Skip to main content

Thank you for visiting You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

RAF inhibitor resistance is mediated by dimerization of aberrantly spliced BRAF(V600E)

This article has been updated


Activated RAS promotes dimerization of members of the RAF kinase family1,2,3. ATP-competitive RAF inhibitors activate ERK signalling4,5,6,7 by transactivating RAF dimers4. In melanomas with mutant BRAF(V600E), levels of RAS activation are low and these drugs bind to BRAF(V600E) monomers and inhibit their activity. This tumour-specific inhibition of ERK signalling results in a broad therapeutic index and RAF inhibitors have remarkable clinical activity in patients with melanomas that harbour mutant BRAF(V600E)8. However, resistance invariably develops. Here, we identify a new resistance mechanism. We find that a subset of cells resistant to vemurafenib (PLX4032, RG7204) express a 61-kDa variant form of BRAF(V600E), p61BRAF(V600E), which lacks exons 4–8, a region that encompasses the RAS-binding domain. p61BRAF(V600E) shows enhanced dimerization in cells with low levels of RAS activation, as compared to full-length BRAF(V600E). In cells in which p61BRAF(V600E) is expressed endogenously or ectopically, ERK signalling is resistant to the RAF inhibitor. Moreover, a mutation that abolishes the dimerization of p61BRAF(V600E) restores its sensitivity to vemurafenib. Finally, we identified BRAF(V600E) splicing variants lacking the RAS-binding domain in the tumours of six of nineteen patients with acquired resistance to vemurafenib. These data support the model that inhibition of ERK signalling by RAF inhibitors is dependent on levels of RAS–GTP too low to support RAF dimerization and identify a novel mechanism of acquired resistance in patients: expression of splicing isoforms of BRAF(V600E) that dimerize in a RAS-independent manner.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Figure 1: Resistance to the RAF inhibitor vemurafenib (PLX4032) is associated with failure of the drug to inhibit ERK signalling.
Figure 2: A BRAF(V600E) variant that lacks exons 4–8 is resistant to the RAF inhibitor vemurafenib.
Figure 3: Identification of splice variants of BRAF(V600E) in human tumours resistant to vemurafenib.

Change history

  • 14 December 2011

    The author affiliation for T.M. was corrected.


  1. 1

    Weber, C. K., Slupsky, J. R., Kalmes, H. A. & Rapp, U. R. Active Ras induces heterodimerization of cRaf and BRaf. Cancer Res. 61, 3595–3598 (2001)

    PubMed  CAS  Google Scholar 

  2. 2

    Rushworth, L. K., Hindley, A. D., O’Neill, E. & Kolch, W. Regulation and role of Raf-1/B-Raf heterodimerization. Mol. Cell Biol. 26, 2262–2272 (2006)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. 3

    Wellbrock, C., Karasarides, M. & Marais, R. The RAF proteins take centre stage. Nature Rev. Mol. Cell Biol. 5, 875–885 (2004)

    Article  CAS  Google Scholar 

  4. 4

    Poulikakos, P. I., Zhang, C., Bollag, G., Shokat, K. M. & Rosen, N. RAF inhibitors transactivate RAF dimers and ERK signalling in cells with wild-type BRAF. Nature 464, 427–430 (2010)

    ADS  Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. 5

    Heidorn, S. J. et al. Kinase-dead BRAF and oncogenic RAS cooperate to drive tumor progression through CRAF. Cell 140, 209–221 (2010)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. 6

    Hatzivassiliou, G. et al. RAF inhibitors prime wild-type RAF to activate the MAPK pathway and enhance growth. Nature 464, 431–435 (2010)

    ADS  Article  CAS  PubMed  Google Scholar 

  7. 7

    Joseph, E. W. et al. The RAF inhibitor PLX4032 inhibits ERK signaling and tumor cell proliferation in a V600E BRAF-selective manner. Proc. Natl Acad. Sci. USA 107, 14903–14908 (2010)

    ADS  Article  PubMed  Google Scholar 

  8. 8

    Flaherty, K. T. et al. Inhibition of mutated, activated BRAF in metastatic melanoma. N. Engl. J. Med. 363, 809–819 (2010)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. 9

    Solit, D. B. et al. BRAF mutation predicts sensitivity to MEK inhibition. Nature 439, 358–362 (2006)

    ADS  Article  CAS  PubMed  Google Scholar 

  10. 10

    Whittaker, S. et al. Gatekeeper mutations mediate resistance to BRAF-targeted therapies. Sci. Transl. Med. 2, 35ra41 (2010)

    Article  CAS  PubMed  Google Scholar 

  11. 11

    Cutler, R. E., Jr, Stephens, R. M., Saracino, M. R. & Morrison, D. K. Autoregulation of the Raf-1 serine/threonine kinase. Proc. Natl Acad. Sci. USA 95, 9214–9219 (1998)

    ADS  Article  CAS  PubMed  Google Scholar 

  12. 12

    Rajakulendran, T., Sahmi, M., Lefrancois, M., Sicheri, F. & Therrien, M. A dimerization-dependent mechanism drives RAF catalytic activation. Nature 461, 542–545 (2009)

