VISTA inhibitor III is a small-molecule ligand for immune checkpoint VISTA with IC50 of 480 nM in FRET assay.
The binding affinity of VISTA inhibitor III to VISTA was confirmed by competitive ELISA with IC50 value of 716 nM, Kd of 247 nM in SPR assay.
VISTA inhibitor III blocks VISTA-VISTA interaction resulting in decreased IL-2 signaling in Promega VISTA bioluminescent cell-based T-cell activation assay.
Similarly, to VISTA antibody, addition of VISTA inhibitor III to co-cultured Jurkat T cells with 8 cancer cell lines selectively restored the proliferation of T cells in the presence of high VISTA expressing cell lines (OVKATE, SKOV3, COV504, RL952, HEC1A) while minimal effect was evident for cancer cell lines with low VISTA expression (A2780, AN3CA, Ishikawa).
VISTA inhibitor III also upregulated interferon gamma (IFNγ) and tumor necrosis factor-alpha (TNFα), cytokines secreted by activated T cells in in the supernatants of co-cultures of purified human T cells with OVKATE and RL952 cell lines with high VISTA expression.