Dose-dependent proteome profiling of KRAS pathway inhibitors sheds new light on drug mechanism of action and KRAS signaling

Kras is a molecular switch controlling the activity of the MAPK pathway. As such, it determines ERK activation state. In this study we identified many novel p-sites that were regulated upon KRAS inhibition. Many of these regulated p-sites were located within a SP/TP motif, the recognition motif of proline directed Kinases such as ERK. In this study we used parallel reaction monitoring (PRM) to determine the ability of ERK to phosphorylate synthetic peptides carrying a subset of these p-sites in a time-resolved manner. We identified that ERK was able to phosphorylate 10 of the tested sequences of which only one was phosphorylated on a mutated control peptide.