Figure 8. Synergistic effect of BIRD-2 and venetoclax in SU-DHL-4 cells, correlating to Bim upregulation in BIRD-2-treated SU-DHL-4 cells.

(A) SU-DHL-4 cells were treated for 24h with various concentrations of venetoclax alone (black bars), BIRD-2 alone (blue bars) or a combination of venetoclax/DMSO Ctrl (0.03%) and BIRD-2 (red bars). Cell death was measured using flow cytometry of Annexin V-FITC/7-AAD-stained cells and plotted as the BIRD-2- or venetoclax-induced apoptotic fraction. The combination index, calculated using the response additive method (CI= (E_venetoclax+ E_BIRD-2)/E_{venetoclax+BIRD-2}) was measured as indication for synergistic or additive cell killing. Data are represented as average ± SEM of N=5. Statistical significance was determined with a two-way ANOVA with a Bonferroni post-hoc test comparing E_venetoclax or E_BIRD-2 with E_{venetoclax+BIRD-2} for the different venetoclax concentrations. (B) Representative western blots showing the Bim_EL-expression levels after 24h treatment of SU-DHL-4 cells with 3 μM BIRD-2 in the presence or absence of 10 μM BAPTA-AM. Vinculin was used as loading control. Data are represented as the average ± SD of N≥3 with ^* p<0.05 obtained via Wilcoxon signed rank test.