Abstract
Activating BRAF mutants and fusions signal as RAS-independent constitutively active dimers with the exception of BRAF V600 mutant alleles which can function as active monomers1. Current RAF inhibitors are monomer selective, they potently inhibit BRAF V600 monomers but their inhibition of RAF dimers is limited by induction of negative cooperativity when bound to one site in the dimer1,2,3. Moreover, acquired resistance to these drugs is usually due to molecular lesions that cause V600 mutants to dimerize4,5,6,7,8. We show here that PLX8394, a new RAF inhibitor9, inhibits ERK signaling by specifically disrupting BRAF-containing dimers, including BRAF homodimers and BRAF–CRAF heterodimers, but not CRAF homodimers or ARAF-containing dimers. Differences in the amino acid residues in the amino (N)-terminal portion of the kinase domain of RAF isoforms are responsible for this differential vulnerability. As a BRAF-specific dimer breaker, PLX8394 selectively inhibits ERK signaling in tumors driven by dimeric BRAF mutants, including BRAF fusions and splice variants as well as BRAF V600 monomers, but spares RAF function in normal cells in which CRAF homodimers can drive signaling. Our work suggests that drugs with these properties will be safe and useful for treating tumors driven by activating BRAF mutants or fusions.
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Acknowledgements
We are grateful to L.M. Desrochers, I. Mellinghoff, and A. Ventura for useful discussions. We thank S. Lowe for the vectors of the retrovirus-based inducible expression system. This research was supported by grants (to N.R.) from the National Institutes of Health (NIH) (P01 CA129243; R35 CA210085); the Melanoma Research Alliance (237059 and 348724); the Commonwealth Foundation for Cancer Research and the Center for Experimental Therapeutics at Memorial Sloan Kettering Cancer Center; and the Stand Up To Cancer – American Cancer Society Lung Cancer Dream Team Translational Research Grant (SU2C-AACR-DT17-15). Support was also received from the NIH MSKCC Cancer Center Support Grant P30 CA008748 and T32 CA009207 (to J.T.). Additional funding was provided by a Conquer Cancer Foundation of the American Society of Clinical Oncology Young Investigator Award (to J.T.) and Career Development Award (to R.Y.). We would like to acknowledge the support of the Arlene and Joseph Taub Foundation and of P. and T. McInerney, without whom this work would not have been possible. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. This work was also supported by grants from the Spanish Ministry of Economy and Competitiveness (SAF2011-30173 and SAF2014-59864-R) (to M.B.). M.B. is the recipient of an endowed chair from the AXA Research Fund.
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Z.Y. and N.R. conceived the project and designed the experiments. Z.Y. and N.R. wrote the manuscript. Y.G., W.S., R.Y., C.Z., M.B., D.M.H., and G.B. provided critical revisions of the manuscript. Z.Y., Y.G., W.S., R.Y., J.T., N.N., Y.Z., C.Z., A.R., A.T., N.M.T., R.M., N.A.O., H.Z., Q.C., B.Q., E.d.S., and D.M.H. established the in vitro and in vivo experimental systems, performed the laboratory experiments, and analyzed the results.
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N.R. is on the scientific advisory board (SAB) and receives research funding from Chugai, on the SAB and owns equity in Beigene, and Fortress. N.R. is also on the SAB of Daiichi-Sankyo, Astra-Zeneca-MedImmune, and F-Prime, and is a past SAB member of Millenium-Takeda, Kadmon, Kura, and Araxes. N.R. is a consultant to Novartis, Boehringer Ingelheim, Tarveda, and Foresight and consulted in the last three years with Eli Lilly, Merrimack, Kura Oncology, Araxes, and Kadman. N.R. owns equity in ZaiLab, Kura Oncology, Araxes, and Kadman. N.R. collaborates with Plexxikon. R.Y. has received research support from Array BioPharma, Genentech, GlaxoSmithKline, and Norvatis; received travel expenses from Array BioPharma; and served on the advisory board for GlaxoSmithKline. D.M.H. has consulted for Atara Biotherapeutics, Chugai Pharma, CytomX Therapeutics, Boehringer Ingelheim, AstraZeneca, Pfizer, Bayer, Debiopharm Group, and Genetech and has received research funding from AstraZeneca, Puma Biotechnology, and Loxo Oncology. Y.Z., C.Z., A.R., and G.B. are employees of Plexxikon Inc.
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Yao, Z., Gao, Y., Su, W. et al. RAF inhibitor PLX8394 selectively disrupts BRAF dimers and RAS-independent BRAF-mutant-driven signaling. Nat Med 25, 284–291 (2019). https://doi.org/10.1038/s41591-018-0274-5
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