Fig. 4: Intrinsic VEN-resistant BCP-ALL cell lines and patient-derived xenograft samples are characterized by increased metabolic activity.
A Six BCP-ALL cell lines were incubated with increasing concentrations (DMSO; 10 nM; 100 nM; 500 nM; 1 μM; 5 μM) of VEN for 72 h before analysis of cell death by FSC/SSC criteria and normalized to the corresponding DMSO control (N = 3, in triplicates). B The basal metabolic profile of six BCP-ALL cell lines is shown. Basal oxygen consumption was measured in an XFe96 flux analyzer (Agilent) (N = 6 biological replicates/N = 1 in five technical replicates, ± standard error of mean). Uncoupled respiration was profiled by injection of 1.25 μM Oligomycin and full respiration capacity was determined by injecting 0.75 μM carbonylcyanide-p-trifluoromethoxyphenylhydrazone (FCCP). Non-mitochondrial respiration was determined by injecting 0.5 μM antimycin A and rotenone, respectively. Oxygen consumption rates (OCRs) were determined by plotting the partial oxygen pressure against time. Basal respiration was analyzed by subtracting the minimal OCR after antimycin A/rotenone injection from the basal OCR. Data were normalized to cell number by Janus Green staining. C, D Metabolic profile of two VENsens (C) compared to two VENins (D) BCP-ALL cell lines after 3 h DMSO or 10 nM and 100 nM VEN treatment is shown (N = 1 in five technical replicates ± standard error of mean). E Six PDX samples were exposed to increasing concentrations (1 nM, 5 nM, 10 nM, 50 nM, 100 nM, 250 nM, 500 nM, 1 μM, 5 μM and 10 μM) of VEN for 24 h. Relative cell death rates were assessed by propidium iodide (PI) staining and normalized to DMSO controls (N = 1, in triplicates). F The basal metabolic profile of three VENsens compared to three VENins PDX samples is shown (mean of N = 3 + 3 biological replicates, N = 1 in five technical replicates ± standard error of mean). G, H Metabolic profiles of two VENsens (G, labelled in green), and two VENins (H, labelled in red) PDX samples after 3 h exposure to DMSO or 500 nM, 1000 nM or 2500 nM VEN (N = 1 in five technical replicates ± standard error of mean).
