Abstract P018: Mapping cellular and microenvironmental determinants of persister populations in pancreatic cancer following neoadjuvant chemoradiation
Authors: Bernard Vincent , Ku Li-Ting , Wang Tianyu , Rajapakshe Kimal , Jiang Dadi , Jacobson Galia , Min Jimin , Tzeng Ching-Wei , Bhutani Manoop S. , Guerrero Paola A. , Ludmir Ethan B. , Koong Albert C. , Taniguchi Cullen M. , Maitra Anirban , Li Ziyi , Koay Eugene J. 
Published: January 2025

Abstract
Introduction: The molecular pathways involved in the response to radiation therapy (RT) in pancreatic ductal adenocarcinoma (PDAC) remain poorly understood. Attempts to combine RT with immunotherapies in PDAC have yielded inconsistent outcomes, highlighting a critical gap in our understanding of immune dynamics and mechanisms influencing RT response. Our study aims to uncover molecular pathways and cellular subtypes associated with radioresponse. To do so, we developed a novel cell-cell interaction platform, "SpaCCI", to map ligand-receptor (LR) interactions, essential for understanding cellular communication within the TME and its contribution to therapeutic resistance. Methods: Single-cell RNA sequencing (scRNA-seq) was performed on 16 matched pre- and post-treatment PDAC biopsies from a Phase I/II dose-escalation clinical trial of stereotactic body radiation therapy. Additionally, spatial transcriptomics (ST) was conducted on resected PDAC (20 treated with neoadjuvant chemoradiation, 8 untreated). We integrated scRNA-seq and ST datasets using cell type decomposition and developed SpaCCI to enhance LR interaction detection in tissues with complex cell mixtures. Results: ScRNA-seq identified five molecular subpopulations of cancer cells associated to RT response, with PDAC_3 enriched post-RT and marked by genes involved in lipid metabolism and ferroptosis defense. Post-RT depleted subpopulations, such as PDAC_0 and PDAC_prolif, saw upregulation of pathways related to rapid proliferation and oxidative phosphorylation. In a murine model, PDAC_3 was similarly enriched in post-RT tumors, with persister cell lines from these tumors showing overexpression of ferroptosis defense genes and resistance to ferroptosis inducers including Erastin and RSL3. ST analysis also revealed spatial immune changes post-RT, with increased cytotoxic and effector memory T cells balanced by regulatory T cells and M2-like macrophages, indicating immune escape. NK and memory B cells correlated with improved survival post-RT, while plasma IgG cells localized outside the tumor bed, suggesting compartmentalized immune responses. Using SpaCCI, we observed differential enrichment of ligand-receptor interactions in RT-treated tumors, particularly among lipid-associated tumor macrophages, cancer cells, myofibroblast-like cancer-associated fibroblasts, and endothelial cells. Persister PDAC_3 subpopulations showed enriched LR SEMA3 signaling with CAFs, myeloid cells, and endothelial cells consistent with axon guidance pathways. Conclusions: This study presents a molecular and cellular atlas of PDAC response to RT, revealing persister cells populations with adaptations in lipid metabolism and ferroptosis defense that could be targeted to enhance RT efficacy. ST highlighted complex immune dynamics within the TME demonstrating a balance between inflammatory and immunosuppressive populations. Our findings provide a foundation for combinatorial therapeutic strategies and suggest that axon guidance pathways may also play an important role in the response to RT in PDAC.
Citation Format: Vincent Bernard, Li-Ting Ku, Tianyu Wang, Kimal Rajapakshe, Dadi Jiang, Galia Jacobson, Jimin Min, Ching-Wei Tzeng, Manoop S. Bhutani, Paola A. Guerrero, Ethan B. Ludmir, Albert C. Koong, Cullen M. Taniguchi, Anirban Maitra, Ziyi Li, Eugene J. Koay.Mapping cellular and microenvironmental determinants of persister populations in pancreatic cancer following neoadjuvant chemoradiation.[abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Translating Targeted Therapies in Combination with Radiotherapy; 2025 Jan 26-29; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(2_Suppl):Abstract nr P018