Fig. 3: Coordinated targeting of SRC with BRAF + EGFR increases efficacy in BRAFV600E CRC cell lines and xenografts.
a, BRAFV600E CRC cell lines treated with vemurafenib ± gefitinib were lysed and immunoblotted with the indicated antibodies. SFK activation is reflected by increased phosphorylation of the SRC activation site Y419 (pY419) and lack of phosphorylation of the inhibitory site Y530 (non-pY530). Active SRC can be deactivated by rephosphorylation of Y530 by CSK. HSP90 serves as a loading control. Molecular weight/size markers are indicated on the right (kDa). The experiment was repeated three times with similar results. b, Shift in vemurafenib sensitivity measured by cell viability assay (left) and calculation of the CI (right) upon treatment of BRAFV600E CRC or melanoma cell lines with vemurafenib + gefitinib ± a SRC inhibitor, dasatinib, for 3 d (n = 4 independent experiments per cell line). c, Colony formation assays in which BRAFV600E CRC cells were treated with an increasing concentration of vemurafenib alone (control) or with a fixed dose of gefitinib ± dasatinib. Data are representative of n = 2 independent repeats. d, Treatment of cell line-derived xenograft mouse models with a vemurafenib progenitor, PLX4720 (PLX); dasatinib; saracatinib; and/or gefitinib for 21 d (n = 7 mice per group). Plotted is the percent change in tumor volume relative to baseline (day 1). Data are displayed as the average for all mice in a specified treatment group ± standard error. e, Treatment of PDX models with vemurafenib ± gefitinib ± dasatinib for 21 d, with data plotted as in d (n = 8 mice per group). All raw and relative tumor volumes and exact P values shown in d,e are available as Source Data; P values are from a two-sided Student’s t test. f,g, GLMs testing the association of change in tumor volume between treatment arms and vehicle over time shown in d,e. Effect size is measured as the GLM standard coefficient. A GLM was applied to each tumor model separately or combined. Results for cell line xenografts and PDXs are shown in f and g, respectively. GLM P values corrected for FDR are shown in g. NT, not tested. h,i, Comparison of the effect sizes and FDR-corrected P values of treatment arms with and without a SRC inhibitor. The same number of mice per group shown in d,e was used for the analyses in f–i (that is, n = 7 mice per treatment group for WiDr and KM20 cell line xenografts and n = 8 mice per treatment group for PDX models 1 and 2).
