Abstract
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.
Highlights
Mitogen-activated protein kinases (MAPKs) play an important role in the regulation of cell proliferation, oncogenic transformation, and drug resistance
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
Using capillary isoelectric focusing (cIEF) immunoassays, this study reported hyper-phosphorylation of the MAPK pathway in breast carcinoma compared with that in breast cancer cell lines
Summary
Mitogen-activated protein kinases (MAPKs) play an important role in the regulation of cell proliferation, oncogenic transformation, and drug resistance. 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 pathwayfocused targeting. The realization of molecular targeted cancer therapy is hindered by multiple challenges, such as the fact that only some human cancers have known kinase-domain mutations[5,6,7,8] and the rapid development of drug resistance due to intrinsic inter- and intra-tumor heterogeneity[9,10] To overcome such challenges, molecular targeted cancer therapy is being applied more broadly, extending beyond specific oncogenic lesions to encompass aberrant signaling pathways whose components are not necessarily mutated[5]. Nanofluidic proteomics has the potential to be a robust method that can identify cancer phenotypes, assist in the design of pathway-focused therapy, and screen for the molecular effects of individual drugs or drug combinations
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