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. 2017 Mar 23;2(6):e90196. doi: 10.1172/jci.insight.90196

Figure 4. Panobinostat synergizes with ibrutinib in killing ABC DLBCL harboring MyD88 mutation.

Figure 4

(A) MTS assay confirming enhanced antiproliferative effects of panobinostat in combination with ibrutinib in ABC cells harboring MyD88 mutation (HBL-1, TMD-8, and OCI-LY-10) but not in ABC cells with MyD88 WT (Ri-1 and U-2932). Cells were incubated with increasing concentrations of panobinostat (0.01, 0.025, 0.05, 0.075 μM) and ibrutinib (0.01, 0.025, 0.05, 0.075 μM), and viability was assessed after 24 hours. Error bars represent SEM of 3 independent experiments. Differences between groups were calculated with 2-way ANOVA with Bonferroni’s test. ***P < 0.0005; ****P < 0.0001. (B) Scatterplots summarizing the results of panobinostat + ibrutinib combinations in our cell line panel after 24 hours of incubation. Cells were incubated with increasing concentrations of panobinostat (0.01, 0.025, 0.05, and 0.075 μM) and ibrutinib (0.01, 0.025, 0.05, and 0.075 μM), and combination index was calculated according to the Chou-Talalay method. All the experiments were repeated 3 times. (C) MTS assay showing enhanced antiproliferative effects of panobinostat in combination with ibrutinib in Ri-1 stably expressing MyD88 mutation but not in Ri-1 with MyD88 WT. Cells were incubated with increasing concentrations of panobinostat (0.01, 0.025, 0.05, and 0.075 μM) and ibrutinib (0.01, 0.025, 0.05, and 0.075 μM), and viability was assessed after 24 hours. Error bars represent SEM of 3 independent experiments. Differences between groups were calculated with 2-way ANOVA with Bonferroni’s test. ***P < 0.0005; ****P < 0.0001. (D) Relative NF-κB-luciferase activity in 2 representative ABC (HBL-1 and TMD-8) cell lines. Cells were treated for 16 hours with indicated concentration of either panobinostat, ibrutinib, or the combination of the 2 drugs. Cells were incubated with 2 ng/ml of TNF-α as positive control for NF-κB activation. Error bars represent SEM of triplicates. Differences between groups were calculated with 2-way ANOVA with Bonferroni’s test. **P < 0.005; ***P < 0.0005; ****P < 0.0001. (E) IL-10 level change in 3 ABC DLBCL cell lines harboring MyD88 mutation (HBL-1, TMD-8, and OCI-LY-10). Cells were treated for 12 hours with indicated concentration of either panobinostat, ibrutinib, or the combination of the 2 drugs. Error bars represent SEM of triplicates. Differences between groups were calculated with 2-way ANOVA with Bonferroni’s test. **P < 0.005; ****P < 0.0001.