Targeted Rejuvenation of Exhausted Chimeric Antigen Receptor T Cells Regresses Refractory Solid Tumors

Abstract

Chimeric antigen receptor (CAR) T-cell therapies have proven to be effective in treating hematologic malignancies but demonstrate only marginal efficacy in eradicating solid tumors. Although several mechanisms can account for these differences, a major cause is thought to derive from CAR T-cell exhaustion, where chronic exposure to tumor antigen can activate feedback pathways that suppress CAR T-cell cytotoxicity. We describe here a strategy to reverse this CAR T-cell exhaustion using a universal anti-fluorescein CAR that concurrently serves as (i) a cancer recognition receptor that enables engagement of multiple cancer cell clones upon addition of a cocktail of bispecific fluorescein-linked tumor-targeting ligands, and (ii) a drug-internalizing receptor that mediates uptake of a CAR T-cell activator comprised of fluorescein linked to an immune stimulant. By attaching a Toll-like receptor 7 agonist (TLR7–1A) to fluorescein, we enable the anti-fluorescein CAR to bind and internalize TLR7–1A, leading to both downregulation of exhaustion markers (i.e., PD-1, TIM3, LAG3) and reactivation of exhausted CAR-T cells without causing the toxicities commonly associated with systemic administration of TLR7 agonists. The resulting rejuvenated CAR-T cells are observed to regress otherwise refractory solid tumors. Moreover, because no other immune cells are altered by this treatment, the data demonstrate that the exhaustion state of the CAR-T cells constitutes a major property that determines the efficacies of CAR T-cell therapies in solid tumors. Implications: A novel strategy for rejuvenating exhausted CAR-T cells is described previously that promotes downregulation of exhaustion markers and renewed eradication of cancer cells in a tumor mass.

Fully equipped CARs to address tumor heterogeneity, enhance safety, and improve the functionality of cellular immunotherapies

“…The proposed mechanism involves internalization of the FITC-TLR7a – CAR complex via receptor-mediated endocytosis. This is followed by TLR-7 activation in the endosomal compartment of AdCAR-T cells ( 24 ).…”

CAR-T cell therapy for hematological malignancies: Limitations and optimization strategies

“…The possibility of rejuvenating exhausted CAR-T cells has also been explored. Luo et al ( 131 ) constructed a fluorescein-targeted-CAR-T cells with both antigen recognition and drug internalization ability. These CAR-T cells were directed toward the folate receptor + cancer cells via a fluorescein-folate bispecific adaptor.…”