Deshmukh, S. Kumar

Abstract Background: Tertiary lymphoid structures (TLS), which are organized ectopic lymphoid aggregates resembling secondary lymphoid organs, have been associated with improved outcomes across various cancer types. However, limited data exist on the prognostic relevance of TLS in HER2+ BC pts treated with trastuzumab. In our prior work, we demonstrated that the presence of functional TLS, defined by lymphoid aggregates with elevated expression of BCL6 or IL21R, correlated with improved outcomes in the NCCTG N9831 trial. In this study, we evaluated TLS-related gene and outcomes in the real-world database. Methods: 1344 HER2+ (HR-HER2+: 560; HR+HER2+: 784) BC samples were analyzed via NGS (592-gene panel, NextSeq; WES/WTS, NovaSeq; Caris Life Sciences). Above (high) or below (low) the 50th percentile expression levels of TLS-related genes were used to calculate overall survival (OS). IL21R-high (H) and -low (L) tumors were classified by RNA expression above or below the 50th percentile. Real-world OS was derived from insurance claims and calculated from date of biopsy to last contact using Kaplan-Meier. Statistical significance was assessed using chi-square and Mann-Whitney U with multiple comparison adjustments (q < 0.05). Results: Among pts with HR-HER2+ BC, high expression of IL21R, CXCL9, CD38, and CXCL10 were associated with improved OS (Table). In contrast, among pts with HR+HER2+ BC, high expression of CXCR3, PDCD1, TBX21, CD200 and IL21R, were associated with improved OS. In both HR-HER2+ and HR+HER2+, IL21R-H groups had 1) higher infiltration of B cells (HR-HER2+: 4.4% vs 2.9%; HR+HER2+: 5.6% vs 4.2%), M1 MΦ (HR-HER2+: 3.9% vs 2.6%; HR+HER2+: 3% vs 2.1%), M2 MΦ (HR-HER2+: 4% vs 3.2%; HR+HER2+: 5.7% vs 4.4%), NK cells (HR-HER2+: 2.8% vs 2.5%; HR+HER2+: 2.7% vs 2.4%), CD8+ T cell (HR-HER2+: 0.9% vs 0%; HR+HER2+: 0.7% vs 0%), Tregs (HR-HER2+: 2.9% vs 1.4%; HR+HER2+: 2.8% vs 1.5%), all q<.05; 2) higher T cell inflamed score (HR-HER2+: 87.5 vs -95; HR+HER2+: 71 vs -86), IFNy score (HR-HER2+: -0.19 vs -0.48; HR+HER2+: -0.27 vs -0.49), all q<.05; 3) higher expression levels of MHC class I genes (HLA-A, HLA-B, HLA-C, HLA-E, FC: 1.6-2.3; all q<.05), MHC class II genes (HLA-DQA1, HLA-DOA, HLA-DPA1, HLA-DMA, FC: 2.2-3.2; all q<.05) compared to IL21R-L group. Conclusion: Consistent with our prior findings, overexpression of TLS-related genes was associated with improved outcomes in HER2+ BC in real-world data. Notably, the prognostic impact of TLS-related genes differed by hormone receptor (HR) status. Among these, IL21R emerged as the only gene significantly associated with clinical outcomes across both datasets and HR subgroups. Higher IL21R expression is associated with more B cell, M1 MΦ, CD8 T cell and NK cells. These findings underscore the potential importance of the IL21/IL21R axis in HER2+ BC and warrant further investigation to validate its role and therapeutic relevance. Citation Format: S. Chumsri, S. Deshmukh, Y. Liu, S. Wu, J. Xiu, N. Norton, S. Shipra Gandhi, E. Torres, M. Lustberg, E. Perez, A. Nassar, G. Sledge, E. Thompson, K. Knutson. Tertiary lymphoid structure (TLS)-related gene expression and outcomes in HER2-Positive (HER2+) breast cancer (BC) in the Real-World Database [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 PS2-09-08.

