通过小激活 RNA 过表达 HIF-1α 增强 CAR-T 细胞在缺氧微环境中的功能

CAR-T cell therapy demonstrates high efficacy in hematological malignancies but remains limited in solid tumors, mainly due to the immunosuppressive hypoxic tumor microenvironment that impairs T cell infiltration and promotes exhaustion.

Driven by clinical transcriptomic and protein evidence implicating hypoxia-inducible factor-1 (HIF-1 ) in metabolic reprogramming, we developed a non-viral CAR-T cell preparation platform using a piggyBac transposon system to co-deliver a CAR construct and a small activating RNA (saRNA) to drive endogenous HIF-1 overexpression (HIF1A OE ). HIF1A OE -CAR-T cells exhibited superior tumoricidal activity under hypoxic conditions (1% O ) and enhanced infiltration in a 3D tumor spheroid model.

Importantly, these beneficial effects were successfully validated across independent antigen-targeting CAR models. Metabolomic analyses and Seahorse metabolic flux profiling revealed that HIF-1 remodels cellular metabolism through a dual mechanism: augmenting aerobic glycolysis by upregulating GLUT1 and preserving mitochondrial integrity via the Nrf2/PGC-1 signaling axis.

Furthermore, combining HIF1A OE -CAR-T cells with an anti-CTLA4 nanobody resulted in synergistic tumour regression and improved infiltration in SCID-NOD mice, without significant toxicity. Collectively, activating endogenous HIF-1 in CAR-T cells is proposed as a promising strategy for the safe and effective treatment of solid tumors within hypoxic microenvironments.