Spatial transcriptomics reveals heterogeneity and pathway dependencies of cancer associated fibroblasts in pancreatic ductal adenocarcinoma

Abstract Pancreatic ductal adenocarcinoma (PDA) is a deadly disease characterized by an immunosuppressive microenvironment and a dense stroma that encapsulates the tumor, making therapeutic targeting particularly challenging. One of the main cellular components of PDA tumor microenvironment (TME) is cancer associated fibroblasts (CAFs), whose function is to sustain tumor growth and provide structural support to the TME. Previously, we established the existence of three transcriptional subtypes of CAFs, myCAF, iCAF and apCAF, each with distinct functions and location in respect to cancer cells. Two of these subtypes, myCAF and iCAF, display a highly plastic potential in vitro and can transdifferentiate into one another upon activation of specific signaling pathways. The plasticity observed supports the existence of transitional CAF sub-states that have previously eluded observation due to technical limitations to their isolation. Through a combination of spatial transcriptomics and fluorescent in situ hybridization (FISH) on tumor samples obtained from KPC (KrasLSL-G12D; p53LSL-R172H; PDX-CRE) and FPC (KrasFrt-LSL-G12V-Frt; p53LSL-R172H; PDX-CRE; Rosa26FlpOERT2) mice, two PDA mouse models displaying a Kras G12D and G12V mutation respectively, we were able to confirm the presence of the previously identified CAF subtypes and two additional CAF sub-states. These sub-states are associated with the expression of specific markers and display different pathway enrichment. Notably, Kras G12D and G12V mutations generate similar organization of the stroma and heterogeneity of the CAF population. Subsequently, to understand the contribution of activating Kras mutation to these CAF phenotypes, we utilized the unique feature of the FPC mouse model which allows the excision of Kras G12V once the tumor has fully developed. Through FISH analysis, we observed deep reorganization of the stroma and a shift in marker expressions within the CAF populations upon Kras excision. Overall, the data here presented offers insight into the diversity of PDA CAFs and the role of mutated Kras in regulating CAF subtypes and stromal organization. Citation Format: Giuseppina Caligiuri, Jennifer Thalappillil, Juliene Hinds, Elise T. Courtois, William F. Flynn, Paul Robson, Alexander Dobin, Youngkyu Park, David A. Tuveson. Spatial transcriptomics reveals heterogeneity and pathway dependencies of cancer associated fibroblasts in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1563.