    ADS  Article  CAS  PubMed  Google Scholar 

  13. 13

    Poulikakos, P. I. & Rosen, N. Mutant BRAF melanomas–dependence and resistance. Cancer Cell 19, 11–15 (2011)

    Article  CAS  PubMed  Google Scholar 

  14. 14

    McArthur, G. et al. Molecular analyses from a phase I trial of vemurafenib to study mechanism of action (MOA) and resistance in repeated biopsies from BRAF mutation–positive metastatic melanoma patients (pts). J Clin Oncol 29, suppl; abstr 8502. (2011)

  15. 15

    Nazarian, R. et al. Melanomas acquire resistance to B-RAF(V600E) inhibition by RTK or N-RAS upregulation. Nature 468, 973–977 (2010)

    ADS  Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. 16

    Johannessen, C. M. et al. COT drives resistance to RAF inhibition through MAP kinase pathway reactivation. Nature 468, 968–972 (2010)

    ADS  Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. 17

    Villanueva, J. et al. Acquired resistance to BRAF inhibitors mediated by a RAF kinase switch in melanoma can be overcome by cotargeting MEK and IGF-1R/PI3K. Cancer Cell 18 683–695 S1535–6108(10)00484–8 (2010)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. 18

    Wagle, N. et al. Dissecting therapeutic resistance to RAF inhibition in melanoma by tumor genomic profiling. J. Clin. Oncol. 29, 3085–3096 (2011)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. 19

    Luco, R. F., Allo, M., Schor, I. E., Kornblihtt, A. R. & Misteli, T. Epigenetics in alternative pre-mRNA splicing. Cell 144, 16–26 (2011)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. 20

    Bollag, G. et al. Clinical efficacy of a RAF inhibitor needs broad target blockade in BRAF-mutant melanoma. Nature 467, 596–599 (2010)

    ADS  Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. 21

    Janakiraman, M. et al. Genomic and biological characterization of exon 4 KRAS mutations in human cancer. Cancer Res 70, 5901–5911 (2010)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. 22

    Nüsse, M., Beisker, W., Hoffmann, C. & Tarnok, A. Flow cytometric analysis of G1- and G2/M-phase subpopulations in mammalian cell nuclei using side scatter and DNA content measurements. Cytometry 11, 813–821 (1990)

    Article  PubMed  Google Scholar 

Download references


We are grateful to M. Baccarini and C. Pritchard for the RAF knockout mouse embryonic fibroblasts. We would like to thank K. Shokat, C. Zhang, S. Chandarlapaty, C. Pratilas and K. Robzyk for useful discussions, C. Liu for her technical expertise and T. J. Riley. We also thank D. Hucks for technical assistance, P. L. Lyle for scoring the tumour sections, and T. Chodon for assistance with tumour procurement. The ITR is supported by the Vanderbilt-Ingram Cancer Center and the T. J. Martell Foundation. This work has been funded by the National Institutes of Health (NIH; R.S.L., N.R., D.B.S.), the Beene Foundation (D.B.S.), the Melanoma Research Alliance (K.T.F., J.A.S. P.B.C., R.S.L., N.R., D.B.S.) and the STARR Foundation (N.R., D.B.S.). R.S.L. was supported by the Burroughs Wellcome Fund, American Skin Association, Joint Center for Translational Medicine, Sidney Kimmel Foundation, and Stand Up to Cancer. T.M. was supported in part by the Intramural Research Program of the NIH, NCI, Center for Cancer Research and P.B.C. was supported in part by the Danny Federici Melanoma Fund. P.I.P. was supported by T32 CACA062948-15.

Author information




P.I.P., M.J., T.G.G., A.R., R.S.L., N.R. and D.B.S. designed experiments and analysed data. P.I.P., Y.P., M.J., X.K., C.N., G.M., H.S., M.A., B.T., M.T.G., M.S., K.B.D. and M.T. performed experiments and analysed data. J.A.W., K.T.F., M.C.K., J.A.S., P.B.C. and R.S.L. provided tumours for analysis. All authors contributed to the writing of the paper.

Corresponding author

Correspondence to David B. Solit.

Ethics declarations

Competing interests

J.A.S., P.B.C., N.R. and D.B.S. have participated on an advisory board for Roche. J.A.S. and P.B.C. have received clinical trial research funding from Roche. P.B.C. has received research support and served on an advisory board for Genentech. A.R. has received honorarium serving on the advisory boards of Roche-Genentech. N.R. and D.B.S. have received research funding from Astra-Zeneca.

Supplementary information

Supplementary Information

This file contains Supplementary Figures 1-13 with legends and Supplementary Tables 1-2. (PDF 1596 kb)

PowerPoint slides

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Poulikakos, P., Persaud, Y., Janakiraman, M. et al. RAF inhibitor resistance is mediated by dimerization of aberrantly spliced BRAF(V600E). Nature 480, 387–390 (2011).

Download citation

Further reading


By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.


Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing