Figure 1.

Differential effects of the RAF inhibitor PLX4032 in BRAF mutant melanoma, thyroid and colorectal cancer cell lines. A, bars represent IC₅₀ values for PLX4032 in 15 BRAF (+) cell lines. The majority of thyroid and colorectal cell lines were comparatively refractory to PLX4032, whereas melanoma lines were uniformly sensitive. A set of BRAF wild type thyroid cancer cell lines were resistant to growth inhibition by the compound. B, western blots of cell lines from (A) treated with 2 μM PLX4032 immunoblotted for pMEK1/2 (Ser217/221) and pERK1/2 (Thr202/Tyr204). Melanoma cell lines had a sustained inhibition of pERK1/2 after exposure to the compound, whereas most thyroid and colorectal cancer cell lines had a rebound of pERK1/2 as early as 6 h post-treatment. As expected, BRAF wild type thyroid cancer cell lines were resistant to MAPK pathway inhibition by PLX4032. C, SW1736 and 8505C were treated as in (B) and collected at the indicated times. 72 h post-treatment cells were re-treated with fresh PLX4032 for 1 h (lane 72 + 1). Immunoblots of pMEK1/2 and pERK1/2 demonstrated that re-addition of PLX4032 did not fully re-inhibit MAPK signaling. D, SW1736 and SK-MEL-28 cells were treated as in (B) and collected at 0, 24 and 48 h post-treatment. Lysates were immunoprecipitated with the RAS binding domain of CRAF (RBD) and immunoblotted for pan-RAS. Activation of RAS increased at 24 and 48 h of treatment in SW1736 cells.