Development of off-the-shelf hematopoietic stem cell-engineered invariant natural killer T cells for COVID-19 therapeutic intervention.

BACKGROUND: New COVID-19 treatments are desperately needed as case numbers continue to rise and emergent strains threaten vaccine efficacy. Cell therapy has revolutionized cancer treatment and holds much promise in combatting infectious disease, including COVID-19. Invariant natural killer T (iNKT) cells are a rare subset of T cells with potent antiviral and immunoregulatory functions and an excellent safety profile. Current iNKT cell strategies are hindered by the extremely low presence of iNKT cells, and we have developed a platform to overcome this critical limitation. METHODS: We produced allogeneic HSC-engineered iNKT ((Allo)HSC-iNKT) cells through TCR engineering of human cord blood CD34(+) hematopoietic stem cells (HSCs) and differentiation of these HSCs into iNKT cells in an Ex Vivo HSC-Derived iNKT Cell Culture. We then established in vitro SARS-CoV-2 infection assays to assess (Allo)HSC-iNKT cell antiviral and anti-hyperinflammation functions. Lastly, using in vitro and in vivo preclinical models, we evaluated (Allo)HSC-iNKT cell safety and immunogenicity for off-the-shelf application. RESULTS: We reliably generated (Allo)HSC-iNKT cells at high-yield and of high-purity; these resulting cells closely resembled endogenous human iNKT cells in phenotypes and functionalities. In cell culture, (Allo)HSC-iNKT cells directly killed SARS-CoV-2 infected cells and also selectively eliminated SARS-CoV-2 infection-stimulated inflammatory monocytes. In an in vitro mixed lymphocyte reaction (MLR) assay and an NSG mouse xenograft model, (Allo)HSC-iNKT cells were resistant to T cell-mediated alloreaction and did not cause GvHD. CONCLUSIONS: Here, we report a method to robustly produce therapeutic levels of (Allo)HSC-iNKT cells. Preclinical studies showed that these (Allo)HSC-iNKT cells closely resembled endogenous human iNKT cells, could reduce SARS-CoV-2 virus infection load and mitigate virus infection-induced hyperinflammation, and meanwhile were free of GvHD-risk and resistant to T cell-mediated allorejection. These results support the development of (Allo)HSC-iNKT cells as a promising off-the-shelf cell product for treating COVID-19; such a cell product has the potential to target the new emerging SARS-CoV-2 variants as well as the future new emerging viruses.