L-Arginine supplementation improves the survival of virus-specific CD8+ T cells during in vitro expansion.

INTRODUCTION: Virus-specific T cell therapy is a promising treatment for life-threatening viral infections following allogeneic hematopoietic stem cell transplantation. However, its efficacy is limited by apoptotic cell loss during in vitro expansion and the cryopreservation process. MATERIALS AND METHODS: Human virus-specific T cells were expanded in vitro with interleukin-7 and interleukin-15 in RPMI 1640-based medium with or without L-arginine supplementation, and their survival, phenotype, mitochondrial function, and cytokine signaling were evaluated. RESULTS: Supplementation of the culture medium with L-arginine enhanced the survival of human virus-specific T cells during in vitro expansion. Supplementation of the RPMI 1640-based culture medium with L-arginine reduced apoptosis and was accompanied by increased expression of the anti-apoptotic protein MCL-1 in virus-specific CD8+ T cells. In parallel, modest differences in exhaustion- and differentiation-associated markers were observed. Consistent with these survival-associated changes, preservation of mitochondrial membrane potential was concurrently observed in non-apoptotic CD8+ T cells. Among several culture-associated factors that may contribute to mitochondrial preservation, L-arginine availability was associated, at least in part, with common gamma-chain cytokine-induced phosphorylation of mammalian target of rapamycin signalling and signal transducer and activator of transcription 5 downstream of interleukin-7 and interleukin-15 during in vitro culture. CONCLUSION: Together, these findings support L-arginine supplementation as a simple and clinically applicable strategy to enhance the durability of virus-specific T cell products during in vitro expansion, as well as cryopreservation and thawing.