Quantitative Interrogation of the Human Kinome Perturbed by Two BRAF Inhibitors

Oncogenic BRAF mutations contribute to the development of a number of cancers, and small-molecule BRAF inhibitors have been approved by the Food and Drug Administration (FDA) for anticancer therapy. In this study, we employed two targeted quantitative proteomics approaches for monitoring separately the alterations in protein expression and ATP binding affinities of kinases in cultured human melanoma cells elicited by two FDA-approved small-molecule BRAF inhibitors, dabrafenib and vemurafenib. Our results showed that treatment with the two inhibitors led to markedly different reprograming of the human kinome. Furthermore, we confirmed that vemurafenib could compromise the ATP binding capacity of MAP2K5 in vitro and inhibit its kinase activity in cells. Together, our targeted quantitative proteomic methods revealed profound changes in expression levels of kinase proteins in cultured melanoma cells upon treatment with clinically used BRAF inhibitors and led to the discovery of novel putative target kinases for these inhibitors.致癌性BRAF突变可诱发多种癌症的发生，小分子BRAF抑制剂已获美国食品药品监督管理局（Food and Drug Administration, FDA）批准用于抗癌治疗。本研究采用两种靶向定量蛋白质组学方法，分别监测两种经FDA批准的小分子BRAF抑制剂——达拉非尼（dabrafenib）与维莫非尼（vemurafenib）——处理培养的人黑色素瘤细胞后，细胞内激酶的蛋白表达水平与ATP结合亲和力的变化情况。研究结果显示，两种抑制剂的处理会使人激酶组发生显著不同的重编程过程。此外，本研究证实维莫非尼可在体外削弱MAP2K5的ATP结合能力，并在细胞内抑制其激酶活性。综上，本研究采用的靶向定量蛋白质组学方法揭示了临床使用的BRAF抑制剂处理培养的黑色素瘤细胞后，细胞内激酶蛋白表达水平发生的显著变化，并发现了这两类抑制剂的新型潜在靶点激酶。