A RAF Inhibitor Model Reveals Allosteric Mechanisms Dictate Activity

RAF inhibitors have been approved for the treatment of BRAF^V600E-mutant tumors, but these inhibitors do not inhibit wild-type BRAF, which acts as a dimer, and RAF dimerization may promote drug resistance. Further, RAF inhibitors paradoxically activate RAF and ERK signaling in cells in which wild-type BRAF dimerization is increased as a result of underlying mechanisms such as RAS mutation. Diverse next-generation RAF inhibitors with varied structural and biochemical properties have been developed, but mechanistic data is lacking to determine the best clinical context for their use, prompting Karoulia, Wu, Ahmed, and colleagues to develop an integrated model of RAF inhibitor action. Comparison of eight structurally diverse RAF inhibitors revealed that five inhibited both monomeric and dimeric BRAF^V600E with similar potency, whereas the other three less potently inhibited dimeric BRAF^V600E. Crystal structures of BRAF bound to inhibitors showed that RAF inhibitors that stabilized the αC helix toward the active IN position (αC-IN) allowed inhibition of both RAF protomers in the dimer by two inhibitor molecules, whereas inhibitors that stabilized the αC helix toward the inactive OUT position, including FDA-approved dabrafenib and vemurafenib, sterically prevented binding of the inhibitor to the second protomer. This suggests that αC-IN inhibitors may more effectively inhibit dimeric RAF, and may guide rational design strategies for selectively targeting dimeric BRAF. Conversely, αC-IN inhibitors promoted RAF binding to RAS (RAF priming) whereas αC-OUT inhibitors did not. This suggests two distinct allosteric mechanisms of RAF inhibitor action: αC-IN inhibitors promote RAF priming and dimer formation and effectively inhibit dimeric RAF, whereas αC-OUT inhibitors promote minimal RAF priming and are ineffective dimeric RAF inhibitors. αC-IN inhibitors exhibited activity in cells expressing various RAF mutations (including BRAF^V600E), and were effective in cells resistant to αC-OUT inhibitors. The identification of allosteric mechanisms of RAF inhibition may guide further rational drug design and clinical application of next-generation RAF inhibitors.

Karoulia Z, Wu Y, Ahmed TA, Xin Q, Bollard J, Krepler C, et al. An integrated model of RAF inhibitor action predicts inhibitor activity against oncogenic BRAF signaling. Cancer Cell 2016 Aug 11 [Epub ahead of print].