Fig. 1. Colorectal cancer cells are highly susceptible to the dual inhibition of BCLxL and MCL1.

a In vitro sensitivity of a panel of colorectal cancer cell lines to the BH3-mimetic drugs. Seventeen CRC lines were treated with 0–10 μM of BH3 mimetics targeting BCLxL, MCL1, and BCL2, alone or in equimolar combinations (1:1 or 1:1:1). Cell viability was determined after 6 h. A discrete heatmap representation of the mean IC₅₀s is shown; red represents potent killing (IC₅₀ < 0.5 μM), whereas blue indicates refractoriness (IC₅₀ > 5 μM). See Supplementary Table 1 for detailed information of the cell lines used. b Killing caused by the combined action of inhibitors targeting BCLxL, MCL1, and BCL2 is mediated by BAX and BAK. The viability of parental HCT 116 cells or a BAX^−/−BAK^−/− HCT 116 sub-clone was determined 48 h after the combined inhibition of BCLxL, MCL1, and BCL2. c LoVo cells are highly refractory to apoptosis induction. Sensitivity (mean IC₅₀s ± SD) of RKO or LoVo cells treated with the indicated combinations of BH3 mimetics for 6–96 h is shown. d Expression of the MCL1-selective peptide¹⁰, BIM2A, enhances the in vivo suppression of tumor growth by the BCLxL inhibitor. The SW480 cells engineered to inducibly express BIM2A or BIM4E (inert control) were inoculated subcutaneously into immunodeficient NSG (NOD SCID IL-2Rγ^−/−) mice and treatment commenced 1 week later with 25 mg/kg of the BCLxL inhibitor A1331852 on weekdays for 2 weeks together with doxycycline-containing food to induce expression of BIM2A or BIM4E. Tumor sizes were monitored every 2–3 days and the data shown represent the mean tumor volumes ± SD of three mice in each group. e Sensitivity of patient-derived tumor organoids to the BH3 mimetics. Patient-derived tumor organoids (n = 5) were treated with 0–1.25 μM of the indicated BH3-mimetic, alone, or in equimolar combinations (1:1 or 1:1:1). Cell viability was determined 24 h later and the mean IC₅₀s of two independent experiments are shown. The relative dependency of each organoid on BCLxL, MCL1, and/or BCL2 is inferred. f Rapid induction of apoptosis by BH3-mimetic treatment. The viability of organoid #5 treated with different BH3 mimetics (see e) was determined by PI (red) staining and imaged at indicated time points (see Supplementary Video 1). Scale bar = 100 µm. Cell viability in a–c, e was determined using CellTiter-Glo assays; data in b, c represent the means ± SD from three independent experiments. See Methods in Supplementary Material for experimental details.