FECH, a novel metabolic target influencing CAR T-cell phenotype and function.

Recent phase I/II clinical trials have demonstrated that chimeric antigen receptor (CAR) T cells targeting the disialogangliosade GD2 represent a promising therapeutic option for pediatric patients with relapsed or refractory high-risk neuroblastoma (NB). However, incomplete and heterogeneous clinical responses highlight the need to improve CAR T-cell efficacy and persistence. We previously demonstrated the therapeutic benefit of combining the dual insulin-like growth factor 1 receptor/insulin receptor (IGF1R/IR) inhibitor linsitinib (LIN) with third-generation GD2.CAR T cells in diffuse intrinsic pontine glioma, where LIN induced tumor cell death and modulated the CAR T-cell phenotype. Here, we extended these findings to NB and explored the mechanisms of LIN-mediated CAR T-cell modulation. LIN, in combination with CAR T cells, significantly enhanced antitumor activity in LIN-sensitive NB cell lines. Mechanistically, we investigated ferrochelatase (FECH), a mitochondrial enzyme involved in heme biosynthesis, and a known off-target of LIN. LIN treatment or selective FECH inhibition with N-methyl protoporphyrin IX reduced intracellular heme, attenuated activation and exhaustion marker expression and promoted central memory characteristics associated with improved in vivo CAR T-cell persistence and functionality. Both treatments similarly decreased ATP production by reducing glycolysis and mitochondrial respiration in chronical antigen-activated CAR T cells. Collectively, these data reveal a dual mechanism of action for LIN, combining direct tumor cell cytotoxicity with metabolic reprogramming of CAR T cells linked to heme biosynthesis. These findings identify heme metabolism as regulator of CAR T-cell phenotype and function and support further investigation of FECH to enhance therapeutic efficacy in NB and beyond.