Abstract Background: Triple-negative breast cancer (TNBC) is an aggressive yet immunogenic subtype of breast cancer (BC) and the only one with FDA-approved immune checkpoint inhibitor (ICI) - pembrolizumab. Currently, PD-L1 (22C3) is the companion biomarker for predicting pembrolizumab benefit, but its utility is limited by spatial/temporal heterogeneity, assay variability, and modest correlation with treatment response. Chemokines such as CXCL9, CXCL10, and CCL5 recruit CD8+ T cells into the tumor microenvironment and have been associated with improved response to ICI, but their predictive value alongside PD-L1 remains unclear. Here, we examined the relationship between these chemokines, PD-L1, and pembrolizumab efficacy in TNBC. Methods: We analyzed 3662 TNBC tumors profiled by NGS (592, NextSeq; WES/WTS, NovaSeq; Caris Life Sciences). Immune cells were estimated using WTS deconvolution (Quantiseq). PD-L1 positivity was defined as >10% CPS (PharmDx 22C3). CXCL9/CXCL10/CCL5-high (H) and -low (L) TNBC were classified by RNA expression above or below 50th percentile. Real-world median overall survival (mOS) was derived from insurance claims and calculated from start of pembrolizumab to last contact using Kaplan-Meier. Statistical significance was assessed by chi-square and Mann-Whitney U with multiple comparison adjustments (q<0.05). Results: TNBC expressed higher levels of CXCL9, CXCL10, and CCL5 (N = 3662, median TPM: 5.3, 14.8, and 15.1) compared to HER2+ (N = 1344; 4.8, 10.1, 10.6) and HR+HER2- BC (N = 6097; 2.7, 6.6, 9.4), all q<0.05. Moderate correlations were observed between PD-L1 CPS and each chemokine (r = 0.41, p<.01). PD-L1-positive tumors had better mOS (24.3 months (m) vs 18.6 m, HR 0.75, 95% CI 0.6-0.92, p=0.007) compared to PD-L1-negative TNBC post-pembrolizumab. Among PD-L1-negative tumors, those with CXCL9-H had better mOS (26.9 m vs 15.8 m for CXCL9-L, HR 0.61, 95% CI 0.45-0.82, p=0.001) post-pembrolizumab. Among PD-L1-positive tumors, CXCL10-H and CCL5-H was associated with improved mOS (CXCL10-H: 25.7 m vs. 20.7 m for CXC10-L, p = 0.03; CCL5-H: 26.7 vs. 18.5 m for CCL5-L, p = 0.03) post-pembrolizumab. PD-L1 negative CXCL9-H, PD-L1 positive CXCL10-H and PD-L1 positive CCL5-H groups showed higher infiltration of CD8+ T cell, B cells, M1/M2 macrophages, Tregs. They also exhibited higher T cell inflamed score, IFNy score and MAPK activation score vs. CXCL9-L, CXCL10-L and CCL5-L groups (Table). Conclusions: In TNBC, a chemokine signature comprising CXCL9, CXCL10 and CCL5 refines PD-L1 based prediction of pembrolizumab benefit. Specifically, high CXCL10 or CCL5 expression in PD-L1-positive tumors, and high CXCL9 expression in PD-L1-negative tumors, were associated with improved survival post pembrolizumab and immune-rich microenvironment. These findings support the use of chemokine profiling to complement PD-L1 and enhance patient selection for ICI therapy in TNBC. Citation Format: S. Gandhi, S. Deshmukh, S. Wu, J. Xiu, G. Sledge Jr., D. Trapani, J. Leone, S. Chumsri, M. Lustberg, C. Hong, S. Yao, K. Takabe, M. Ernstoff, M. Opyrchal, P. Chalasani, C. Paulos, G. Lesinski, A. Madabhushi, S. Badve, K. Kalinsky, P. Kalinski. Chemokine signatures improve PD-L1-based prediction of pembrolizumab response in triple-negative breast cancer [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 PS2-08-14.

