VEGFR-2 redirected CAR-T cells are functionally impaired by soluble VEGF-A competition for receptor binding

VEGFR-2 redirected CAR-T cells are functionally impaired by soluble VEGF-A competition for receptor binding

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Background The adoptive transfer of chimeric antigen receptor (CAR)-T cells has emerged as a potent immunotherapy against some hematological malignancies but not yet for epithelial-derived solid tumors.One critical issue is the paucity of broadly expressed solid tumor antigens (TAs), and another is the presence of suppressive mechanisms in MENS T-SHIRTS the tumor microenvironment (TME) that can impair CAR-T cell homing, extravasation and effector functions.TAs expressed by endothelial cells of the tumor vasculature are of clinical interest for CAR therapy because of their genomic stability and accessibility to circulating T cells, as well as their expression across multiple tumor types.

In this study, we sought to explore limitations to the efficacy of second-generation (2G) murine CAR-T cells redirected against the vascular endothelial growth factor receptor-2 (VEGFR-2) with the well-characterized single-chain variable fragment DC101.Methods Primary murine T cells were retrovirally transduced to express a 2G anti-VEGFR-2-CAR, and the in vitro binding to VEGFR-2, as well as reactivity against TA-expressing cells, was evaluated in the absence versus presence of exogenous VEGF-A.The CAR-T cells were further tested in vivo for tumor control alone and in combination with anti-VEGF-A antibody.

Finally, we performed ex vivo phenotypic analyses of tumor-infiltrating CAR-T cells for the two treatment groups.Results In line with previous reports, we observed poor control of B16 melanoma by the 2G anti-VEGFR-2 CAR-T cells as a monotherapy.We further showed that VEGFR-2 is not downregulated by B16 melanoma tumors post treatment, but that its soluble ligand VEGF-A is upregulated and furthermore competes in vitro with the CAR-T cells for binding to VEGFR-2.

This competition resulted in impaired CAR-T cell adhesion and effector function in vitro that could be restored in the presence of anti-VEGF-A antibody.Finally, we demonstrated that coadministration of anti-VEGF-A antibody in vivo promoted CAR-T cell persistence and tumor control and was associated with reduced frequencies Personal Care of PD-1+ Ki67- and LAG-3+ Ki67- CAR-T cells in the TME.Conclusions This study represents the first example of impaired function of a vasculature-targeted CAR by an angiogenic ligand and rationalizes the use of combinatorial therapies that target the tumor vasculature and augment CAR-T cell effector function.