Systemically administered low-affinity HER2 CAR T cells mediate antitumor efficacy without toxicity

Abstract

BackgroundThe paucity of tumor-specific targets for chimeric antigen receptor (CAR) T-cell therapy of solid tumors necessitates careful preclinical evaluation of the therapeutic window for candidate antigens. Human epidermal growth factor receptor 2 (HER2) is an attractive candidate for CAR T-cell therapy in humans but has the potential for eliciting on-target off-tumor toxicity. We developed an immunocompetent tumor model of CAR T-cell therapy targeting murine HER2 (mHER2) and examined the effect of CAR affinity, T-cell dose, and lymphodepletion on safety and efficacy.MethodsAntibodies specific for mHER2 were generated, screened for affinity and specificity, tested for immunohistochemical staining of HER2 on normal tissues, and used for HER2-targeted CAR design. CAR candidates were evaluated for T-cell surface expression and the ability to induce T-cell proliferation, cytokine production, and cytotoxicity when transduced T cells were co-cultured with mHER2+ tumor cells in vitro. Safety and efficacy of various HER2 CARs was evaluated in two tumor models and normal non-tumor-bearing mice.ResultsMice express HER2 in the same epithelial tissues as humans, rendering these tissues vulnerable to recognition by systemically administered HER2 CAR T cells. CAR T cells designed with single-chain variable fragment (scFvs) that have high-affinity for HER2 infiltrated and caused toxicity to normal HER2-positive tissues but exhibited poor infiltration into tumors and antitumor activity. In contrast, CAR T cells designed with an scFv with low-affinity for HER2 infiltrated HER2-positive tumors and controlled tumor growth without toxicity. Toxicity mediated by high-affinity CAR T cells was independent of tumor burden and correlated with proliferation of CAR T cells post infusion.ConclusionsOur findings illustrate the disadvantage of high-affinity CARs for targets such as HER2 that are expressed on normal tissues. The use of low-affinity HER2 CARs can safely regress tumors identifying a potential path for therapy of solid tumors that exhibit high levels of HER2.

Antibody Recognition of Human Epidermal Growth Factor Receptor-2 (HER2) Juxtamembrane Domain Enhances Anti-Tumor Response of Chimeric Antigen Receptor (CAR)-T Cells

“…We hypothesize that the narrow binding space of trastuzumab–CAR to the proximal HER2 domain IV may exclude the phosphatase CD45 from the CAR zone, thereby enhancing T cell activation. The toxicity of “on-target off-tumor” is influenced by HER2 CAR affinities [ 30 ]. The low-affinity HER2-CAR (nanomolar level) is adequate to eliminate tumors safely and effectively, while the high-affinity HER2-CAR (picomolar level) induces lethal on-target off-tumor toxicity.…”

Universal CAR 2.0 to overcome current limitations in CAR therapy

“…Otherwise, the antigen low expressing cancer cells may escape the CAR-mediated recognition whereas the antigen negative cancer cell subsets cannot be recognized by the CAR T cells and in due course initiate the relapse ( 91 , 105 ). Identifying the optimal CAR sensitivity and -kinetics depend on multiple factors, including the differential antigen expression in healthy versus cancerous tissue ( 106 ), the tumor microenvironment, the level of inflammation, and the responsiveness of the specific CAR design ( 12 , 102 ). The activation state and exhaustion of CAR T cells are influenced by changing conditions during the anticancer response, including the cancer burden and the cytokine milieu ( 107 , 108 ).…”