IM-02 * A scFv-BASED CAR TO REDIRECT T CELLS TO IL13R 2-POSITIVE PEDIATRIC GLIOMA

Abstract

The intent of this project is to develop antigen-specific T cells as an effective immunotherapy for diffuse intrinsic pontine glioma (DIPG) and glioblastoma (GBM), the most aggressive, uniform fatal, primary human brain tumors in children. IL13Rα2 is expressed at a high frequency in DIPG and GBM but not in normal brain, making it is a promising target for CAR T-cell immunotherapy. While other investigators have generated IL13Rα2-targeted CARs using mutated forms of IL13 as CAR binding domains, our studies indicate that these CARs also recognize IL13Rα1, raising significant toxicity concerns. To overcome this limitation we have developed a high affinity IL13Rα2-specific scFv that does not recognize IL13Rα1. The goal of this project was to generate scFv-based IL13Rα2-specific CAR (IL13Rα2-CAR T cells) and evaluate their function. We constructed a panel of IL13Rα2-CARs that contain our IL13Rα2-specific scFv as an ectodomain, a short hinge (SH) or long hinge (LH), a CD28 transmembrane domain, and endodomains that contain signaling domains derived from CD3ζ and co-stimulatory molecules. IL13Rα2-CAR T cells were generated by retroviral transduction. CAR cell surface expression varied depending on used hinge and endodomain. In cytotoxicity assays, IL13Rα2-CAR T cells only killed target cells that expressed IL13Rα2 and not IL13Rα1 confirming specificity. While all IL13Rα2-CAR T cells secreted significant levels of IFNγ in co-culture assays with the IL13Rα2+ glioma cell line U373, only short hinge CAR T cells secreted significant amounts of IL2. In vivo, injection of IL13Rα2.SH.CD28.ζ-CAR T cells into U373-bearing mice resulted in regression of glioma xenografts as judged by bioluminescence imaging. IL13Rα2.LH.CD28.ζ- or IL13Rα2.Δ-CAR T cells had no antitumor effects. In conclusion, for the first time we have generated a CAR that only recognizes IL13Rα2 and not IL13Rα1. Our results warrant further active exploration of IL13Rα2-specific scFv-based CAR T cells for the adoptive immunotherapy of pediatric brain tumors.