Cytometric analysis authenticates adjuvanticity of interferon-γ and suggests potential synergy with toll-like receptor agonists

Abstract Interferon-γ (IFN-γ) is a cytokine that plays an important role in immune regulation, especially in the activation and differentiation of immune cells. Toll-like receptors (TLRs) are a family of pattern-recognition receptors that sense structural motifs related to pathogens and alert immune cells to the invasion. Both IFN-γ and TLR agonists have been investigated as immunoadjuvants to augment the efficacy of cancer immunotherapies and vaccines against infectious diseases. In this study, we aimed to explore the potentiality of IFN-γ and TLR agonists being applied as dual adjuvant systems. In brief, murine dendritic cells were treated with IFN-γ and/or the TLR agonists, polyinosinic-polycytidylic acid (poly I:C) or resiquimod (R848). Subsequently, the cells were stained for an activation marker, cluster of differentiation 86 (CD86), and the percentage of CD86-positive cells was measured by flow cytometry. IFN-γ efficiently stimulated a considerable number of the dendritic cells, while the TLR agonists by themselves could merely activate a few compared to the control. The combination of IFN-γ with poly I:C or R848 triggered a higher amount of cell activation than IFN-γ alone. For instance, 10 ng/mL IFN-γ with 100 μg/mL poly I:C achieved 59.1% cell activation, which was significantly higher than the 33.4% positive cells obtained by 10 ng/mL IFN-γ (p= 0.0004). These results suggested that IFN-γ and TLR agonists could be applied as a complementary, dual adjuvant system to promote dendritic cell activation and antigen presentation. Additionally, there might be a synergy between the two classes of adjuvants, and further investigation is warranted to ascertain the interaction of their adjuvant activities. This work was supported by the NIDA/NIH award UG3DA048775.