MODELING TUMOR HETEROGENEITY IN PDAC USING 3D STROMAL-ORGANOID COCULTURES

Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal form of pancreatic cancer with limited treatment options. Additionally, PDAC patients are often diagnosed with late-stage disease, thus lowering their chances to obtain substantial treatments with therapeutics or surgery. Therefore, new therapies must be developed in order to increase survival and treatment options for patients in need. In order to address these limitations, research models such as 3D organoid cultures are being used as an alternative to traditional models such as genetically engineered murine models and monolayer cell lines. Organoid cultures, a type of 3D culture system, contain organ-like units capable of self-organization, mimicking the organ environment. We combined traditional and organoid cell culture methods in order to study the interactions of PDAC cells and pancreatic stellate cells (PSCs), a resident fibroblast population in the pancreas. Studies suggest that PSCs act as an immune surveillance barrier for drug delivery. However, opposing studies suggest that that stromal cells may potentially provide growth factors or metabolites that contribute to the proliferation of tumor cells in vivo and in organoid cultures. Thus, understanding the interactions between these cell types is crucial for development of drug therapies to enhance outcomes for patients. We hypothesized that the development of a PDAC organoid cell co-culture with primary PSCs would provide further insight on the specific metabolites which contribute to tumorigenesis and/or metastasis. In order to examine this concept, we used a transgenic PDAC tumor mouse model containing Kras and p53 mutations (LSL-KrasG12D, p53 fl/fl, Pdx-1-Cre, or “KP-/-C”) to isolate tumor organoids. Tumor cells were resuspended in matrigel and plated in complex organoid media. Prior to this process, murine DNA samples were collected for genotyping and used for PCR analysis to identify mice with the necessary mutations. Results showed that PDAC tumor organoids were highly responsive to co-cultures with PSCs, and displayed increase in cellular proliferation when exposed to metabolites alanine and pyruvate. Future studies will work to characterize metabolites and co-culture PDAC with endogenous fibroblasts in order to further understand tumor heterogeneity.