Targeting the Raf kinases in human cancer: the Raf dimer dilemma

Abstract

The Raf protein kinases are key intermediates in cellular signal transduction, functioning as direct effectors of the Ras GTPases and as the initiating kinases in the ERK cascade. In human cancer, Raf activity is frequently dysregulated due to mutations in the Raf family member B-Raf or to alterations in upstream Raf regulators, including Ras and receptor tyrosine kinases. First-generation Raf inhibitors, such as vemurafenib and dabrafenib, have yielded dramatic responses in malignant melanomas containing B-Raf mutations; however, their overall usefulness has been limited by both intrinsic and acquired drug resistance. In particular, issues related to the dimerisation of the Raf kinases can impact the efficacy of these compounds and are a primary cause of drug resistance. Here, we will review the importance of Raf dimerisation in cell signalling as well as its effects on Raf inhibitor therapy, and we will present the new strategies that are being pursued to overcome the 'Raf Dimer Dilemma'.

Figure 1

Raf dimerisation in normal and oncogenic signalling. (A, B) Ras-dependent Raf dimerisation is required for Raf kinase activation mediated by normal growth factor-induced signalling (A) and by oncogenic signalling driven by mutant RTK or Ras proteins (B). In these cases, B-Raf/C-Raf heterodimers predominate. (C) Constitutive Ras-independent self-dimerisation is required for oncogenic signalling mediated by non-V600 B-Raf mutants. (D) In contrast, V600 B-Raf mutants can signal as Ras-independent monomers. RBD=Ras-binding domain.

Figure 2

Structural features of a B-Raf dimer bound to the ATP-competitive Raf inhibitor, dabrafenib. Shown is a crystal structure of a B-Raf dimer bound to dabrafenib. The αC-helices and the conserved dimer interface RKTR motif are indicated and dabrafenib is depicted in green. The αC-helix regulatory spine residue L505 is shown in yellow and residues comprising the conserved RKTR dimer interface motif are shown in red, with R506 indicated.

Figure 3

Effect of Raf dimerisation on Raf inhibitor therapy: overcoming the Raf dimer dilemma. (A) Raf dimerisation can impact the effectiveness of first-generation Raf inhibitors by increasing the dimerisation potential of the Rafs in the presence of activated Ras, thereby promoting paradoxical ERK activation. In addition, all Raf inhibitors in the ‘DFG IN/αC OUT’ class exhibit negative cooperativity in binding to the second protomer of a Raf dimer, making these compounds ineffective inhibitors of dimeric Raf complexes. (B, C) Efforts to overcome the Raf dimer dilemma have led to the development of compounds that bind monomeric and dimeric Raf with equal potency and inhibit all Raf family members (B, pan-Raf monomer/dimer inhibitors) and drugs that alter the position of the αC-helix to such an extent as to prevent/disrupt Raf dimerisation (C, non-dimer promoting Raf inhibitors). RBD=Ras-binding domain.