Extracellular signal-regulated kinases 1 and 2 (ERK1/2) are ubiquitously expressed in almost all tissues and cell types. Accordingly, ERK1/2 regulate a diversity of cellular processes by phosphorylating more than 150 substrate proteins. With its constitutive activation being frequently implicated in human malignancies, ERK has been an appealing target in anti-cancer drug development. However, currently available ERK1/2 inhibitors are ATP-competitive. Since these ERK inhibitors block all ERK functions, the potential for side effects in normal tissues and cells may limit their potential of being developed into therapeutic agents. To overcome this limitation, we used a combination of computational and experimental methods to develop small molecular weight inhibitors that target ERK-substrate docking domains. Here we report the identification, characterization and optimization of a compound with a thienyl benzenesulfonate scaffold that specifically inhibits substrates containing an F-site or DEF (docking site for ERK, FXF) motif. Our X-ray crystallography data showed that this compound bound to a pocket in the vicinity of the F-site docking domain on ERK. Biological analysis further demonstrated that this compound and its analogs could preferentially inhibit phosphorylation and function of F-site-containing ERK substrates, and that these compounds preferentially inhibited growth of melanoma cells harboring a V600E B-Raf mutant. Additionally, this compound was shown to inhibit the phosphorylation of specific PKC isoforms via an ERK dependent mechanism. These findings represent the first identification and validation of chemicals that selectively inhibit ERK substrates and signaling events through ATP-independent mechanisms. These compounds have potential utility for elucidating the complex biological roles of ERK1/2 and development into novel anti-cancer agents.

Citation Format: Jun Zhang, Taiji Oashi, Ramin Samadani, Kerrick Nevels, Kimberly Burkhard, Deva Priyakumar, Prabhu Raman, Steven Fletcher, Edvin Pozharsky, Paul Shapiro, Alexander MacKerell. A docking domain directed inhibitor targeting ERK regulated F-site substrates. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4554. doi:10.1158/1538-7445.AM2013-4554