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Fig. 3 | Journal of Experimental & Clinical Cancer Research

Fig. 3

From: Metabolic synthetic lethality by targeting NOP56 and mTOR in KRAS-mutant lung cancer

Fig. 3

IRE1α-mediated UPR fuels mTOR signaling via p38 MAPK. A, H358 cells expressing scrambled control or NOP56-specific shRNAs were treated with different inhibitors, with bar graphs illustrating sensitivity increase after NOP56 KD. Fold changes of IC50 values were presented as IC50 of rapamycin in H358 cells expressing scrambled shRNA (sh_Scrambled) compared to that in H358 cells expressing NOP56-targeted shRNAs (shNOP56 a/b). Data presented as mean (n = 2). B, Clonogenic assay of H358 and H460 cells expressing control shRNA (sh_Scr) or NOP56-specific shRNAs (sh-a, sh-b) after treatment with indicated doses of rapamycin or everolimus. Representative images are shown. The heatmap (right) indicates the percentage of viable cells after the treatment, based on quantification of clonogenic results (left). Data are presented as mean ± SD (n = 3). C, Growth inhibition of H358 and H460 cells expressing NOP56-specific shRNAs (sh NOP56a, sh NOP56b) or control shRNAs (sh Scram) after treated for 72 h with the mTOR inhibitors (rapamycin, everolimus). Data are presented as mean ± SD (n = 3). D, NOP56 silencing significantly enriched the mTOR gene signature in KRAS-mutant cancer cells. GSEA was performed based on the GEO dataset GSE15212. E, Negative correlation of NOP56 mRNA levels with mTOR gene signature (mTOR pathway score) as determined in a TCGA cohort of patients with KRAS-mutant lung adenocarcinoma. Pearson and Spearman coefficient, as well as the significance (p-value), were determined using R software (Cor.test function). F, NOP56 expression is a predictive marker of sensitivity (IC50) to rapamycin in KRAS-mutant cancer cell lines (n = 18) but not in KRAS-wide-type cancer cell lines (n = 82). Drug response profiles were downloaded from the GDSC (Genomics of Drug Sensitivity in Cancer) database. G, Immunoblots of H358 cells after transfection with scramble control siRNAs (si-Control) for 72 h (−) or NOP56-specific siRNAs (si-NOP56) for different time points (24 h, 48 h, 72 h and 96 h). H, Immunoblots of H358 and H460 cells expressing scrambled control or NOP56-specific shRNAs. I, Immunoblots of H358 cells expressing scramble control or the NOP56-specific shRNAs after treated with the p38 inhibitor SB203580 (5 μM) for 24 h. J, Clonogenic assay of H460 and H358 cells expressing control or NOP56-specific shRNAs after treated with the p38 inhibitor SB203580. Representative images are shown

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