InVivoMAb anti-mouse VISTA

Endemic typhus caused by Rickettsia (R.) typhi is an emerging febrile disease that can be fatal due to multiple organ pathology. Here we analyzed the requirements for protection against R. typhi by T cells in the CB17 SCID model of infection. BALB/c wild-type mice generate CD4+ TH1 and cytotoxic CD8+ T cells both of which are sporadically reactivated in persistent infection. Either adoptively transferred CD8+ or CD4+ T cells protected R. typhi-infected CB17 SCID mice from death and provided long-term control. CD8+ T cells lacking either IFNγ or Perforin were still protective, demonstrating that the cytotoxic function of CD8+ T cells is not essential for protection. Immune wild-type CD4+ T cells produced high amounts of IFNγ, induced the release of nitric oxide in R. typhi-infected macrophages and inhibited bacterial growth in vitro via IFNγ and TNFα. However, adoptive transfer of CD4+IFNγ-/- T cells still protected 30-90% of R. typhi-infected CB17 SCID mice. These cells acquired a TH17 phenotype, producing high amounts of IL-17A and IL-22 in addition to TNFα, and inhibited bacterial growth in vitro. Surprisingly, the neutralization of either TNFα or IL-17A in CD4+IFNγ-/- T cell recipient mice did not alter bacterial elimination by these cells in vivo, led to faster recovery and enhanced survival compared to isotype-treated animals. Thus, collectively these data show that although CD4+ TH1 cells are clearly efficient in protection against R. typhi, CD4+ TH17 cells are similarly protective if the harmful effects of combined production of TNFα and IL-17A can be inhibited.

Immune checkpoint V-domain Ig suppressor of T-cell activation (VISTA) exhibits distinct expression patterns and non-redundant immunoregulatory mechanisms in different autoimmune diseases. This study aims to investigate the expression of VISTA in patients with autoimmune uveitis and experimental autoimmune uveitis (EAU) mice, and explore its clinical significance and preliminary mechanisms in disease development. We found that VISTA expression on 12 subsets of peripheral blood immune cells was lower in autoimmune uveitis patients than in healthy volunteers, especially on neutrophils. The expression of neutrophil VISTA in active uveitis patients markedly increased when intraocular inflammation was ameliorated, indicating a significant correlation with disease activity. In vitro treatment of neutrophils from autoimmune uveitis patients with a VISTA antagonist markedly aggravated cell activation and neutrophil extracellular traps formation, whereas a VISTA agonist produced the opposite effect. Moreover, VISTA was constitutively expressed in the outer segments of retina in healthy mice, and decreased in EAU mice, reaching the lowest level of expression when the disease was at a peak stage. Taken together, this study investigates the relationship between neutrophil VISTA and the development of autoimmune uveitis, and provides new insights into the mechanisms and therapeutic roles of VISTA in autoimmune diseases.

V-domain immunoglobulin suppressor of T-cell activation (VISTA, VSIR) is a key immune checkpoint receptor under investigation as a target for cancer immunotherapy. However, a better understanding of the signaling mechanisms of VISTA is needed to optimize the therapeutic potential. In this study, we identified a conserved four amino acid (NPGF) intracellular motif in VISTA that suppresses cell proliferation by constraining cell-intrinsic growth receptor signaling. A class of triple-negative breast cancers (TNBC) with high VISTA expression and low proliferative index was identified and characterized. The NPGF motif bound to the adapter protein NUMB and recruited Rab11 endosomal recycling machinery. The NPGF motif sequestered NUMB at endosomes, which interfered with EGFR trafficking and signaling to suppress tumor growth. These tumor-suppressive effects did not require canonical VISTA ligands or a functioning immune system. Mutation of the VISTA NPGF domain reverted VISTA-induced growth suppression in multiple breast cancer mouse models. The NPGF motif was also required for response of VISTA+ TNBCs to VISTA-blocking antibodies. These results define a mechanism by which VISTA recruits adapter proteins to control malignant epithelial cell growth and signaling. They also define distinct intracellular residues that are critical for response to therapeutic antibodies that could be exploited to improve immunotherapy.

Targeted therapies have improved survival for lung adenocarcinoma patients. However, similar advances are lacking for lung squamous carcinoma (LUSC). Advances in immunotherapy have shown some promise, but the overall response rate remains low in LUSC. Here, we demonstrate that the mTORC2 signaling pathway represents an actionable target in LUSC to improve anti-tumor immune responses. We show that genetic alterations affecting the mTORC2 pathway are common among patients with LUSC tumors, and targeting mTORC2 reduces LUSC tumor growth in mouse models. Transcriptomics reveal that mTORC2-deficient LUSC cells exhibit reduced expression of glycolytic and hypoxia-related genes. In agreement, loss of mTORC2 signaling decreases lactate levels in tumor-interstitial fluid, creating reduced acidity within the tumor microenvironment. Interestingly, mTORC2-deficient LUSC cells also exhibited reduced expression of the pH-sensitive VISTA ligand PSGL-1 in a HIF-2α dependent mechanism. LUSC patients, but not those with LUAD, display a positive correlation in expression between HIF-2α and PSGL-1, suggesting a distinct association among mTORC2, HIF-2α, and immune responses in LUSC. Indeed, mTORC2 loss-of-function enhanced CD8+ T cell activation in tumors, while use of anti-VISTA immunotherapy reduced LUSC tumor burden only in the presence of intact mTORC2 signaling. Collectively, these data describe an important role of mTORC2 signaling in LUSC tumors and demonstrate the therapeutic potential of targeting the mTORC2/PSGL-1/VISTA axis in patients that are non-responsive to current therapies.