(A) Synergy between AZD6244 and AZ628 was determined using the Bliss independence model in colorectal and melanoma cell lines with genetic lesions in BRAF, NF1, KRAS or NRAS (red dots indicate mutation, black indicate wild type). The sum of excess over Bliss was calculated across the matrix of concentrations tested. Values in excess of 2, indicated by the dotted red line, indicated a synergistic response.
(B) A375 cells were infected with lentiviral expression vectors encoding ORFs for LacZ and KRASG12V or shRNAs for luciferase and NF1. After 72 h, cells were treated with 1 μM AZD6244, 0.1 μM AZ628 or a combination of both compounds for 96 h. Cell proliferation was assessed using CellTiter-Glo.
(C) A375 cells were infected with lentiviral expression vectors as in (B). After 72 h, cells were treated with 1 μM AZD6244, 0.1 μM AZ628 or a combination of both compounds for 18 h. Cell lysates were analyzed for the indicated proteins.