Background VISTA is a negative checkpoint regulator prominently expressed in the TME of a wide variety of cancers. In a preclinical model of colorectal cancer, monotherapy of small tumors (40 mm³) with anti-VISTA results in markedly slowed tumor growth. Mice bearing significantly larger tumors (600 mm³) are resistant to anti-PD-1 and anti-CTLA4 treatment and all mice die following treatment, indicating checkpoint resistance. Inclusion of anti-VISTA leads to complete rejection of 50% of tumors.

Methods The underlying therapeutic mechanisms of leading to enhanced anti-tumor immunity in both models was investigated by high-dimensional scRNAseq of the CD45+ immune infiltrate of tumors 10 days after treatment initiation.

Results In both modes, anti-VISTA treatment stimulated several pathways involving myeloid activation and antigen-presentation. Multi-spectral imaging of anti-VISTA treated tumors supported increased antigen presentation, and suppression assays showed that the myeloid infiltrate was less suppressive to T cells. Transcriptional analysis of tumor-specific CD8 T cells showed that anti-VISTA therapy induced T cell pathways highly distinct from the anti-exhaustion effects of anti-PD-1 therapy.

Conclusions These data document the unique and complementary impact of targeting VISTA in contrast to PD-1 and CTLA-4 in both the myeloid and T cell lineages. These mechanistic insights strongly support the use of anti-VISTA to overcome the checkpoint resistance seen in contemporary treatments involving PD-1.

Ethics Approval All mouse studies described in this work were carried out in accordance with the principles of the Guide for the Care and Use of Animals and were approved by the Institutional Animal Care and Use Committee of Dartmouth College, NH, USA (protocol 2012).