PROJECT SUMMARY – RP3 Immunotherapies can induce durable responses in advanced cancer patients and are the most exciting new cancer treatments. However, as current immunotherapies directly reinvigorate preexisting anti-tumor T cells, they are ineffective in T cell-poor (“cold”) cancers, such as pancreatic ductal adenocarcinoma (PDAC). New immunotherapies that stimulate de novo immunity are therefore needed for PDAC. To identify such strategies, we discovered that compared to the majority of PDACs that are T-cell poor and “cold,” T-cell rich “hot” tumors from rare long-term PDAC survivors are infiltrated by higher densities of group 2 innate lymphoid cells (ILC2s), a recently discovered lymphocyte with tissue-specific anti-tumor function. Higher densities of tumor ILC2s and higher expression of the ILC2-activating cytokine interleukin-33 (IL33) correlated with higher CD8+ T cell density, greater T cell cytolytic activity, and 2-fold longer survival in human PDAC. Using PDAC mouse models, we found that systemic delivery of recombinant IL33 (rIL33) activated tumor ILC2s to produce the chemokine Ccl5, recruit intratumoral dendritic cells, amplify intratumoral CD8+ T cells 10-fold, and restrict tumor growth. In addition, like activated T cells, rIL33-activated ILC2s upregulated the inhibitory immune checkpoint PD-1, and combining rIL33 with PD-1 antibody blockade maximally restricted tumor growth and prolonged survival in PDAC mouse models. Thus, our laboratory studies identify rIL33 as a novel therapeutic cytokine that stimulates de novo anti-tumor immunity in PDAC both as a mono- and as a combination therapy with PD-1 blockade. However, as there are currently no drugs that target the IL33-ILC2 axis, and as human PDAC tumors are immunologically heterogeneous, the appropriate drug delivery strategy and target patient cohorts to rationally translate these findings to PDAC patients are unknown. We will address these key knowledge gaps through three specific aims to determine if rIL33 has anti-tumor efficacy (1) in immunologically heterogeneous PDAC molecular subtypes, (2) in immunologically heterogeneous PDAC primary tumors and metastases, and (3) as half-life extended clinic- compatible drugs. We will complete these aims and thereby establish the principles to translate rIL33 to patients using mechanistic studies in orthotopic PDAC mice deficient in components of the IL33-ILC2 axis, patient-derived organoid models of the tumor immune microenvironment, cellular and transcriptional analyses of matched multi- site human PDAC primary tumors and metastases, and IND-enabling toxicology studies. Through these studies, we will identify the target patient cohorts, the mechanisms of action, and the optimal drug to inform the design of a first-in-human rIL33 clinical trial in PDAC patients by the end of the study period. This work will therefore critically expand our knowledge of preclinical and translational biology of the novel IL33-ILC2 therapeutic axis, accel...