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Data and code from: RAF isoform selectivity of MEK inhibitors and rational design of a covalent ARAF-MEK inhibitor

Eck, Michael1; Chakraborty, Sayan1; Jiang, Jie1; Scott, David1; Jeon, Hyesung1; Jänne, Pasi1

Published Oct 02, 2026 on Dryad. https://doi.org/10.5061/dryad.h1893201p

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Abstract

Aberrant activation of the RAS/RAF/MEK/ERK pathway is a frequent cause of cancer. Allosteric MEK inhibitors block this pathway by binding RAF-MEK complexes to prevent activation of MEK by RAF. However, how MEK inhibitor potency varies across the three RAF isoforms remains poorly understood. We profiled seven allosteric MEK inhibitors and observed a striking hierarchy of sensitivity: all most potently inhibited CRAF-driven MEK activation while relatively sparing ARAF-driven activation. Point mutations near the inhibitor binding site in ARAF and CRAF markedly altered inhibitor sensitivity. Through rational design we developed TWG-07-148, a more potent ARAF-MEK inhibitor. Our cryo-EM structure shows how this acrylamide-containing analog of MEK inhibitor trametinib covalently targets Cys514, a residue unique to ARAF. Our studies highlight the importance of the activating RAF isoform as a determinant of MEK inhibitor sensitivity and provide proof-of-concept for development of MEK inhibitors that more effectively block ARAF-driven signaling via covalent targeting of Cys514.