Fig. 6: RORγ inhibition in CRISPR/Cas9 t(4;11) cells acts synergistically in combination with ATV.

CRISPR/Cas9 t(4;11) cells were simultaneously treated with increasing concentrations of XY018 alone or in combination with FS (A) or ATV (B) for 7 days. The percentage of living cells (annexin V−, PI−) was evaluated with flow cytometry. IC₅₀ values were mapped as isobologram and the Chou-Talalay method was used to measure the CI for identification of synergistic, additive or antagonistic effects. C Apoptosis was measured in CRISPR/Cas9 t(4;11) cells treated with 15 µM XY018 alone or in combination with 15 µM FS or ATV. DMSO-treated or single-treated cells were used as control, respectively. Representative dot plots (top) and pooled data (bottom) show different apoptotic stages of three independent donors (n = 3) measured in technical duplicates. Bars represent the mean ± SD. One-way ANOVA. *p < 0.05. D RT-qPCR analysis of SREBF2 gene expression in the CRISPR/Cas9 t(4;11) cells treated with 15 µM XY018, 15 µM FS or 15 µM ATV alone or in combination for 72 h. Bars represent the mean of three independent donors (n = 3) ± SD. One-way ANOVA. *p < 0.05. E Heat map display of fold changes (in log2) in genes of cholesterol homeostasis in CRISPR/Cas9 t(4;11) cells treated as described in (D). The expression of indicated genes was measured by RT-qPCR for which DMSO-treated cells were set as 1. Experiments were performed with three independent donors (n = 3) in technical duplicates. One-way ANOVA. *p < 0.05. CRISPR/Cas9 t(4;11) and CD34+ control cells were treated with indicated concentrations of XY018 alone or in combination with FS (F) and ATV (G) for a total of 48 h. The percentage of living cells (annexin V−, PI−) was evaluated with flow cytometry, and vehicle-treated cells were set to 100 as an internal control. Bars represent the mean of three independent donors (n = 3) ± SD. One-way ANOVA. *p < 0.05. ns=not significant.