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. Author manuscript; available in PMC: 2018 Jul 25.
Published in final edited form as: Cell Rep. 2017 Jul 25;20(4):999–1015. doi: 10.1016/j.celrep.2017.07.006

Figure 2. Landscape view and validation of sensitizers to MEK/ERK inhibitors across KRAS mutant cancers.

Figure 2

A) Comparison of hit frequency across tissues and drugs. A gene is considered a hit if it scores reproducibly in two cell lines per tissue. B) Hierarchical clustering of the Z-scored DM for the 3 most active sgRNAs per gene in each replicate for GDC-0623 (MEKi) and SCH772984 (ERKi) screens. For each condition, cells were grown either in vehicle or low doses of the indicated inhibitor (see Supplemental Table 3 for doses) for 3–4 weeks and then results were de-convoluted by deep sequencing (boxes highlight representative areas of heat, indicating groups of possible tissue-specific sensitizers). C) Table with representative processes and corresponding target genes that modulate sensitivity to MEK/ERK inhibition uncovered by the screens. D–F) Crystal violet staining of 7 day colony growth in cell lines treated with the indicated, candidate sensitizers. Cells were treated with the indicated inhibitors in combination with AZD6244 (MEKi) or ERKi (SCH772984) at the listed concentrations (ERK5i, XMD8-92; MDM2/4i, MI-773; EGFRi, Gefitinib; mTORC 1/2, Torin1; SRCi, dasatinib; CDK1i RO-3306). Data are a representative image of each experiment performed in duplicate. (G–I, top) Pharmacologic validation of 12 sensitizers. Mutant and wild type cells were tested in 8-point GI50 assays with either SCH772984 (ERKi) alone or in the presence of a constant background concentration of the indicated drugs. Relative viability was measured at 72 hours post-treatment using Cell Titer Glo. Dotted line indicates ERKi GI50 value for DMSO treated KRAS mutant cells. Data are mean ± SEM of three replicate experiments. (G–I, bottom) Similar to top, log2 transformed GI50 values for two KRAS mutant cell lines and one KRAS WT cell line per tissue. Data are normalized to DMSO treated samples for each cell line. (DNMT1i, Azacitadine 0.5μM; EZH2i UNC1999, 0.5μM; CDK2i Roscovitine, 5μM; CDK9i LDC000067, 2μM; CDK7i BS-181, 2μM; SRCi dasatinib, 0.2μM; IGFRi, GSK1838705A 1μM; mTORC1 Rapamycin, 0.1μM; mTORC 1/2, Torin1, 0.2μM; RAFi LY3009120, 1μM; CDK1i RO-336 5μM; CDK4/6i PD0332991, 2μM). *p<0.05. See also Figure S1, S2 and Tables S3, S4.