Fig. 4. ATM loss is sufficient to drive immune changes and improve anti-PD1 response in preclinical bladder models but ATM mutations are not associated with immune properties or anti-PD1 response in clinical bladder cohorts.

(A) Growth curves for ATM-deleted and WT ATM tumor xenografts. The ATM-deleted tumors had a larger response to anti-PD1 treatment than WT ATM xenografts (linear regression model). (B) Flow cytometry showed no differences in the number of CD8⁺ T cells in ATM-deleted versus WT ATM tumors. Data are plotted as the means ± SD, n = 5 tumors. (C) Flow cytometry showed a significantly higher number of CD4⁺ T cells in ATM-deleted compared to WT ATM tumors, **P < 0.01. (D) Flow cytometry showed significantly fewer FOXP3⁺ CD4⁺ T cells in ATM-deleted compared to WT ATM tumors, *P < 0.05. (E) Principal components analysis (PCA) of RNA sequencing (RNA-seq) counts from ATM-deleted and WT ATM tumors. (F) RNA-seq–based immune cell fraction estimation using TIMER revealed a trend toward increased CD4⁺ T cells in ATM-deleted compared to WT ATM tumors (Student’s t test; BH correction). (G) A gene expression signature of transforming growth factor–β signaling in fibroblasts was higher in WT ATM tumors than in ATM-deleted tumors. (H) RNA-seq analysis of immune cell subsets in ATM-mutant versus ATM nonmutant tumors from the TCGA (left) and IMvigor210 (right) bladder cancer cohorts. There were no significant differences in any immune cell subsets (Wilcoxon rank sum test). (I) Quantification of multiplexed immunofluorescence data from institutional bladder cancer cases showed no difference in CD8, PD1, or FOXP3 staining between ATM-mutant and nonmutant bladder tumors. (J) OS of patients with ATM-mutant and nonmutant bladder tumors in the Imvigor210 cohort (Kaplan-Meier method). (K) TTF (left) and OS (right) for ATM-mutant and ATM nonmutant institutional bladder cancer cases following anti-PD1/PD-L1 treatment (Kaplan-Meier method).