Abstract 2178: Multimodal analysis of the PDAC tumor microenvironment in response to radiation and immunotherapy reveals insights into treatment resistance

Abstract Pancreatic ductal adenocarcinoma (PDAC) is a deadly disease with few treatment options and poor prognosis. Immunotherapy is effective in other cancers but shows worse outcomes in PDAC due to a desmoplastic stroma, an abundance of immunosuppressive cells, and a lack of antigen-presenting dendritic cells, which result in low immune infiltration. We previously reported a study combining dual immune checkpoint blockade (ICB) and radiation therapy in metastatic PDAC patients who progressed on at least one line of chemotherapy and showed an 18% overall response rate (ORR) and 29% disease control rate (DCR). Here, we performed a follow-up phase 2 study on previously treated PDAC patients and observed 3.6% ORR, 10.7% DCR, 2.3 months median progression-free survival, and one complete response. To uncover mechanisms of treatment resistance, we profiled 33 samples from 23 patients with 13 paired pre and on treatment biopsies across distinct tissue sites including liver metastases and primary pancreas. Using single nucleus sequencing (>240k cells), single-cell RNA sequencing (>160k cells) with TCR (scTCR) sequencing (>25k cells), and Visium spatial transcriptomics of a subset of samples (n=15), we comprehensively characterized the cell states and gene programs in the TME of metastatic PDAC prior to and after radiation and ICB. Post radiation, tumor cells were enriched for classical states, exhausted CD8 and proliferating T cells were expanded, and interferon-related programs were upregulated in epithelial subtypes, C1QC+ and MHCII+ macrophages, and CD14+/CD16+ monocytes, suggesting cell type-specific responses to treatment. Expanded T cell clonotypes showed phenotypic shifts toward CD8 exhausted and cytotoxic activated states. Using a published gene signature, we identified tumor-reactive T cells, which correlated with known proxies for tumor reactivity including CXCL13 expression. Only 2.2% of T cells were tumor-reactive, and 71% of these came from the single patient who displayed complete response to treatment. Moreover, bulk TCR sequencing of serial peripheral blood samples showed long-term persistence of tumor-reactive clonotypes in this patient over a year post-treatment. Spatial transcriptomics demonstrated co-localization of CXCL13+ T cell, dendritic cell, and inflammatory chemokine gene programs, and separate co-localization of macrophage, CCL8+ CAF, Th17/Tc17, Class II MHC antigen presentation, and interferon stimulated gene programs. Together, our multimodal dataset offers valuable insight into radiation and immunotherapy-induced shifts in the TME. Citation Format: Sarah Kang, Aparna R. Parikh, Milan Parikh, Julie L. Koenig, Leon Pappas, Moshe Sade-Feldman, Lynn Bi, Nicole Carzo, Tarin M. Grillo, Islam Baiev, Olanike Asupoto, Irena Gushterova, Tom LaSalle, Anna Gonye, David P. Ryan, David T. Ting, Dana Pe'er, Theodore S. Hong, Nir Hacohen, Ryan J. Park, Arnav Mehta. Multimodal analysis of the PDAC tumor microenvironment in response to radiation and immunotherapy reveals insights into treatment resistance [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 2178.