Mechanistic investigations of granzyme A adjuvant effects on γ 9δ 2T cells

Abstract

Abstract γδ T cells are ideal effector cells against cancer and pathogens for numerous reasons: they are found in blood, mucosa, and lymphoid organs, develop memory responses independent of MHC restriction, present antigen, produce cytokines, and are cytolytic. An obstacle for the development of biomedical interventions targeting γδ T cells is the need for an adjuvant that promotes their expansion and is suitable for human use. The adjuvant routinely used, IL-2, has multiple drawbacks limiting its use for vaccination. Our lab has shown that memory γ 9δ 2T cells, induced by BCG vaccination, are expanded in vitro by M. tuberculosis, M.bovis(BCG), and our lab’s vaccine candidate 6-O-methylglucoselipopolysaccharide (mGLP), in the presence of IL-2. Additionally, the expanded γ 9δ 2T cells inhibit intracellular mycobacterial growth. This inhibition is dependent on granzyme A (GzmA) production by γ 9δ 2T cells, and GzmA alone can inhibit intracellular mycobacterial growth. We recently identified that GzmA can also promote expansion of γδ T cells, without exogenous IL-2, in response to mGLP and HMBPP, a phosphoantigen that stimulates all γ 9δ 2T cells. To identify γδ T cell adjuvants, in vitro expansion assays were performed using human peripheral blood mononuclear cells, dendritic cells, antigen, and adjuvant. The absolute numbers of expanded effector γδ T cells were calculated using flow cytometry after 7 days of culture. We identified that the ability of GzmA to promote γδ T cell expansion is dependent upon TLR4, CD14, and GzmA homodimerization, but independent of GzmA enzymatic activity. This new information has broad implications for vaccine design, promoting γδ T cell responses, and understanding the role of GzmA in infectious disease and cancer. Supported by grants from NIH (R01 AI048391) and Bill & Melinda Gates Foundation (OP1118659)