Improving CAR T Cell Persistence in Pancreatic Cancer Using an In Vivo CRISPR Knockout Screen

Abstract

Chimeric antigen receptor (CAR) T cell therapy has demonstrated remarkable success in treating a subset of hematologic malignancies. However, efficacy against solid tumors has remained limited owing, in part, to the poor CAR T cell persistence in the immunosuppressive solid tumor microenvironment. Preclinical CRISPR screening studies have improved our understanding of the mechanisms that regulate CAR T cell exhaustion. However, such work has primarily been conducted in vitro, failing to reproduce the complex challenge of the hypoxic, cytokine-depleted vivo tumor microenvironment. Here, we perform in vivo CAR T cell CRISPR knockout screens (14 or 28-day CAR engraftment) in pancreatic adenocarcinoma using a curated Mario guide library to identify genes that enhance CAR T cell persistence in vivo. Results suggest a temporal progression to the enrichment of gene hits. The short-term, 14-day Mario screen may have identified hits that improve the short-term enrichment of Mesothelin CAR T cells. However, the 28-day screen identified gene knockouts implicated in the JAK-STAT signaling pathway, such as SOCS1 and PTPN2, which demonstrate more durable tumor control.