Fig. 5: PARP1 inhibition boosts the in vivo and anticlonogenic effect of CHK1 inhibitors in CRC-SCs.

a Clonogenic survival of neoR-CRC-SCs left untreated or pretreated for 72 h with 100 nM prexasertib (CHK1i) alone or together with 5 µM olaparib (OLA) or 300 nM talazoparib (TZ) and then cultivated in drug-free medium as indicated. Representative images and quantitative data are reported. Results are means ± SEM and individual data points from 3 independent experiments. ^*P < 0.05, ^**P < 0.01, ^***P < 0.001 (one-way ANOVA and Bonferroni post-hoc test) compared to untreated conditions. See also Supplementary Fig. S5b. b In vivo growth of neoR-CRC-SCs (#19neoR) xenografted into immunodeficient NSG mice left untreated or treated with vehicles (Control), 5 mg/kg CHK1i, 50 mg/kg OLA or 5 mg/kg CHK1i + 50 mg/kg OLA as indicated (see Materials and Methods). NSG mice employed per group: Control- and OLA-groups: 7; CHK1i- and CHK1i + OLA-groups: 8). Tumor size curves are reported as means ± SEM. Arrows correspond to the first and last treatment, while line to the treatment period. ^*P < 0.05, ^**P < 0.01, ^***P < 0.001 (two-way ANOVA and Bonferroni post-hoc test for multiple time-points) and ^#P < 0.05, ^##P < 0.01, ^###P < 0.001 (Kruskal–Wallis ANOVA followed by Dunn’s post-hoc test for the last time point, vertical lines), as illustrated. c Western-blot analysis in CRC-SCs intrinsically resistant (innR), or moderately (SENS^MED) or highly (SENS^HIGH) sensitive to CHK1i exposed to 100 nM CHK1i for 24 h using antibodies against PARP1 and Cofilin (to ensure equal lane loading). cPARP1, cleaved PARP1. d Cell viability (evaluated by CellTiter-Glo^® assay) of innR-CRC-SCs treated with CHK1i alone or in the presence of OLA or TZ as illustrated. Data are reported as means ± SEM from 7 (on the left) and 8 (on the right) independent experiments. ^*P < 0.05, ^**P < 0.01, ^***P < 0.001 (one-way ANOVA and Bonferroni or Dunnett T3 post-hoc test) as indicated. All significant P values are shown in Supplementary Table S4. Supplementary figures associated: Supplementary Fig. S5.