Abstract Background: Male breast cancer (MaBC) is a rare malignancy, accounting for approximately 1% of all breast cancer cases. Our previous studies suggest that MaBC is biologically distinct from female breast cancer (FeBC), exhibiting a unique mutational profile and immune-genomic microenvironment. While the impact of these molecular alterations on site-specific metastases has been explored in FeBC, it remains poorly understood in MaBC. This gap limits accurate risk stratification and prediction of disease trajectory in male patients. In this study, we investigated site-specific genomic and immune alterations across metastatic sites in MaBC. Methods: 238 MaBC samples were tested by NGS (592, NextSeq; WES, NovaSeq) and WTS (NovaSeq; Caris Life Sciences, Phoenix, AZ). We performed analysis in hormone receptor-positive (HR+) and HER2-negative (HER2-) subtypes which are most common in male. HR+HER2- was defined as estrogen receptor (ER) or progesterone receptor (PR) positive and HER2 IHC 1+ or 2+ with negative chromogenic in situ hybridization (CISH) assay. Immune cells were estimated using WTS deconvolution (Quantiseq). Gene expression profiles were analyzed for T-cell inflamed score, interferon-gamma (IFN-gamma) score and MAPK Activation Score. We classified patients by site of biopsy/specimen site. Statistical significance was assessed with chi-square and Mann-Whitney U tests, p<0.05 is considered statistically significant. Results: Our cohort included 86 HR+HER2- MaBC samples biopsied from primary breast cancer (pBC) (n=39), bone (n=15), liver (n=8), lung (n=8), lymph nodes (n=7), and skin (n=9) mets. We report here only the data on statistically significant differences, but the overall landscape has been depicted across all defined-sites investigated. pBC showed a significantly lower prevalence of PIK3CA mutations (23.6% vs. 53.3%, p<0.05) compared to bone mets. ESR1 mutations were absent in pBC but present in 25% of liver and 25% of lung mets (all comparisons p<0.05). Immune profiling revealed reduced infiltration of M2 macrophages (4.3% vs. 6%, p<0.05) and neutrophil (2.2% vs. 6.9%, p<0.05) in pBC relative to bone Mets. Further, CD4+ T cells were lower in pBC compared to lymph nodes (0% vs. 2.4%, p<0.05). pBC had lower expression of immune checkpoint genes (PDCD1LG2, HAVCR2 and CD47) compared to other mets. pBC exhibited lower MAPK pathway activation score (-1.6 vs. 0.17, p=0.03) and a trend toward lower T cell-inflamed gene expression scores (-65 vs. 8, p=0.06) compared to bone mets. Bone mets had higher infiltration of M2 macrophage compared to lung (6% vs. 4.4%, p<0.05) and lymph node (6% vs. 4.5%, p<0.05). Neutrophil infiltration was elevated in bone versus liver (6.9% vs. 2%, p<0.05) but T regulatory cells were lower in bone compared to lymph nodes (1% vs. 2.4%, p<0.05). Bone showed reduced expression of CD47 (median TPM 50.6 vs. 109.9, p<0.05) compared to skin. Further bone had lower IFN-gamma scores (-0.41 vs. -0.17, p<0.05) compared to lymph node. Data on prior number and types of cancer treatment to give a context to the findings will be presented at the meeting. Conclusions: Our study highlights that each MaBC metastatic site harbors a unique genomic and immune landscape that can influence tumor behavior and potentially impact patient prognosis. Better understanding of metastatic site-specific molecular features in larger cohorts can help in the future research efforts and guiding treatment decisions. Citation Format: D. Trapani, S. Deshmukh, S. Wu, J. Xiu, N. Lin, G. Curigliano, P. Spanheimer, P. Advani, P. Barreto Coelho, M. Lustberg, G. Sledge Jr, S. Tolaney, J. Leone. Site specific Genomic and Immune Landscapes Underpinning in metastatic Male Breast Cancer [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 PS2-08-04.