Dabrafenib; Preclinical Characterization, Increased Efficacy when Combined with Trametinib, while BRAF/MEK Tool Combination Reduced Skin Lesions
Figure 6
Dabrafenib-induced MAPK activation in wild-type BRAF/mutant RAS cells is CRAF-dependent and abrogated by MEK inhibition.
In order to assess the individual contributions of ARAF, BRAF, and CRAF to paradoxical MAPK activation caused by dabrafenib in HCT-116 (wild-type BRAF, mutant KRAS) cells, phospho and total MEK (pMEK, tMEK) and ERK (pERK, tERK) were evaluated by immunoblot after a 72-hour incubation with siRNA towards ARAF, BRAF, or CRAF, or medium (none), and treatment with 0, 100, or 300 nM dabrafenib for 1 h (A). Sensitivity of this paradoxical activation to MEK inhibition was evaluated in HCT-116 cells, following treatment with 0, 100, or 300 nM dabrafenib for 1 h in the presence of 0, 0.5, 5.0, or 50 nM MEK inhibitor (trametinib). Lysates were immunoblotted for total ERK (tERK) and dual-phosphorylated ERK (ppERK) (B).