The MAPK signaling pathway, comprised of H/K/N-RAS, A/B/C-RAF, MEK1/2, and ERK1/2, plays a major role in the regulation of cellular functions such as cell-cycle regulation, proliferation, survival, and migration. The pathway is activated by extracellular signals that in turn induces the small G protein RAS to exchange GDP for GTP, and results in activation of the RAF/MEK/ERK cascade. The MAPK pathway is activated in many human cancers such as those harboring mutations in RAS or RAF. RAF709 is a novel ATP-competitive kinase inhibitor with high potency and selectivity against RAF kinases. In in vitro biochemical assays, RAF709 exhibited potent inhibitory activity targeting BRAF, BRAFV600E, and CRAF with IC50 values 1.5 nM, 1 nM and 0.4 nM, respectively. The cellular selectivity of RAF709 was evaluated in a Ba/F3 cell panel using wild-type or Ba/F3 cells rendered IL3 independent by stably expressing 38 different kinase oncogenes. Consistent with on-target activity against RAF kinases, RAF709 was active in Ba/F3 cells expressing the BRAFV600E oncogene with an IC50 of 0.52 µM with little activity observed in cells expressing 36 additional kinases, these data demonstrated that RAF709 is an active and highly selective inhibitor targeting the RAF kinases. In A375 (BRAFV600E) cells, RAF709 showed robust activity inhibiting mutant BRAF monomer-driven ERK activation with IC50 of 44 nM. And in HCT116 cells (KRASG13D), RAF709 pretreatment exhibited activity inhibiting RAF dimer-driven signaling with an IC50 of 79 nM for pERK. These data suggested that RAF709 inhibits both RAF monomers and dimers with similar potency. Nude mice bearing Calu-6 xenograft tumors were treated with a single dose of RAF709 across a wide dose range (from 10 to 200 mg/kg). The result showed that RAF709 at 100 mg/kg and 200 mg/kg was able to suppress pMEK to greater than 50% for more than 16h, which demonstrated that RAF709 has antitumor activity in tumors harboring RAS mutations in vivo[2].
作用机制
RAF709 binds to the BRAF kinase in a characteristic of a type II inhibitor binding
mode. RAF709 in complex with the BRAF kinase domain revealed that the protein adopts an inactive conformation with the DFG out and the aC-helix in.
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