PROJECT 1 - ABSTRACT There are two distinct pathways to invasive neoplasia in the pancreas – the more common non-cystic (pancreatic intraepithelial neoplasia or PanIN) and cystic (mostly, intraductal papillary mucinous neoplasm or IPMN). Both PanIN and IPMNs occur at significantly higher rates in the population than invasive cancer, reiterating that only a fraction of precursor lesions progress to pancreatic ductal adenocarcinoma (PDAC), but determining which patient will progress is currently not possible. PanINs and IPMNs are both characterized by the common occurrence of oncogenic KRAS mutations, while the latter also harbors concomitant “hotspot” GNAS mutations in 2/3rd of cases. How these early epithelial alterations and their crosstalk with the tumor microenvironment (TME) impacts progression to cancer remains understudied. The formation of early pancreatic precursors is accompanied by the expansion of a heterogeneous fibroblast population. In advanced disease, fibroblasts become cancer associated fibroblasts (CAFs). Little is known about precursor lesion associated fibroblasts (PAFs), including prevalent differences between the two precursor subtypes. Our preliminary data show that fibroblast reprogramming occurs in early pancreatic lesions, and leads to expression of IL6, and IL33. While IL6 is secreted and known to regulate the immune microenvironment, IL33 is prevalently nuclear in fibroblasts. Interestingly, IPMNs also have extremely elevated epithelial nuclear IL33. IL33 contains a DNA binding domain and is known to regulate transcription. Based on these observations, we propose to study the mechanisms of fibroblast reprogramming and comparing fibroblast gene expression in PanINs and IPMNs (Aim 1). Further, we will study the role of fibroblast IL33 in lesion progression and establishment of the precursor lesion microenvironment (PME) in PanIN and IPMN (Aim 2). Lastly, we will determine how epithelial IL33 in IPMNs regulates the PME as well as progression to malignancy (Aim 3). We will integrate the mouse model and in vitro studies with primary human tissue and organoids generated in the Wood laboratory (see Project 3). Together, these studies will shed light on the composition and role of PAFs in different premalignant lesions of the pancreas.