College of Pharmacy, Roseman University of Health Sciences, 10530 Discovery Drive, Las Vegas, NV, 89135, USA.
College of Medicine, Roseman University of Health Sciences, 10530 Discovery Drive, Las Vegas, NV, 89135, USA.
Sci Rep. 2018 Jul 3;8(1):9991. doi: 10.1038/s41598-018-28335-8.
Mitogen-activated protein kinases (MAPKs) play an important role in the regulation of cell proliferation, oncogenic transformation, and drug resistance. This study examined the capability of nanofluidic proteomics to identify aberrations in the MAPK signaling cascade, monitor its drug response, and guide the rational design of intervention strategies. Specifically, the protein post-translational modification (PTM) profiles of MEK1, MEK2, and ERK1/2 were measured in breast carcinoma and breast cancer cell lines. Nanofluidic proteomics revealed hyper-phosphorylation of MAPKs in breast carcinoma and breast cancer cells treated with kinase inhibitors that interfere with cell cycle regulation, such as dinaciclib, an inhibitor of cyclin-dependent kinases, and rigosertib, an inhibitor of polo-like kinase 1. A pMEK1 (Thr286) phosphor-isoform, which serves as a biomarker of cell cycle-regulated negative feedback phosphorylation in breast cancer cells, was detected in breast carcinoma. Inhibition of the MAPK pathway with dabrafenib, a B-Raf inhibitor, or trametinib, a MEK1/2 inhibitor, suppressed both the positively regulated phosphorylation of MAPKs and the negatively regulated phosphorylation of MEK1. Interestingly, the combinations of dabrafenib and rigosertib or trametinib and rigosertib permitted the suppression of positively regulated MAPK phosphorylation together with the promotion of negatively regulated MEK1 phosphorylation. The effectiveness of protein PTM-guided drug combinations for inhibition of the MAPK pathway remains to be experimentally tested. Via protein PTM profiling, nanofluidic proteomics provides a robust means to detect anomalies in the MAPK signaling cascade, monitor its drug response, and guide the possible design of drug combinations for MAPK pathway-focused targeting.
丝裂原活化蛋白激酶（MAPKs）在细胞增殖、致癌转化和耐药性的调节中发挥着重要作用。本研究考察了纳流控蛋白质组学在鉴定 MAPK 信号级联中的异常、监测其药物反应以及指导基于 MAPK 通路的干预策略的合理设计中的能力。具体来说，测量了 MEK1、MEK2 和 ERK1/2 的蛋白翻译后修饰（PTM）谱，这些蛋白存在于乳腺癌和乳腺癌细胞系中。纳流控蛋白质组学揭示了在乳腺癌和乳腺癌细胞中，激酶抑制剂处理后 MAPKs 的过度磷酸化，这些激酶抑制剂干扰细胞周期调节，如抑制周期蛋白依赖性激酶的 dinaciclib 和抑制 Polo 样激酶 1 的 rigosertib。在乳腺癌细胞中检测到 pMEK1（Thr286）磷酸化同工型，它是乳腺癌细胞中细胞周期调控负反馈磷酸化的生物标志物。用 B-Raf 抑制剂 dabrafenib 或 MEK1/2 抑制剂 trametinib 抑制 MAPK 通路，抑制了 MAPKs 的正向调节磷酸化和 MEK1 的负向调节磷酸化。有趣的是，dabrafenib 和 rigosertib 或 trametinib 和 rigosertib 的组合允许抑制正向调节的 MAPK 磷酸化，同时促进负向调节的 MEK1 磷酸化。蛋白质 PTM 指导的药物组合抑制 MAPK 通路的有效性仍有待实验验证。通过蛋白质 PTM 谱分析，纳流控蛋白质组学为检测 MAPK 信号级联中的异常、监测其药物反应以及指导针对 MAPK 通路的药物组合设计提供了一种强大的方法。
