Abstract PD7-02: Transcriptomic profiling of the tumor microenvironment indicates differential response to neoadjuvant chemotherapy with and without pembrolizumab in early TNBC

Abstract Background: Neoadjuvant chemotherapy (CT) with pembrolizumab (P) has significantly improved pathologic complete response (pCR) and survival outcomes compared to CT alone in patients (pts) with stage II-III TNBC. Predictive markers of response to the addition of P to CT are lacking. We compared tumor microenvironment (TME) composition inferred by gene expression at baseline (BL) and post-treatment (if residual disease, RD) in pts who received neoadjuvant CT vs CT-P. Methods: From the prospective DFCI Multicenter TNBC Registry, FFPE research or archival BL biopsies and surgical samples (if RD) from pts who received neoadjuvant CT or CT-P were reviewed for tumor content. RNA expression was quantified using whole transcriptome sequencing (WTS) of isolated RNA. TME cell type abundance was estimated using TIMER3 deconvolution methods. Comparison between groups was performed using linear models with tumor abundance as a covariate and paired samples as a random effect intercept. Associations with recurrence free (RFS) and overall survival (OS) used a Cox proportional hazards model adjusted for stage and pCR and a likelihood ratio test. Results: From 5/2019-1/2024, 125 pts with TNBC were identified: 50 CT, 75 CT-P. Median age at diagnosis was 51 (IQR: 42-61). Anatomic stage I, II and III was 13 (26.0%), 32 (64.0%) and 5 (10.0%) in the CT group; 5 (6.7%), 53 (70.7%) and 17 (22.7%) in the CT-P group. Neoadjuvant regimens included an anthracycline in 41/50 and 63/75, taxane in 50/50 and 73/75, and platinum in 10/50 and 72/75 pts, respectively. Germline BRCA1/2 status was known in 109 pts: 5/46 (10.9%) CT and 14/63 (22.2%) CT-P had pathogenic variants. BL biopsies were analyzed with WTS in 104 pts; 20/37 (54.1%) CT and 39/67 (58.2%) CT-P had pCR. Among 45 pts with RD, 43 RD samples were analyzed (35 with paired BL biopsy). Pts who experienced pCR had higher BL abundance of almost exclusively proinflammatory TME cell types (FDR0.05) that included both lymphoid (cytotoxic CD8, memory CD4 and CD8, gamma delta T, B, NK, neutrophils) and myeloid (antitumor-like tumor-associated macrophages (TAMs), dendritic) cell subtypes compared to pts with RD. Higher dendritic cells at BL were significantly associated with pCR to CT-P (p=1.6e-04) while a weaker signal was seen with CT alone (p=0.47) (difference in cell type abundance in pCR vs RD between CT-P vs CT, FDR 0.20). The RD TME was characterized by a wound-healing response (higher total and suppressive TAMs, cancer-associated fibroblasts (CAFs), neutrophils, mast cells; lower antitumor-like TAMs) and proinflammatory lymphocytes (higher CD8 T cells, cytotoxic cells, activated NK cells; lower Tregs) (FDR0.05) compared to BL samples. When comparing RD to BL samples in pts who received CT and CT-P, the difference in total TAM abundance was weaker in the CT-P group (FDR0.15), driven by lower suppressive TAMs in the TME after CT-P. In pts with RD, higher TAMs in the RD TME were associated with a favorable trend for RFS (p=0.1) and improved OS (p=0.003), which was significant for CT-P (p=0.01) and not CT (p=0.1). Conclusions: Proinflammatory TME in TNBC at BL was associated with pCR to neoadjuvant therapy, and higher dendritic cells identified pts more likely to benefit from the addition of P to CT. Combined proinflammatory (higher cytotoxic cells, lower Tregs) and anti-inflammatory (higher CAFs and suppressive TAMs) changes were observed after treatment. Lower suppressive TAMs after CT-P suggests presence of a more favorable TME compared to RD after CT alone. Additional analyses will be presented exploring associations with survival, and both tumor-intrinsic and TME changes in BL and RD samples with CT vs CT-P. Whole genome, single-nucleus RNA and methylation profiling are ongoing to further identify markers of response to neoadjuvant CT with and without P. Citation Format: J. Gomez Tejeda Zanudo, S. Kirschner, A. Patel, E. Christoforou, A. Barkell, J. Baginska, B. Binboga Kurt, B. Koca, O. Cunningham, C. Stever, T. Parker, T. Rahman, B. Cross, H. Rimmer, T. Carr, M. Luo, I. Martino, B. Drummey, S. A. Virani, K. Santos, J. Bsat, C. Snow, N. Tung, S. Lo, M. Faggen, N. Sinclair, N. Ahmad, M. Constantinou, S. Sinclair, J. L. Meisel, T. A. King, J. L. Guerriero, E. A. Mittendorf, N. U. Lin, E. P. Winer, E. C. de Bruin, S. M. Tolaney, A. Moeini, A. C. Garrido-Castro. Transcriptomic profiling of the tumor microenvironment indicates differential response to neoadjuvant chemotherapy with and without pembrolizumab in early TNBC 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 PD7-02.