Abstract 6210: Spatial compartmentalization of tumor and immune cellular architecture defines opposing prognostic roles in prostate cancer

Abstract Background: Understanding the role of immune cells in prostate cancer (PCa) remains challenging, with conflicting reports on immune infiltration's prognostic value. We hypothesized that spatial localization, rather than immune cells absolute bulk abundance, drives their association with clinical outcomes. To test this, we generated a clinical cohort of PCa patients integrated with multiplex IHC from spatially profiled immune cells in both the tumor and surrounding stroma (n=41). Methods: IHC multiplex assay was utilized to assess CD3+, CD8+, CD20+, FoxP3+, and CD68+ cells within cytokeratin-positive (CKpos; epithelial nests) and cytokeratin-negative (CKneg; stroma) compartments. Cell counts, densities, and ratios were correlated with time to metastasis (TTM), biochemical recurrence (BCR), BMI, tumor burden, intratumoral heterogeneity, PSA, and TMPRSS2:ERG/ETV4 fusions. After data collection validation and curation, statistical analysis of correlations between continuous variables was done using Spearman’s or Pearson’s correlation, while Chi square test was used for categorical variables. Results: Our association analysis revealed a pattern where immune infiltration within CKpos tumor nests (cancer cells) showed consistently favorable associations with clinical outcome (r=0.36-0.52, P 0.001), with CD8+ T-cells showing the strongest correlation with prolonged TTM and delayed BCR. In contrast, the same immune populations in CKneg compartment (stroma) demonstrated opposite correlations (r=-0.31 to -0.47, P 0.05), with stromal FoxP3+ T-cells most strongly associated with accelerated metastasis. Interestingly, identical immune subsets displayed opposite prognostic directions within the same patient samples, despite strong cross-compartment correlations (r=0.66-0.79, P 0.001), indicating coordinated recruitment but compartment-specific roles. Higher BMI correlated exclusively with increased stromal (non-protective) lymphocytic infiltration (r=0.39-0.59, P 0.05). Tumor burden inversely correlated with intratumoral T-cell and stromal macrophage ratios. TMPRSS2:ERG and ETV4/TMPRSS2 fusions were significantly associated with elevated stromal CD8+ infiltration, and higher pre-treatment PSA correlated with stromal CD68+ density, linking tumor biology with macrophage recruitment. Conclusion: This work demonstrates that the spatial location of immune cell populations determines their association with outcomes in PCa. Notably, our data indicate that high-burden aggressive tumors might be associated with suppression in protective intratumoral immunity. Our current work is aimed at validating these observations in a larger cohort. These findings highlight the need for a shift towards spatially-resolved tumor microenvironment and immune biomarkers for risk stratification and patient selection for immunotherapies. Citation Format: Ahmed Elsehemy, Xinpei Ci, Magnus Lam, Matthew Ramotar, Alejandro Berlin, Theodorus van der Kwast, Housheng Hansen He. Spatial compartmentalization of tumor and immune cellular architecture defines opposing prognostic roles in prostate cancer abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 6210.