In vitro modeling of tumor-stroma-crosstalk in pancreatic cancer leads to new insights into drug resistance and discovery

Pancreatic ductal adenocarcinoma (PDAC) remains a devastating disease despite decades of research. The 5-year-survival rate of PDAC patients remains at a dismal 11%(1) and worse still has not significantly improved over time. One key component which sets PDAC apart is the special role of the stromal cells in the tumor and the dense and restrictive extracellular matrix (ECM) which partially derives from them. Given the importance of the stromal component of PDAC, this research project aimed to investigate the role of stromal cells in drug-cancer-cell-interactions, as well as include the tumor-stroma-crosstalk when screening for tumor suppressive drugs. Paper I serves as an introduction to the cell culture models and their application in PDAC and discusses the benefits and drawbacks of different of them. Paper II describes a global assessment of the impact of crosstalk in PDAC. By choosing cell lines from human and mouse we could differentiate the RNA expression changes in the cancer cells and in the stromal cells. A key protein identified in this study was cellular communication network factor 1 (CCN1) which was further investigated in Paper III. Panc1-CCN1-KO cell lines were created and characterized. These newly established cell lines were more resistant to Gemcitabine. This increased resistance could be linked to the correlated downregulation of the Gemcitabine-metabolizing enzyme deoxycytidine kinase (DCK). Restoring CCN1 expression also restored DCK expression suggesting involvement in the same signaling pathway. Paper IV describes a newly developed method for spheroid viability, suited for the application in a high throughput drug screen. This image-based assay classifies treatment outcomes based on morphological features. This assay was further trained to identify shifts in apparent cell morphology shifts. Paper V describes the preliminary results of the high throughput screen (HTS). The results were selected based on targeting Panc1 cells over human pancreatic stellate cells (hPSCs) with the newly developed screening tool described in Paper IV. The compound CAY10581 showed synergistic effects with Gemcitabine in preliminary experiments. List of scientific papers I. Gündel B, Liu X, Lohr M, Heuchel R. Pancreatic Ductal Adenocarcinoma: Preclinical in vitro and ex vivo Models. Front Cell Dev Biol. 2021;9:741162. https://doi.org/10.3389/fcell.2021.741162 II. Liu X, Gündel B, Li X, Liu J, Wright A, Löhr M, Heuchel R. 3D heterospecies spheroids of pancreatic stroma and cancer cells demonstrate key phenotypes of pancreatic ductal adenocarcinoma. Translational oncology. 2021;14(7):101107. https://doi.org/10.1016/j.tranon.2021.101107 III. Gündel B, Liu X, Pfützenreuter A, Engelsberger V, Weiskirchen R, Löhr J-M, Heuchel R. The Crosstalk Analysis between mPSCs and Panc1 Cells Identifies CCN1 as a Positive Regulator of Gemcitabine Sensitivity in Pancreatic Cancer Cells. Int J Mol Sci. 2024;25(17):9369. https://doi.org/10.3390/ijms25179369 IV. Gündel B, Gündel A, Axelsson H, Timpone A, Löhr M, Heuchel R. Applying an automated image analysis to quantify pancreatic cancer spheroid viability and classify morphology in a high throughput drug screen. [Manuscript] V. Gündel B, Heffinger L, Timpone A, Axelsson H, Gündel A, Löhr M, Heuchel R. Reporting on the outcomes of a phenotypic high throughput screen on Panc1/hPSC heterospheroids. [Manuscript]