Engineering natural killer cells to express immunoglobulin A Fc receptor CD89 unlocks IgA-mediated cytotoxicity responses 2309809

Abstract Introduction Natural killer (NK) cells are critical effector cells of the innate immune system. Classically, NK cells mediate antibody-dependent responses via CD16, the IgG Fc receptor. However, IgG is the minority of Ig expressed at mucosal sites, whereas IgA is > 90% of Ig in mucosal secretions. Thus, harnessing NK cells for mucosal responses via IgA is an attractive therapeutic target. In this study, we demonstrate highly efficient CD89 knock-in that enables potent IgA- and IgG-mediated responses in NK cells. Methods Primary human NK cells were expanded and activated with irradiated K562-mbIL15 cells for 7 days before nucleofection with a transposon vector including the CD89 open reading frame. Knock-in efficiency was determined 4 days post-nucleofection and CD89+ NK cells were FACS purified and expanded with feeder cells. Functional assays were carried out using CD89+ NK cells and monoclonal rituximab antibodies expressing IgG, IgA1, or IgA2 Fc regions. CD89+ NK cells were co-cultured with Raji cells (10:1 E:T) and combinations of rituximab antibodies for 6-hours before staining and data acquisition via flow cytometry. Results CD89+ NK cells retained baseline phenotypes compared to non-nucleofected NK cells, including natural cytotoxicity and co-stimulatory receptors, intracellular signaling molecules, and markers of activation. Most importantly, CD89+ NK cells also retained CD16 expression, allowing them to respond to both IgG and IgA antibody activation. CD89+ NK cells display similar activation profiles as measured by CD107a, interferon-γ, and MIP-1β when activated with IgG, IgA1, or IgA2. As expected, CD89- NK cell controls only responded to IgG and not to IgA, confirming CD89-specific activation. Conclusion NK cells represent a promising avenue for cell-based therapeutics due to various benefits compared to T cell-based therapies. Through our current study we have shown consistent and robust engineering of NK cells to express CD89 to give them the ability to simultaneously respond to IgA and IgG. Funding Source NIAID grants R21AI184422 and P01AI162242 Topic Categories Innate Immune Responses and Host Defense: Molecular Mechanisms (INM)