IL-21 maintains CD62L expression during NKT-cell ex vivo expansion and enhances antitumor activity of NKT cell therapy in vivo

Abstract

Abstract Vα24-invariant Natural Killer T cells (NKTs) have potent antitumor properties and are being developed for cellular immunotherapy of cancer. Such therapy requires extensive ex vivo expansion of NKTs while preserving their longevity and function. In a recent report from our group, CD62L+ subset of NKTs has been shown to have longer persistence in vivo and stronger antitumor activity than CD62L− counterpart. However, the requirements for the preservation of CD62L+ NKTs during ex vivo expansion remain largely unknown. The gene expression analysis comparing CD62L+ and CD62L− NKTs revealed a significantly higher expression level of IL-21R mRNA in the former. FACS analysis confirmed the finding at the cell surface protein level. These led us to hypothesize that IL-21 preferentially supports CD62L+ NKTs. To test this hypothesis, we expanded primary human peripheral blood NKTs using in vitro stimulation with their cognate antigen, α-galactosylceramide. The culture was supplemented with IL-2, IL-21, or both cytokines. We found that in contrast to IL-2, IL-21 alone failed to support NKT-cell expansion. A combination of IL-2 and IL-21 led to similar absolute numbers of NKTs compared with IL-2 alone. However, the proportion of CD62L+ NKT-cell subset was significantly higher in the culture with IL-2/IL-21 combination compared with IL-2 alone after both primary and secondary expansions. Furthermore, after transfer to NSG mice, IL2/IL-21-expanded NKTs persisted significantly longer and had higher therapeutic efficacy in a lymphoma model compared with IL-2-expanded NKTs. Our results suggest that inclusion of IL-21 in the NKT-cell expansion protocol, which currently uses IL-2 alone, would increase antitumor potential of NKT cell therapy.