that does not express P-gp, was used exclusively as a control and enabled the discerning between P-gp and MRP1 inhibition. Moderate to highly hydrophobic poloxamines T304, T904 and T1301 showed inhibitory activity against P-gp and BCRP but not against MRP1 in both hepatic cell lines. A remarkable dependence of this effect on the copolymer concentration and hydrophobicity was found. No inhibitory effect against these ABC pumps was observed with the hydrophilic T1107. These findings further evidence the potential usefulness of these Trojan horses as both drug nanocarriers and ABC inhibitors in hepatic MDR tumors and infections that involve the activity of these efflux transporters.
原发性肝细胞癌是全球第三大常见致命癌症，每年有超过 50 万人死亡。大约 40%的 HCC 患者表现出跨膜蛋白的肿瘤过度表达，这些蛋白属于三磷酸腺苷结合盒蛋白超家族（ABC），将药物泵出细胞。这些外排转运蛋白的过度表达赋予细胞多药耐药表型，这被认为是癌症患者治疗抵抗的一个重要原因。本研究旨在研究不同浓度的 pH 和温度响应 X 型聚（乙二醇）-聚（丙二醇）嵌段共聚物（泊洛沙姆，Tetronic，PEO-PPO）对三种不同 ABC 蛋白（即 P-糖蛋白（P-gp 或 MDR1）、乳腺癌耐药蛋白（BCRP）和多药耐药相关蛋白 MRP1）功能活性的抑制作用，这三种 ABC 蛋白在两种人肝癌细胞系 HepG2 和 Huh7 中表达。首先，通过使用 Annexin V 和碘化丙啶（PI）进行凋亡分析，彻底评估了不同共聚物（在不同浓度下）对两种肝癌细胞系的细胞毒性。因此，鉴定了存活细胞（AV-/PI-）、早期凋亡细胞（AV+/PI-）和晚期凋亡细胞（V-FITC+/PI+）。结果表明，中等至高度疏水性和中等分子量的共聚物（例如 T904）具有最强的细胞毒性。然后，使用 DiOC2、罗丹明 123 和长春碱作为这些泵的差异底物。人宫颈癌上皮细胞系 HeLa 不表达 P-gp，仅用作对照，能够区分 P-gp 和 MRP1 的抑制作用。中等至高度疏水性泊洛沙姆 T304、T904 和 T1301 在两种肝细胞系中均显示出对 P-gp 和 BCRP 的抑制活性，但对 MRP1 没有抑制作用。发现这种效应强烈依赖于共聚物浓度和疏水性。亲水性 T1107 对这些 ABC 泵没有抑制作用。这些发现进一步证明了这些特洛伊木马作为药物纳米载体和 ABC 抑制剂在肝多药耐药肿瘤和涉及这些外排转运蛋白活性的感染中的潜在用途。
