Skip to main content
Fig. 3 | Genome Medicine

Fig. 3

From: A role for the unfolded protein response stress sensor ERN1 in regulating the response to MEK inhibitors in KRAS mutant colon cancers

Fig. 3

A genetic screen for resistance to MEK inhibitors in ERN1 knockout colon cancer. a Schematic outline of the genome-scale CRISPR/Cas9 knockout screen for resistance to MEK inhibition. Two different MEK inhibitors, selumetinib and trametinib, were used, each in two replicates, and compared to the untreated control population. b, c MA plots of the selumetinib (b) and trametinib screens (c). Horizontal dashed line indicates an arbitrarily imposed threshold of log2 (fold change of treated over untreated) of 7 and vertical dashed line indicates mean number of reads in untreated samples of 50. Highlighted in color are the sgRNAs targeting DUSP4, DET1, COP1, CBFB, RUNX2, and STK40, that are found above these two thresholds (with the p adjusted of ≤ 0.1) in both the selumetinib (b) and the trametinib (c) screen. d, e Functional validation of DET1 and COP1 in LoVo ERN1KO background. d Colony formation assays of DET1 and COP1 KO cells in the presence and absence of the MEK inhibitor AZD6244 (selumetinib) are shown relative to control cells having NT gRNA. Shown is a representative example of at least three biological replicates. e Live cell proliferation assay of DET1 and COP1 KO cells in the presence and absence of 1 μM AZD6244 compared to control cells expressing NT gRNA. Error bars indicate standard deviation calculated from three replicate experiments. f Western blot analysis of DET1 and COP1 expression in DET1 and COP1 knockout cells using antibodies against ERN1, DET1, COP1, JUN, p-ERK, and HSP90 as control both in the presence and absence of the MEK inhibitor AZD6244. g Median-centered log(IC50) of five different MEK1 inhibitors in high (top 25%) and low (bottom 25%) expressing DET1 (left) and COP1 (right) CRC cell lines in the GDSC100 data set [42]. Cell lines with high DET1 or COP1 expression have significantly lower IC50s (p = 0.004 for both DET1 and COP1). Log(IC50) estimates were median-centered over all cell lines to make them comparable between MEK inhibitors

Back to article page