Extended Data Fig. 10: Functional rescue experiments and in vivo validation of WRN dependency in a MSI colorectal cancer cell line.
From: Prioritization of cancer therapeutic targets using CRISPR–Cas9 screens
a, Expression of wild-type mouse Wrn rescued the viability effect of WRN knockout in MSI cell line SW48. MSS cell line SW620 was used a negative control. Box-and-whisker plots represent the median and 1.5× interquartile range. Data represent two independent biological replicates completed in technical triplicate. b, Western blots confirmed expression of Flag-tagged protein using all variants of the Wrn vector. Images are representative of experiments performed in triplicate. c, WRN knockout induced by doxycycline treatment in WRN sgRNA-expressing HCT116 (HCT116-WRN) cells measured by western blot for two separate clonal lines. Data are representative of two independent experiments. d, Growth curves of HCT116 parental, HCT116 sgNon (non-essential sgRNA) and WRN sgRNA-expressing HCT116 cells grown in the absence (black line) or presence of doxycycline (2 μg ml−1; yellow line). Data are mean ± s.d. of 10 technical replicate wells for each condition (1 image per well) and representative of two independent experiments. e, Growth curves of WRN sgRNA-expressing HCT116 (clone b) subcutaneous tumours from mice treated with doxycycline (50 mg kg−1; yellow line) or vehicle (grey line). Tumour growth suppression was observed (P = 0.03, two-way ANOVA comparing doxycycline versus vehicle). The number of mice in each cohort is indicated. Data are mean ± s.e.m. f, Representative KI-67 immunohistochemistry assessment of WRN sgRNA-expressing HCT116 (clone b) tumours explanted after one week of doxycycline treatment (left). Scale bar, 50 μm; 40× magnification. Quantification of KI-67 staining (right). Data are mean ± s.d. of 10 fields from three different samples (n = 30) and means were compared using a two-sided Welch’s t-test. Source data for all western blots are shown in Supplementary Fig. 1.
