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Abstract 3266: Distinct immune stimulatory functions of anti-human VISTA antibodies are determined by the antibody isotype

The clinical success of immune modulatory antibodies in oncological indications has demonstrated the importance of harnessing the immune system for the treatment of cancer. VISTA (PD-1H) is an Ig superfamily cell surface molecule mainly expressed on myeloid cells, but also to some extent on NK cells and T cells. Substantial published data supports the view that VISTA-binding antibodies or administration of VISTA itself can provide an inhibitory signal to T cells though the mechanism of action remains unclear including whether this activity is mediated directly or indirectly. We present here novel findings regarding the impact of different IgG isotypes on the activity of anti-human VISTA antibodies with immune stimulatory effects on primary human immune cells. The immune stimulatory effects of IgG antibodies were found to differ between variants with an identical antigen-binding domain but with different IgG isotypes (IgG4 and IgG1). First, we observed that both variants increased ongoing immune reactions, while the IgG1-variant was additionally able to activate resting immune cells. In a mixed leukocyte reaction (MLR) with healthy donor PBMCs, the ongoing immune reaction was effectively augmented by both, anti-VISTA-IgG4 and anti-VISTA-IgG1 isotype antibodies, as observed by increased levels of cytokines, e.g. TNF. However, in an experimental culture system of unstimulated PBMC of single healthy donors, only VISTA-IgG1 isotype antibodies induced increased levels of activation markers, e.g. upregulation of HLA-DR on resting myeloid cells. Second, we showed that Fc-receptor binding was a necessary interaction for immune activation as preventing the Fc-gamma-receptor binding to either of the two isotypes abolished the effects. The effects observed were specific as non-VISTA-binding isotype control mAbs were inactive. We then investigated the effects of anti-VISTA-IgG4 isotype antibodies on cells from AML patient samples by pharmacoscopy, a novel microscopy analysis method able to robustly quantify cell-cell proximity as a measure of cellular interactions. Here, VISTA-IgG4 isotype antibodies augmented interactions between multiple immune cell populations as well as between immune cells and AML cancer cells. This augmented interaction was abolished by blockade of Fc-gamma-receptor interactions, confirming our previous results. In summary, the immune stimulatory effects of anti-VISTA antibodies are defined by the antibody isotype and interaction with Fc-gamma-receptors, highlighting the importance of understanding these interactions when designing immune stimulatory antibody therapeutics for IO applications. Citation Format: Sven Mostbock, Helen Wu, Timothy Fenn, Christina Taubert, Gregory Vladimer, Anne Vogt. Distinct immune stimulatory functions of anti-human VISTA antibodies are determined by the antibody isotype [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3266.