A, Fluorescence intensity for U2932 cells expressing the indicated sgRNAs and stained with 50 nM MitoTracker Deep Red FM dye for the 30min (n=3, mean ± SD, one-way ANOVA; (**p<0.001, ns=not-significant). B, Total NAD+ and NADH levels were determined in U2932 cells expressing the indicated sgRNAs. (n=3; mean ± SD, two-tailed Student’s t-test; **p<0.01, ****p<0.0001). C, U2932 cells expressing the indicated sgRNAs were treated for 4 days with the indicated concentrations of IACS-010759. Live cell numbers were plotted (n=3, mean ± SD). IC50 is shown in the bar graph. D, U2932 cells expressing the indicated sgRNAs were treated for 4 days with DMSO or 10 nM IACS-010759. Live cell numbers were plotted (n=3; mean ± SD,two-tailed Student’s t-test; **p<0.01, ****p<0.0001 ). E, Oxygen consumption rates (OCR) of U2932 expressing the indicated sgRNAs pretreated with DMSO or with 15 nM IACS-010759. Measurements were taken using the Seahorse XFe96 Analyzer (n=6; mean ± SD; two-tailed Student’s t-test; ****p<0.0001, ns=not-significant). F, Quantification of ATP levels in U2932 cells expressing the indicated sgRNAs. Cells were treated with DMSO or 5 nM IACS-010759 for 16h (n=3; mean ± SD; two-tailed Student’s t-test; *p<0.05, ***p<0.001, ****p<0.0001).