Abstract 2843: Resistance to ERK inhibitors as a result of an acquired novel P-loop mutation of ERK2
Authors: Clarke Paul A. , Roe Toby , Swabey Kate , McAndrew Craig , Boxall Kathy , Westwood Isaac , van Montfort Robert , Al-Lazikhani Bissan , Workman Paul 
Published: August 2015

Abstract
Genomic aberrations frequently target and activate the MAPK and PI3K pathways in human cancers and combined inhibition of these pathways is being explored widely in the clinic. In a panel of 45 human colorectal cancer cell lines we found the presence of a KRAS mutation resulted in reduced sensitivity to PI3K and MEK inhibitors when used as single treatments. In addition, prolonged exposure to the individual PI3K (GDC-0941) or MEK (GDC-0973) inhibitors resulted in resistance that was associated with acquisition of mutations of the respective pathway being targeted. Resistance to individual targeted agents is challenge in clinic and can be overcome by combining treatments and in our colorectal cancer panel combined MEK/PI3K inhibition exhibited potent synergy in the KRAS mutant lines. However, as with the single treatments prolonged exposure to the combination regime also resulted in resistance through acquisition of a mutation of the allosteric drug-binding site of MEK2 that abrogates binding of the inhibitor. The clinical development of small-molecule ERK inhibitors is of considerable interest as ERK signaling represents a key downstream effector of RAS mutations and ERK inhibitors may retain activity in cells resistant to RAF and MEK inhibitors. Consistent with the latter, the combination resistant line retained sensitivity to an ERK inhibitor (VTX-11e) that also rescued the synergistic interaction when combined with PI3K inhibition. However, prolonged exposure of this MEK/PI3K resistant line to the combination of VTX-11e/GDC0941 resulted in the acquisition of additional resistance to the ERK inhibitor component of the combination. Resistance to VTX-11e was a result of an ERK2 mutation, novel to protein kinases, in a conserved residue of the kinase domain P-loop. Recombinant wild-type and P-loop mutant ERK2 had similar Vmax and ATP Km values, but exhibited a 20-fold increase in IC50 for VTX-11e. Stable expression of the P-loop mutant ERK2 in parent cells resulted in resistance to VTX-11e. Development of an additional VTX-11e resistant line, in this case by prolonged exposure to a single treatment resulted in the acquisition of the same ERK2 mutation. The repeated development of resistance to the MEK/ERK inhibitor arm of the combination is intriguing as the parent cells were not particularly sensitive to treatment with MEK or ERK inhibitors alone, with a GI50 30-fold higher than BRAF mutant cell lines that are sensitive to these inhibitors. Our data show that dual combination of agents, at least for MEK and PI3K pathway inhibition, is insufficient to block the acquisition of resistance, an observation similar to that of clinical trials of BRAF and MEK inhibitors in melanoma. Finally we recommend sequencing of P-loop of the ERKs in clinical studies of ERK inhibitors to determine whether these mutations will occur in the clinic.
Citation Format: Paul A. Clarke, Toby Roe, Kate Swabey, Craig McAndrew, Kathy Boxall, Isaac Westwood, Robert van Montfort, Bissan Al-Lazikhani, Paul Workman. Resistance to ERK inhibitors as a result of an acquired novel P-loop mutation of ERK2. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2843. doi:10.1158/1538-7445.AM2015-2843