Conditioned CAR-T cells by hypoxia-inducible transcription amplification (HiTA) system significantly enhances systemic safety and retains antitumor efficacy

Abstract

BackgroundHypoxia is a striking feature of most solid tumors and could be used to discriminate tumors from normoxic tissues. Therefore, the design of hypoxia-conditioned Chimeric Antigen Receptor (CAR) T cells is a promising strategy to reduce on-target off-tumor toxicity in adoptive cell therapy. However, existing hypoxia-conditioned CAR-T designs have been only partially successful in enhancing safety profile but accompanied with reduced cytotoxic efficacy. Our goal is to further improve safety profile with retained excellent antitumor efficacy.MethodsIn this study, we designed and constructed a hypoxia-inducible transcription amplification system (HiTA-system) to control the expression of CAR in T (HiTA-CAR-T) cells. CAR expression was determined by Flow cytometry, and the activation and cytotoxicity of HiTA-CAR-T cells in vitro were evaluated in response to antigenic stimulations under hypoxic or normoxic conditions. The safety of HiTA-CAR-T cells was profiled in a mouse model for its on-target toxicity to normal liver and other tissues, and antitumor efficacy in vivo was monitored in murine xenograft models.ResultsOur results showed that HiTA-CAR-T cells are highly restricted to hypoxia for their CAR expression, activation and cytotoxicity to tumor cells in vitro. In a mouse model in vivo, HiTA-CAR-T cells targeting Her2 antigen showed undetectable CAR expression in all different normoxic tissues including human Her2-expresing liver, accordingly, no liver and systemic toxicity were observed; In contrast, regular CAR-T cells targeting Her2 displayed significant toxicity on human Her2-expression liver. Importantly, HiTA-CAR-T cells were able to achieve significant tumor suppression in murine xenograft models.ConclusionOur HiTA system showed a remarkable improvement in hypoxia-restricted transgene expression in comparison with currently available systems. HiTA-CAR-T cells presented significant antitumor activities in absence of any significant liver or systemic toxicity in vivo. This approach could be also applied to design CAR-T cell targeting other tumor antigens.

Current advances and challenges in CAR T-Cell therapy for solid tumors: tumor-associated antigens and the tumor microenvironment

“…This HiTA system showed a significant improvement in hypoxia-restricted transgene expression over existing systems and showed significant antitumor activity in the absence of significant hepatic or systemic toxicity observed in vivo. This approach can also be applied to the design of CAR T cells directed against other tumor antigens [ 97 ]. Deepak et al developed “switch” CAR systems, the activity of which could be controlled in vivo and which bound specific peptides; the specific peptides bound to Fab molecules, and the Fab bound to tumors.…”

Enhancing CAR T-cell therapies against solid tumors: Mechanisms and reversion of resistance

“…To target the hypoxic tumor microenvironment, hypoxiainducible CAR T-cells provide tumor cytotoxicity only within the low oxygen condition and avoid on-target, off-tumor effect (85)(86)(87). Normalization of tumor vascularity can reverse hypoxia and benefit CAR T-cell therapies (88,89).…”

Advances in Promoting the Efficacy of Chimeric Antigen Receptor T Cells in the Treatment of Hepatocellular Carcinoma

“…Another team reported the feasibility of another hypoxia-inducible transcription amplification system (HiTA-System) against hypoxia, and it does not have any prominent hepatic or systemic toxicity in vivo and exhibits significant antitumor activity. The transactivator of this system is co-regulated by hypoxia response elements (HREs) and an oxygen-dependent degradation domain (ODD), which can specifically bind to its target sequence upstream of genes, subsequently initiating their transcription and translation under hypoxia [ 21 ].…”