American Association for Cancer Research
10780432ccr173674-sup-193706_2_supp_4715385_p7rnjp.ppt (2.02 MB)

Supplementary Figure S1-S6 from ERK Mutations and Amplification Confer Resistance to ERK-Inhibitor Therapy

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posted on 2023-03-31, 21:48 authored by Bijay S. Jaiswal, Steffen Durinck, Eric W. Stawiski, Jianping Yin, Weiru Wang, Eva Lin, John Moffat, Scott E. Martin, Zora Modrusan, Somasekar Seshagiri

Fig. S1. ERK inhibitors and their effect on MAPK mutant cell lines. A. Structure of ERK inhibitors used in this study. B. Dose response curve of ERK inhibitors on the viability of A375, IPC298, SKMEL30, HCT116, MIA PaCa2 and Panc1 cells. Fig. S2. Western blot analysis of parental, VI-3-R and G994-R HCT116 (A) or MIA PaCa2 (B) cells treated with indicated ERK inhibitors. Fig. S3. Western blot analysis of HCT116 cells (A) and SKMEL30 (B) stably expressing indicated ERK1 or ERK2 mutants. Fig. S4. Copy number and expression analysis of MITF in SKMEL30-ERKi-R cells. A. MITF copy gain in SKMEL30-V11e-R, -VI-3-R and -G994-R. B. MITF copy number vs. expression in SKMEL30-V11e-R, VI-3-R and G994-R cells. Fig. S5. Dose response curve of ERK2 overexpressing IPC298 cells treated with MEK inhibitors GDC-0973 or AZD6244 (A) and ERK2 amplified IPC298-G994-R (B) or MIA PaCa2-S984-R (C) cells treated with indicated ERK-inhibitors from alternate scaffold class. Fig. S6. Dose response curve of parental or ERKi-resistant HCT116 (A) and MIA PaCa2 (B) cells treated with indicated inhibitors. The data shown are same as in Fig. 6B for HCT116 (A) and Fig. 6C for MIA PaCa2 (B) grouped by drug treatment.



Purpose: MAPK pathway inhibitors targeting BRAF and MEK have shown clinical efficacy in patients with RAF- and/or RAS-mutated tumors. However, acquired resistance to these agents has been an impediment to improved long-term survival in the clinic. In such cases, targeting ERK downstream of BRAF/MEK has been proposed as a potential strategy for overcoming acquired resistance. Preclinical studies suggest that ERK inhibitors are effective at inhibiting BRAF/RAS-mutated tumor growth and overcome BRAF or/and MEK inhibitor resistance. However, as observed with other MAPK pathway inhibitors, treatment with ERK inhibitors is likely to cause resistance in the clinic. Here, we aimed to model the mechanism of resistance to ERK inhibitors.Experimental Design: We tested five structurally different ATP-competitive ERK inhibitors representing three different scaffolds on BRAF/RAS-mutant cancer cell lines of different tissue types to generate resistant lines. We have used in vitro modeling, structural biology, and genomic analysis to understand the development of resistance to ERK inhibitors and the mechanisms leading to it.Results: We have identified mutations in ERK1/2, amplification and overexpression of ERK2, and overexpression of EGFR/ERBB2 as mechanisms of acquired resistance. Structural analysis of ERK showed that specific compounds that induced on-target ERK mutations were impaired in their ability to bind mutant ERK. We show that in addition to MEK inhibitors, ERBB receptor and PI3K/mTOR pathway inhibitors are effective in overcoming ERK-inhibitor resistance.Conclusions: These findings suggest that combination therapy with MEK or ERBB receptor or PI3K/mTOR and ERK inhibitors may be an effective strategy for managing the emergence of resistance in the clinic. Clin Cancer Res; 24(16); 4044–55. ©2018 AACR.