Abstract 1359: iNOS inhibition reprograms the tumor microenvironment enhancing antitumor immunity in obesity associated triple negative breast cancer (TNBC)

Abstract Purpose of the study: Obesity fosters an adverse tumor microenvironment (TME) characterized by immune suppression, metabolic dysfunction, and pro-tumoral cytokines. Elevated nitric oxide (NO) levels produced by inducible nitic oxide synthase (iNOS) exacerbate vascular dysfunction and metastasis. This study investigates whether combining the NOS inhibitor N(G)-monomethyl-L-arginine (L-NMMA) with docetaxel enhances antitumor efficacy in an obese TNBC model. Experimental Procedures: An orthotopic TNBC model was developed by injecting E0771 cells into the mammary fat pad of C57BL/6 mice fed either a normal diet (ND) or high-fat diet (HFD). Upon tumors reaching 100 mm3, mice were randomized into vehicle and L-NMMA treatment groups. Immune phenotyping and TME composition were assessed using digital spatial transcriptomics. Results: HFD mice showed significantly accelerated tumor growth and elevated pro-inflammatory cytokines (TNF-α, IL-6, IL-17) and G-CSF levels, alongside reduced T-cell recruiting chemokines (Eotaxin, RANTES), indicative of a pro-tumoral inflammatory state. L-NMMA combined with docetaxel reduced tumor growth and lung metastases compared to single treatments. Heatmap clustering revealed HFD tumors exhibit a distinct immune profile compared to ND or HFD+LNMMA tumors. L-NMMA treatment decreased immunosuppressive macrophages and exhaustion markers, shifting the TME toward an immune-supportive phenotype like ND tumors. L-NMMA-treated HFD tumors displayed decreased expression of proliferation (Ki67) and survival markers (p53, pAktSer473). Immune profiling revealed fewer cancer stem cells (CSCs; CD44+CD24-), M2 macrophages (F4/80+CD206+), and exhausted CD8+ T cells (PD1+) in LNMMA-treated tumors, alongside increased CD4+ T cells, dendritic cells, and plasma cells. Moreover, epithelial-mesenchymal transition (EMT) markers shifted, with elevated epithelial cells (CD71+) and reduced mesenchymal cells (Vim+), further supporting an antitumor shift. Conclusion: L-NMMA effectively remodels the obesity-altered TME, reducing tumor growth and metastasis while promoting an immune-supportive environment. Targeting iNOS with L-NMMA in combination with chemotherapy emerges as a promising therapeutic strategy to enhance TNBC treatment, particularly for obese patients. Citation Format: Ivonne Uzair, Fotis Nikolo, Sean Hynes, Gianmarco Melone, Karina Ortega, Jenying Deng, Wei Qian, Jianying Zhou, Liliana Guzman, Christoforos Thomas, Keith Chan, Jenny Chang. iNOS inhibition reprograms the tumor microenvironment enhancing antitumor immunity in obesity associated triple negative breast cancer (TNBC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 1359.