Novel NK activating multimeric immunotherapeutic complexes (NaMix) induce cytotoxic activity against HIV-1 infected cells in vitro and in vivo
Combined Antiretroviral Therapy (cART) has transformed HIV-1 infection from a lethal disease into a chronic, manageable infectious disease. However, HIV-1 persists in viral reservoirs of latently infected CD4+ T cells containing integrated replication-competent viral DNA. cART does not eradicate these reservoirs and treatment interruption will ultimately lead to a viral rebound.
NK cells play a key role in immune-surveillance against cancer cells and virus infected cells. We developed new recombinant molecules allowing targeted multimerization of IL-15 on NK cells using single-chain fragment variables (scFvs) of the checkpoint inhibitors NKG2A or KIR2DL (NKG2A-NaMix or KIR-NaMix). We exploited the oligomerization domain of C4 binding protein (C4bp) as a scaffold to multimerize both IL-15 and the NKG2A or KIR2DL scFvs. The NaMix molecules significantly improved the cytotoxic activity of NK cells against HIV-1 positive ACH-2 cells (p<0.0001) and resistant Raji cancer cells (p<0.05) compared to controls by increasing their degranulation capacity and IFN-γ expression and secretion. Interestingly, targeting the NKG2A receptor had a stronger effect compared to the KIR2DL receptors due to its higher expression on NK cells. We further studied the effect of the lead NKG2A-NaMix in humanized NSG tg-huIL-15 mice (having functional NK cells) infected with HIV-1 and treated with cART. We observed enhanced degranulation and killing of human NK cells isolated from the spleen of mice treated with the molecule compared to PBS. Furthermore, human NK cells from mice stimulated with NKG2A-NaMix showed a higher HLA-DR expression indicating a stronger activation status.
In conclusion, a combined strategy blocking the inhibitory receptor NKG2A and multimerizing IL-15 at the surface of NK cells increased NK cell activity and cytotoxicity against HIV-1 infected cells in vitro and in vivo, and is a promising immunotherapeutic strategy towards a functional cure for HIV-1.

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