IDR-induced CAR condensation improves the cytotoxicity of CAR-Ts against low-antigen cancers

Abstract

SummaryChimeric antigen receptor (CAR)-T cell-based therapies demonstrate remarkable efficacy for the treatment of otherwise intractable cancers, particularly B-cell malignancies. However, CAR-T cells respond to only a subset of cancers and are limited by low signaling efficiency: a few hundred or more antigen molecules are required to activate a CAR T cell whereas a single peptide-loaded MHC molecule is sufficient to trigger the activation of a normal T cell. This low antigen sensitivity not only limits the current CAR-T therapy to high antigen-expressing cancers, but also results in high frequencies of relapse, during which high antigen-expressing cancers downregulate their antigen levels to escape CAR-T’s attacking. To improve the antigen sensitivity of CAR-T cells, we engineered CARs targeting CD19, CD22, and HER2 by including intrinsically disordered regions (IDRs) that promote signaling condensation. The “IDR CARs” triggered enhanced membrane-proximal signaling in the CAR-T synapse, which led to an increased release of cytotoxic factors, a higher killing activity towards low antigen-expressing cancer cells in vitro, and an improved anti-tumor efficacy in vivo. No elevated tonic signaling was observed in IDR CAR-Ts. Together, we demonstrated IDRs as a new tool set to enhance CAR-T cytotoxicity and broaden CAR-T’s application to low antigen-expressing cancers.