Figure 4. Leveraging the landscape of sensitizers to suppress resistance.

A) Time to progression (TTP) assay in CRC240 cells treated with the ERKi (VX-11e, 1μM) + SRCi (dasatinib, 1μM) combination. Data are mean ± SEM of three replicates. B) TTP for several candidate CRC combinations tested in CRC119 cells. MEKi, AZD6244 1μM; IGFRi, GSK1838705A 1μM; SRCi, dasatinib 0.5μM; ERKi, SCH772984 0.1μM; AKTi, MK2206 5μM. C) Immunoblots of indicated targets in CRC119 cells treated with DMSO, MEKi (AZD6244, 1μM), SRCi (dasatinib, 0.5μM), or the combination for 14 days and probed at the indicated times. Blots are cropped for clarity. D) Pairwise combinations of sensitizers in CRC119 cells to identify triple combinations (left). Triple combinations (red) were tested for their ability to shift the GI50 curves to a greater extent than either of the two body combinations (grey; center). Log2 fold shifts from baseline are shown for all combinations tested, where negative values indicate leftward shift of the curve as in the center plot. Data are mean ± SD of three replicate experiments. Drug identities as above except: RAFi, LY3009120 0.05μM; ERKi 0.05μM; SRCi 1μM. E) CRC119 annexin V+ cells after 48 hours of treatment with the indicated combinations (MEKi, SRCi, and AKTi identity same as above). F) TTP assay for a candidate triple combination (MEKi AZD6244, 0.2μM; SRCi dasatinib, 1μM; AKT MK2206, 10μM). G) HCT116 xenograft treated with vehicle, MK-2206 (15 mg/kg, daily), dasatinib (15 mg/kg, daily) and AZD6244 (15 mg/kg, daily), or the triple combination. For average tumor volumes, each arm only plots the data up to the point at which the first mouse in the group reached the humane endpoint. To the right, a survival curve showing percent of mice with tumors less than 4X the starting volume at a given time. To the right of survival curve are the mouse weights for the triple combination group over the course of the study. Data are mean ± SEM. *p>0.05. See also Figure S4.