Allogeneic human double negative T cells as a novel immunotherapy for acute myeloid leukemia and its underlying mechanisms.

Purpose: To explore the potential of ex vivo expanded healthy donor derived allogeneic CD4 and CD8 double negative cells (DNTs) as a novel cellular immunotherapy for leukemia patients. Experimental Design: Clinical grade DNTs from peripheral blood of healthy donors were expanded and their anti-leukemic activity and safety were examined using flow-cytometry based in vitro killing assays and xenograft models against AML patient blasts and healthy donor-derived hematopoietic cells. Mechanism of action was investigated using antibody-mediated blocking assays and recombinant protein treatment assays. Results:  Expanded DNTs from healthy donors target a majority(36/46) of primary AML cells including 9 chemotherapy-resistant patient samples in vitro and significantly reduce the leukemia load in patient-derived xenograft models in a DNT-donor unrestricted manner. Importantly, allogeneic DNTs do not attack normal hematopoietic cells or affect hematopoietic stem/progenitor cell engraftment and differentiation, nor cause xenogeneic graft-versus-host disease in recipients. Mechanistically, DNTs express high levels of NKG2D and DNAM-1 that bind to cognate ligands preferentially expressed on AML cells. Upon recognition of AML cells, DNTs rapidly release IFN which further increases NKG2D and DNAM-1 ligands expression on AML cells. IFN pretreatment enhances the susceptibility of AML cells to DNT-mediated cytotoxicity, including primary AML samples that are otherwise resistant to DNTs, and the effect of IFN treatment is abrogated by NKG2D and DNAM-1 blocking antibodies. Conclusion: This study supports healthy donor-derived allogeneic DNTs as a therapy to treat patients with chemotherapy-resistant AML and also reveals interrelated roles of NKG2D, DNAM-1, and IFN in selective targeting of AML by DNTs.

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