Abstract PO-080: Patient-derived organoids and cancer associated fibroblasts as a co-culture model to explore cell type interactions in pancreatic cancer
Authors: Zimmerman Jacquelyn W. , Stein-O'Brien Genevieve , Burkhart Richard A. , Fertig Elana J. , Jaffee Elizabeth M. 
Published: November 2021

Abstract
Introduction: Pancreatic ductal adenocarcinoma (PDAC) remains challenging to treat in part due to the dense desmoplastic stroma characteristic of the tumor microenvironment (TME). Often, less than 50% of the tumor mass is composed of malignant ductal epithelial cells. The relationship between the TME and the ductal tumor is important in tumorigenesis, metastasis, and treatment resistance. Cancer associated fibroblasts (CAFs) are an abundant cell type in the TME and have a dynamic role in both promoting and inhibiting tumor progression. Efforts to target fibroblast function therapeutically have yielded mixed results. While pre-clinical data have generated some optimism that CAF inhibition may improve clinical outcomes, this has not translated to the clinical setting. One likely reason for these inconsistent findings is the evolving understanding of CAF heterogeneity. Expanding our understanding of the interactions between the epithelial tumor and CAFs may identify new modalities for modulating the TME to augment treatment efficacy and improve patient outcomes. This requires the generation of human model systems to better interrogate intercellular interactions. Methods: We have developed an ex vivo model system to investigate interactions between ductal epithelial tumor cells and CAFs using matched patient-derived organoids (PDOs) and CAFs extracted following dissociation of surgical specimens. Flow cytometry was used to evaluate the viability of both cellular components as well as associated proliferation and cell identity. Moreover, we have used immunohistochemistry to further characterize this system while preserving the spatial relationship of the cell populations. Most recently, we employed the MULTI-seq multiplex single cell RNA sequencing (scRNA-seq) pipeline to explore the transcriptional dynamics of this system at intervals over 96 hours by examining 4 conditions with 1,500 cells sequenced per condition per time point. Results: Flow cytometry demonstrated that both PDOs and CAFs remain viable in this co-culture construct and that both cell types can be reliably identified using epithelial markers to identify organoid cells and an exclusion panel to identify the CAFs. We have further validated these findings using immunohistochemistry and demonstrated that in co-culture both PDOs and CAFs remain viable and continue to proliferate. We have also demonstrated successful extraction of both cell types from the Matrigel scaffold in which they are grown for use in downstream high dimensional genomics assays. Conclusions: We have previously demonstrated the utility of a PDO model to explore clinically relevant biological mechanisms in PDAC. However, the addition of CAFs to this model provides a more comprehensive representation of the in vivo tumor. This ex vivo system is viable and flexible for multiple end point assays and can be used to explore relevant cellular interactions and mechanisms. Our ongoing investigation will specifically focus on PDO-CAF interactions and transcriptional dynamics over time as we interrogate the scRNA-seq data.
Citation Format: Jacquelyn W. Zimmerman, Genevieve Stein-O'Brien, Richard A. Burkhart, Elana J. Fertig, Elizabeth M. Jaffee. Patient-derived organoids and cancer associated fibroblasts as a co-culture model to explore cell type interactions in pancreatic cancer [abstract]. In: Proceedings of the AACR Virtual Special Conference on Pancreatic Cancer; 2021 Sep 29-30. Philadelphia (PA): AACR; Cancer Res 2021;81(22 Suppl):Abstract nr PO-080.