Liu Yang, Liu Hailan, Han Baoguang, Zhang Jian-Ting
Department of Pharmacology and Toxicology, Walther Oncology Center/Walther Cancer Institute and IU Cancer Center, Indiana University School of Medicine, Indianapolis, Indiana 46202, USA.
Cancer Res. 2006 Mar 15;66(6):3248-55. doi: 10.1158/0008-5472.CAN-05-3801.
Multidrug resistance (MDR) is a major obstacle to successful cancer treatment. To understand the mechanism of MDR, many cancer cell lines have been established, and various mechanisms of resistance, such as ATP-binding cassette (ABC) transporter-mediated drug efflux, have been discovered. Previously, a MDR cell line MCF7/AdVp3000 was selected from breast cancer cell line MCF7 against Adriamycin, and overexpression of ABCG2 was thought to cause MDR in this derivative cell line. However, ectopic overexpression of ABCG2 in MCF7 cells could not explain the extremely high drug resistance level of the selected MCF7/AdVp3000 cells. We hypothesized that MCF7/AdVp3000 cells must have other resistance mechanisms selected by Adriamycin. To test this hypothesis, we compared the global protein profiles between MCF7 and MCF7/AdVp3000 cells. Following two-dimensional gel electrophoresis and matrix-assisted laser desorption/ionization-time of flight mass spectrometry analysis, 17 protein spots with differential levels between the two cell lines were identified. Although 14-3-3sigma, keratin 18, keratin 19, ATP synthase beta, protein disulfide isomerase, heat shock protein 27, cathepsin D, triose-phosphate isomerase, peroxiredoxin 6, and electron transfer flavoprotein were increased, nm23/H1, peroxiredoxin 2, nucleophosmin 1/B23, and inorganic pyrophosphatase were decreased in MCF7/AdVp3000 cells. The differential levels of these proteins were validated using Western blot. Furthermore, functional validation showed that the elevated 14-3-3sigma expression contributes considerably to the observed drug resistance in MCF7/AdVp3000 cells. We, thus, conclude that these proteins likely contribute to the resistance selected in the MCF7/AdVp3000 cells, and their altered expression in tumors may cause clinical resistance to chemotherapy.
多药耐药（MDR）是癌症治疗成功的主要障碍。为了了解MDR的机制，已经建立了许多癌细胞系，并发现了各种耐药机制，如ATP结合盒（ABC）转运蛋白介导的药物外排。此前，从乳腺癌细胞系MCF7中筛选出对阿霉素耐药的MDR细胞系MCF7/AdVp3000，ABCG2的过表达被认为是该衍生细胞系产生MDR的原因。然而，MCF7细胞中ABCG2的异位过表达并不能解释所选MCF7/AdVp3000细胞极高的耐药水平。我们推测MCF7/AdVp3000细胞必定具有其他由阿霉素选择的耐药机制。为了验证这一假设，我们比较了MCF7和MCF7/AdVp3000细胞之间的整体蛋白质谱。经过二维凝胶电泳和基质辅助激光解吸/电离飞行时间质谱分析，鉴定出两个细胞系之间17个蛋白点水平存在差异。虽然在MCF7/AdVp3000细胞中14-3-3σ、角蛋白18、角蛋白19、ATP合酶β、蛋白质二硫键异构酶、热休克蛋白27、组织蛋白酶D、磷酸丙糖异构酶、过氧化物酶体增殖物激活受体6和电子传递黄素蛋白增加，但nm23/H1、过氧化物酶体增殖物激活受体2、核磷蛋白1/B23和无机焦磷酸酶减少。这些蛋白质的差异水平通过蛋白质印迹法得到验证。此外，功能验证表明，14-3-3σ表达的升高对MCF7/AdVp3000细胞中观察到的耐药性有很大贡献。因此，我们得出结论，这些蛋白质可能对MCF7/AdVp3000细胞中选择的耐药性有贡献，它们在肿瘤中的表达改变可能导致临床化疗耐药。
