Abstract 654: Transcriptomic analysis ofTGF-β-targeting oncolytic adenovirus and immune checkpoint inhibitor therapy in a triple-negative breast cancer mouse model

Abstract Immunotherapy has shown promise in cancer treatment, but resistance remains a major challenge. Combining therapies that target specific pathways, such as TGF-β signaling, can enhance the effectiveness of immune checkpoint inhibitors (ICIs). Our oncolytic adenovirus (OAd), mHAdLyp.sT, containing a TGF-β decoy to reverse immune resistance to ICI treatment and the LyP-1 peptide to enhance tumor targeting, has been shown to significantly enhance anti-tumor responses and reduced lung metastasis in a 4T1 triple negative breast cancer (TNBC) mouse model when systemically delivered in combination with anti-PD-1 and anti-CTLA-4 antibodies. We also showed previously that mHAdLyp.sT systemically primes an inflamed immune cell phenotype for ICI treatment by both flow cytometry and mass cytometry (CyTOF). In this study, we investigated how the mHAdLyp.sT-ICIs combination affects the expression of key genes in major signaling and immune regulatory pathways by RNA-seq analysis, both in tumors and systemically in the spleen, at the early response stage (day 16) and experiment terminal point (day 25) in the TNBC mouse model. In tumor, both ICIs and the mHAdLyp.sT-ICIs combination treatments significantly induced immune-related genes beneficial for anti-tumor effects compared to the untreated control group at both day 16 and day 25. However, at day 16, while ICI treatment significantly escalated the expression of several proinflammatory cytokines, such as IL-6, IL-10, and IL-11, which are known to potentiate tumor resistance to immunotherapy, mHAdLyp.sT-ICIs combination treatment did not trigger this response. In the spleen, all treatments exhibited slight effects on the immune response at day 16, while at day 25, mHAdLyp.sT-ICIs combination induced a strong response with extensive rescued changes across various crucial signaling pathways in tumor progression and metastasis, including those involved in TGF-β signaling, cell adhesion, migration, angiogenesis, extracellular matrix organization, cell differentiation, gene expression regulation, protein phosphorylation, inflammation, and apoptosis. Notably, mHAdLyp.sT-ICIs combination treatment more effectively inhibited immunosuppressive genes such as Myd88 and Ptgir than ICI alone, confirming that TGF-β inhibition by mHAdLyp.sT enhanced anti-tumor immune responses and helped overcome ICI resistance. We are confident in advancing this new treatment option for TNBC and other metastatic cancers that are projected to be resistant to immunotherapy in a clinical setting. Citation Format: Nguyễn Thị Thanh Nhàn, Beniamin Filimon, Hailie Fowler, Sheetal Ganesan, Xinkun Wang, Bruce Brockstein, Weidong Xu. Transcriptomic analysis ofTGF-β-targeting oncolytic adenovirus and immune checkpoint inhibitor therapy in a triple-negative breast cancer mouse model [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 654.