Abstract PS3-13-22: Epithelial Podoplanin Co-expression in Postpartum Breast Cancer: An Exploratory TME Analysis

Abstract Background: Postpartum breast cancer (PPBC) - diagnosis within 10 years postpartum - shows more aggressive and ∼3-fold higher metastasis risk than nulliparous or ≥10 years postpartum cases. Human data on the PPBC tumor microenvironment (TME) are limited. We evaluated epithelial, macrophage, and endothelial wound-healing phenotypes in the PPBC TME. Objective: Identify TME phenotype (s) associated with PPBC status. Study Population: From the international Breast Cancer Family Registry (BCFR), we included 180 invasive ductal carcinoma cases diagnosed ≤50 years within the New York subset who had a H 31, 40, and 58 cases diagnosed 5, ≥5 - 10, ≥10 years postpartum, respectively, and 51 nulliparous cases. Methods: Multiplex immunofluorescence panel design, staining, and multispectral imaging (Opal/Vectra Polaris) were conducted blinded to patient clinicopathologic data for Podoplanin (PDPN), Macrophages (CD68), Endothelial Cells (CD31/CD34 antibody mixture), panCK, SEMA7A, PD-L1, and DAPI. Slides were scanned with a 10X objective. Tumor containing regions were annotated by pathologists; cell phenotyping, cell counts, and signal intensities were quantified in HALO (full magnification), with fluorescent intensities normalized to 0-255. Phenotypes were defined by co-/anti-coincidence logic. Within tumor regions, we computed densities (cells/mm2) for markers and 20 colocalized phenotypes, normalized each by DAPI+ nuclear density to yield unitless ratios, and log-transformed these ratios to approximate normality. Phenotypes with ≥25% zero values were dichotomized as present/absent. Continuous predictors were z-scored within cross validation (CV) folds. We defined PPBC cases as 10 years postpartum; nulliparous or ≥10 years postpartum were considered non-PPBC cases. Statistical Analysis: We examined clinicopathologic, epidemiologic, and the tumor phenotype predictor variables stratified by PPBC status (chi2, t-tests). We then fit a LASSO-penalized logistic regression; λ was tuned by repeated 5-fold CV with 5 repeats, optimizing for AUC (λₘin). Continuous predictors were standardized (z-scored) within CV folds to prevent information leakage. Binary indicators for categorical predictors remained 0/1 (not standardized). Predictors with non-zero coefficients at the selected λ comprised the active set. For interpretability, we refit an unpenalized logistic regression model on the active set (‘post-LASSO’) and reported odds ratios (OR) and 95% CIs per 1-SD increase. Sensitivity analyses (i) restricted the sample to parous participants and including parity-related predictors (e. g. , breastfeeding), and (ii) redefined PPBC cases to 5 years postpartum in the post-LASSO model. Results: PPBC and non-PPBC cases were similar across predictors, except PPBC cases were younger at diagnosis (p0. 001), and among parous women, PPBC cases were older at first FTP prior to diagnosis (p0. 001). At λₘin, only age at diagnosis and PanCK+PDPN+ were retained. Macrophage- and endothelial/lymphatic-centric phenotypes shrank to zero. In post-LASSO models, age at diagnosis was inversely associated with PPBC (ORper 1-SD 0. 39, 95% CI (0. 26, 0. 58) and PanCK+PDPN+ was positively associated (ORper 1-SD 1. 72, 95% CI (1. 21, 2. 52) ). When restricted to the parous subsample, PanCK+PDPN+ was retained by LASSO (post-Lasso ORper 1-SD 2. 04, 95% CI (1. 11, 4. 16) ) ; however, when redefining PPBC cases to 5 years postpartum, PanCK+PDPN+ was null (post-Lasso ORper 1-SD 0. 93, 95% CI (0. 47, 2. 04) ). Conclusion: PPBC tumors showed enrichment of PanCK+PDPN+ epithelial cells, consistent with epithelial PDPN upregulation seen in wound-healing-like programs. PanCK+PDPN+ was retained among parous women, but not within a more conservative definition of PPBC. Findings are exploratory and warrant validation. Citation Format: J. A. McDonald, A. J. Kociolek, S. Tun Hein Aung, Y. Liao, B. Vasikj Bjelogrlic, C. Loomis, J. M. Genkinger, M. Terry, K. L. Gardner. Epithelial Podoplanin Co-expression in Postpartum Breast Cancer: An Exploratory TME Analysis abstract. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32 (4 Suppl): Abstract nr PS3-13-22.