Fig. 1: Chronic activation within the tumor microenvironment of BCG-unresponsive tumors promotes adaptive resistance.
A, Targeted mRNA gene-set enrichment analysis showing statistically significant differences between BCG-naïve (N=20) and BCG-unresponsive cases (N=20). All gene sets are significant at p<0.05 by Kruskal-Wallis or independent two-sided t-test.
B, Longitudinal protein analysis of urine supernatants across four timepoints in BCG-naïve patients with NMIBC (N=27) with comparisons between BCG-naïve and 6th induction dose timepoints (* p<0.05, ** p<0.001).
C, An additional validation cohort of patients with NMIBC receiving BCG therapy at Aarhus University (N=66) showing BCG-naïve vs BCG-exposed timepoints in urine samples (p-values in both cohorts assessed via independent two-sided t-test or Kruskal-Wallis with Benjamini-Hochberg correction for multiple comparisons).
D, Differential protein expression analysis of urine before (top row) and after (bottom row) BCG exposure and sampled before (left-side) and after (right-side) transurethral resection of bladder tumors. Number of patients included in analyses: (Top-left) BCG-naïve when tumor present, non-responders, N= 47; responders, N= 60. (Top-right) BCG-naïve after tumor resection, non-responders, N= 5; responders, N= 6. (Bottom-left) BCG-exposed when tumor present, non-responders, N= 49; responders, N= 34. (Bottom-right) BCG-exposed after tumor-resection, non-responders, N= 19; responders, N= 26. P-values assessed with two-sided Wilcoxon rank sum. Proteins listed in orange text indicate unadjusted p-values <0.05. BCG-unresponsive NMIBC was defined as high-grade recurrence within two years after receiving BCG therapy.
E, Median fluorescence intensity (MFI) for PD-L1 and HLA-E staining on primary CD45- cells from patients with NMIBC and immortalized bladder tumor lines ex vivo or after 24 hours of incubation with recombinant human (rh) IFN-γ stimulation (all comparisons, p<0.05). All stimulation experiments were performed in triplicate.