Inefficient exploitation of accessory receptors reduces the sensitivity of chimeric antigen receptors

Abstract

Chimeric antigen receptors (CARs) can re-direct T cells to target abnormal cells but their activity is limited by a profound defect in antigen sensitivity, the source of which remains unclear. Here, we show that CARs have a >100-fold lower antigen sensitivity compared to the TCR when antigen is presented on antigen-presenting-cells, but nearly identical sensitivity when antigen is presented as purified protein in isolation. Given that the TCR uses other, accessory, receptors to achieve high sensitivity, we screened prominent accessory receptors by presenting their purified ligands together with antigen. We found that ligating the adhesion receptor CD2 or LFA-1 improved antigen sensitivity for the TCR by >100-fold, whereas for CARs the improvement was <10-fold. We reproduced these findings using target cells where the CD2 and/or LFA-1 interaction were abrogated. Sensitivity can be partially restored by fusing the CAR variable domains to the TCR CD3ε subunit (also known as a TRuC) and fully restored when exchanging the TCRαβ variable domains for those of the CAR (also known as a STAR). Our study localises the defect in CAR sensitivity to inefficient use of accessory receptors and suggests approaches to increase sensitivity.

Structural principles of peptide-centric chimeric antigen receptor recognition guide therapeutic expansion

“…S26). This could be consistently rationalized by an improvement in the dissociation rate (table S4), in accordance with kinetic proofreading models for TCR activation (25)(26)(27)(28)(29)(30). The differences observed were not a result of pMHC complex stability (fig.…”

“…In an analogous manner to TCRs, both antigen density and pHLA dissociation rate from the 10LH CAR can contribute to our observation that HLA-A*23:01, which has approximately two orders of magnitude reduced binding to 10LH relative to HLA-A*24:02 (2.6 μM versus 11 nM K D ), shows potent CAR T cell killing of tumor cells (14), because both interactions show similar k d values by SPR (0.0085 s −1 versus 0.0022 s −1 ). These results allow us to establish a relationship between pHLA dissociation rate from the scFvexpressed as CAR dwell time-and CAR T cell activation, cytokine release, and cytotoxicity, established by previous studies focusing on both TCRs and CARs (28)(29)(30).…”

Mechanical forces impair antigen discrimination by reducing differences in T‐cell receptor/peptide–MHC off‐rates

“…It is well established that engagement of the T‐cell adhesion receptors CD2 (which binds CD58) and LFA‐1 (which binds ICAM‐1) improve the sensitivity of T cells to antigens (Huse, 2017 ; Siller‐Farfán & Dushek, 2018 ; Zhu et al , 2019 ; preprint: Burton et al , 2021 ). Recently, we reported that this improvement in sensitivity was progressively lost as the antigen affinity was lowered and consequently, engagement of these receptors increased the ability of T cells to discriminate antigens (Pettmann et al , 2021 ) but the mechanism remains unclear.…”

Mechanical forces impair antigen discrimination by reducing differences in T cell receptor off-rates

“…It is well established that engagement of the T cell adhesion receptors CD2 (which binds CD58) and LFA-1 (which binds ICAM-1) improve the sensitivity of T cells to antigens (9, 18, 23, 36). Recently, we reported that this improvement in sensitivity was progressively lost as the antigen affinity was lowered and consequently, engagement of these receptors increased the ability of T cells to discriminate antigens (4) but the mechanism remains unclear.…”