Solid tumors such as pancreatic ductal adenocarcinoma (PDAC) are comprised of transformed cancer cells along with a participatory population of stromal cells. These include cancer-associated fibroblasts (CAFs), which are known to provide fibrotic stroma (desmoplasia), which supports tumor proliferation. My preliminary data suggest that fibroblasts in solid tumor stroma are locally-recruited and activated, and proliferate in a highly clonal manner. Such clonality suggests the presence of tissue-resident fibroblast progenitors. Changes in chromatin accessibility are suspected to play a critical role in driving fibroblast activation. Characterization of activated fibroblasts is a key step toward developing novel stromal therapies against cancer. In this project, I aim to 1) quantitate the local, clonal expansion of activated fibroblasts in PDAC using the Rainbow mouse, a stochastic, multi-colored, lineage-tracing model. I will use the EMBLEM (Epigenome and Mitochondrial Barcode of Lineage from Endogenous Mutations) protocol, which analyzes somatic mutations in mitochondrial DNA from single cell (sc)-assay for transposase-accessible chromatin (ATAC)-seq data to track cell lineages, in order to identify and characterize putative fibroblast progenitors. To further explore the biology underlying fibroblast activation in PDAC, I will 2) conduct sc-RNA-seq on mouse and human PDAC CAFs to identify functionally relevant changes in gene expression that occur with fibroblast activation. I will correlate CAF gene expression patterns with changes in chromatin accessibility based on the nuclear reads obtained in my sc-ATAC-seq dataset. DNA- binding elements such as Ets1 which are known to trigger Focal adhesion kinase (FAK)-mediated signaling, and FAK pathway signaling is suggested to play a significant role in CAF-associated desmoplasia in solid tumors. My preliminary data suggest that PDAC CAFs show significant upregulation of CXCL5 and CXCL10 expression, which are downstream chemokine targets of the FAK pathway. As such, I will 3) determine functional effects and explore the therapeutic potential of modulating these chemokines in activated PDAC CAFs using in vitro and in vivo assays. I will correlate modulation of these putative therapeutic targets with specific changes in chromatin accessibility using protein-indexed (Pi)-ATAC, which permits correlation of chromatin accessibility with protein expression at a single cell level. The results of these aims will expand our understanding of CAF biology and heterogeneity in PDAC. This project is expected to identify novel stromal targets that could be exploited therapeutically either primarily or adjunctively in the setting of solid tumors.
The proliferation of transformed epithelial cells in cancer is accompanied by an expansion of fibroblasts, which appear to develop from progenitor cells that are activated in the local environment and deposit dense scar tissue making tumors firm. A comprehensive understanding of what drives the activation of these fibroblasts in pancreatic cancer, which shows one of the most profound fibrotic responses of any solid tumor type, will expand our knowledge of fibroblast biology and heterogeneity. This knowledge has the potential to unveil novel targets for cancer therapies that modulate tumor fibroblasts and could be used independently or alongside currently available chemo- or immunotherapies.
