Fig. 2

AZD1775 has antileukemic effects against primary and patient-derived ALL samples in vitro, independent of p53 status. A Sensitivity of primary (N = 4) and patient-derived xenograft (N = 23) ALL samples to AZD1775 exposure for 96 h in ex vivo co-culture with hTERT-immortalized MSCs. Drug responses were determined by fluorescence image-based microscopy in at least technical duplicate, relative to respective DMSO controls (N = 1 for each respective sample). IC50 values are based on live cell enumeration fitted with a 4-parameter non-linear regression curve. MSC drug sensitivity is based on at least three independent experiments. B Evaluation of apoptosis by Annexin-V staining with flow cytometry in PDX samples (N = 8) following exposure to respective AZD1775 IC50 fractions for 48 h (N = 1 for each respective sample). Error bars show mean ± SD C Immunoblot of apoptotic marker cleaved PARP (Asp214) in PDX samples (N = 3) treated with respective AZD1775 IC50 fractions for 24 h. ɑ-tubulin was used as loading control. D Immunoblot of cell cycle markers in PDX samples (N = 3) treated with respective AZD1775 IC50 fractions for 24 h. ɑ-tubulin was used as loading control. E No correlation was observed between AZD1775 IC50 values and WEE1 protein expression levels determined by semi-quantitative immunoblotting (n = 10). The non-parametric one-tailed Spearman test was used to determine the correlation coefficient. F Statistical significance between adavosertib IC50s for cell lines, primary, and primary-derived ALL blast samples with (N = 27) or without (N = 5) TP53 mutations were compared using Mann–Whitney U test. Dots in all panels represent individual samples