Tumor-Intrinsic Gain-of-Function p53 <sup>R172H</sup> Mutation Drives Accumulation of Neutrophils in the Pancreatic Tumor Microenvironment that Promotes Resistance to Immunotherapy

Abstract

The immune microenvironment plays a critical role in cancer development, progression and resistance to therapy. Recent reports indicate different tumor genotypes can profoundly influence the composition of the immune microenvironment. After oncogenic KRAS, TP53 is the most frequently mutated gene in pancreatic ductal adenocarcinoma (PDAC). KRASG12D mutations in tumor cells can modify the immune landscape of PDAC, however, the immune effects of neomorphic TP53 mutations have not been defined. Focusing on the most common TP53 mutation in PDAC, we sought to evaluate the non-cell-autonomous role of mutant p53 in modulating the immune microenvironment of pancreatic cancer by comparing the immune profiles generated by KRASG12D-mutated mouse pancreatic ductal epithelial cells (PDEC) genetically engineered to express the Trp53 R172H gain-of-function mutation to their p53 wildtype control. We found orthotopically implanted tumors derived from KrasG12D/+;Trp53R172H/+ PDEC had a distinct immune profile in comparison to KrasG12D/+; Trp53 +/+ tumors, characterized by an influx of intratumoral CD11b+ Ly6G+ neutrophils and concomitant decreases in CD3+ T cells, CD8+ T cells, and CD4+ T helper 1 (Th1) cells. Analysis of publicly available human PDAC cohorts revealed enrichment of genes in neutrophil-related pathways in TP53 -mutated tumors. Knockdown of CXCL2, a neutrophil chemoattractant, in the tumor epithelial compartment of KrasG12D/+; Trp53R172H/+ PDEC tumors reversed the neutrophil phenotype. Depleting neutrophils in mice bearing G12D/+; Trp53R172H/+ PDEC tumors augmented sensitivity to combined CD40 immunotherapy and chemotherapy. Collectively, these data link mutant p53 to the presence of intratumoral neutrophils in pancreatic cancer and suggests that tumor genotypes could inform patient selection for immunotherapy.