Department of Chemistry, Xavier University of Louisiana, 1 Drexel Drive, New Orleans, LA 70125, USA.
Breast Cancer Res. 2012 Mar 14;14(2):R45. doi: 10.1186/bcr3144.
Acquired tamoxifen resistance involves complex signaling events that are not yet fully understood. Successful therapeutic intervention to delay the onset of hormone resistance depends critically on mechanistic elucidation of viable molecular targets associated with hormone resistance. This study was undertaken to investigate the global proteomic alterations in a tamoxifen resistant MCF-7 breast cancer cell line obtained by long term treatment of the wild type MCF-7 cell line with 4-hydroxytamoxifen (4-OH Tam).
We cultured MCF-7 cells with 4-OH Tam over a period of 12 months to obtain the resistant cell line. A gel-free, quantitative proteomic method was used to identify and quantify the proteome of the resistant cell line. Nano-flow high-performance liquid chromatography coupled to high resolution Fourier transform mass spectrometry was used to analyze fractionated peptide mixtures that were isobarically labeled from the resistant and control cell lysates. Real time quantitative PCR and Western blots were used to verify selected proteomic changes. Lentiviral vector transduction was used to generate MCF-7 cells stably expressing S100P. Online pathway analysis was performed to assess proteomic signatures in tamoxifen resistance. Survival analysis was done to evaluate clinical relevance of altered proteomic expressions.
Quantitative proteomic analysis revealed a wide breadth of signaling events during transition to acquired tamoxifen resistance. A total of 629 proteins were found significantly changed with 364 up-regulated and 265 down-regulated. Collectively, these changes demonstrated the suppressed state of estrogen receptor (ER) and ER-regulated genes, activated survival signaling and increased migratory capacity of the resistant cell line. The protein S100P was found to play a critical role in conferring tamoxifen resistance and enhanced cell motility.
Our data demonstrate that the adaptive changes in the proteome of tamoxifen resistant breast cancer cells are characterized by down-regulated ER signaling, activation of alternative survival pathways, and enhanced cell motility through regulation of the actin cytoskeleton dynamics. Evidence also emerged that S100P mediates acquired tamoxifen resistance and migration capacity.
获得性他莫昔芬耐药涉及复杂的信号事件，目前尚未完全了解。成功的治疗干预以延迟激素耐药的发生，关键取决于与激素耐药相关的可行分子靶标的机制阐明。本研究旨在研究通过用 4-羟他莫昔芬（4-OH Tam）长期处理野生型 MCF-7 细胞系获得的他莫昔芬耐药 MCF-7 乳腺癌细胞系中的全局蛋白质组变化。
我们用 4-OH Tam 培养 MCF-7 细胞超过 12 个月，以获得耐药细胞系。使用无凝胶、定量蛋白质组学方法来鉴定和定量耐药细胞系的蛋白质组。纳流高效液相色谱与高分辨率傅立叶变换质谱联用，用于分析来自耐药和对照细胞裂解物的等压标记的分级肽混合物。实时定量 PCR 和 Western blot 用于验证选定的蛋白质组变化。慢病毒载体转导用于生成稳定表达 S100P 的 MCF-7 细胞。在线途径分析用于评估他莫昔芬耐药中的蛋白质组特征。生存分析用于评估改变的蛋白质组表达的临床相关性。
定量蛋白质组分析显示，在获得性他莫昔芬耐药过渡期间存在广泛的信号事件。共发现 629 种蛋白质发生显著变化，其中 364 种上调，265 种下调。总的来说，这些变化表明雌激素受体（ER）和 ER 调节基因的抑制状态、存活信号的激活以及耐药细胞系迁移能力的增加。发现蛋白质 S100P 在赋予他莫昔芬耐药性和增强细胞迁移能力方面发挥关键作用。
我们的数据表明，他莫昔芬耐药乳腺癌细胞蛋白质组的适应性变化的特征是 ER 信号转导下调、替代存活途径激活以及通过调节肌动蛋白细胞骨架动力学增强细胞迁移能力。证据还表明，S100P 介导获得性他莫昔芬耐药和迁移能力。
