Monoclonal Antibodies and Nanobodies to the Immune Checkpoint ligand VISTA
The V-domain immunoglobulin suppressor of T-cell activation know as VISTA is a newly discovered immune checkpoint molecule regulating T-cell activation. It is widely expressed within the hematopoietic compartment, with the highest expression found on myeloid cells. VISTA acts as both a receptor and a ligand on immune cells. In terms of sequence homology, PD-L1 is the closer homologue of the VISTA extracellular domain. The VISTA pathway however operates through a distinct mechanistic pathway than PD-1:PD-L1 immune axis. VISTA does not bind to itself, nor to PD-1 or PD-L1 and despites the fact that VISTA has been shown to bind to several ligands (VSIG3, PSGL-1 and Syndecan-2), it remains unclear if any of these ligands are true receptors. Activating the VISTA pathway either with VISTA as a ligand or with agonistic mAbs to VISTA blocks T cell activation and proliferation events. As such, we and others have shown that these agents are immunosuppressive in many murine models of inflammatory diseases. In order to further study the mechanism of action of VISTA, our laboratory has recently generated nanobodies and mouse mAbs to the human VISTA IgV domain with most of them behaving as agonistic mAbs. We have also generated rat mAbs that recognize both murine and human VISTA IgV domain. These reagents are available for collaboration (1).
Designing a bivalent protein construct incorporating the minimal V domain of the Poliovirus Receptor that serves as a potent immunosuppressant 2
PVR (poliovirus receptor) functions as a ligand that signals through TIGIT and CD96 to induce suppression of T-cell and NK-cell responses. In contrast, PVR also binds to CD226, resulting in a co-stimulatory signal. To date, TIGIT antibody antagonists have been developed to restore immune functions and allow PVR to signal though CD226 in the context of cancer immunotherapy. Due to PVR receptor heterogeneity, agonizing either of these pathways with a recombinant form of the PVR extracellular domain represents a therapeutic strategy for either immunosuppression or activation. We have now developed a minimal murine PVR-Fc fusion construct, consisting of only the IgV domain of PVR (vdPVR-Fc), and assessed its ability to dampen inflammatory responses in a murine model of psoriasis. vdPVR-Fc bound to TIGIT, CD96 and CD226 with similar low nanomolar affinities as defined by surface plasmon resonance. vdPVR-Fc was also able to suppress the in-vitro proliferation of murine CD4+ and CD8+ T-cells in mixed splenocyte cultures. Importantly, vdPVR-Fc delayed the onset, and reduced inflammatory responses (scaling and thickness) in a murine model of psoriasis. Collectively, our results suggest that the minimal IgV domain of PVR is sufficient to dampen immune responses in-vitro and attenuate symptoms of psoriasis in-vivo (2).
What are Bispecific Proteins ?
Bispecific proteins have two binding domains for binding two distinct targets. These proteins have versatile properties that make them advantageous for use as tools in biochemical techniques or as therapeutics. Our current research includes designing novel soluble reagents for improving in vitro techniques and to design and assess novel bispecific therapeutics with immunomodulatory properties.
T-CEP (T-Cell Expansion Protein) constructs
T-CEP (T-Cell Expansion Protein) combines a single chain variable fragment (ScFV) against CD28 with a ScFV against CD3 in one small soluble protein to produce a highly potent small ex vivo T-cell activator and expander. This technology improves on currently available soluble method for T-cell expansion, and has potential for application in the field of adoptive cell therapy, where efficient expansion of effective T-cells is needed (3).
Figure 1. Side view of CD28-CD3 ScFV bispecific construct ribbon structure as predicted using Deepmind Alphafold 2.0 under a Google Colab iteration [CD28 ScFV, CD3 ScFV, scFv heavy and light chain, general linker, 6x His Tag] (https://colab.research.google.com/github/sokrypton/ColabFold/blob/main/beta/AlphaFold2_advanced.ipynb#scrollTo=KAZj6CBZTkJM)
Bispecific proteins for tissue-specific targeting and accumulation
The use of the bispecific protein format for tissue-specific accumulation of therapeutic cargo is also being explored. By selecting two appropriate domains, we are studying the effectiveness of delivering immunomodulatory proteins to a specific antigen as means of potentiating the therapeutic effect and reducing systemic side effects in the treatment of inflammatory diseases.
Selected publications from our group:
1. Ma YV, Sparkes A, Romão E, Saha S, Gariépy J. (2021) Agonistic nanobodies and antibodies to human VISTA. MAbs. 13(1):2003281
2. Saha S, Sparkes A, Matus EI, Lee P, Gariépy J. (2023) The IgV domain of the poliovirus receptor alone is immunosuppressive and binds to its receptors with comparable affinity. Sci Rep. 13(1):4609
3. Matus EI, Sparkes A, Gariépy J (2020) A soluble activator that favors the ex vivo expansion of CD8+CD27+ T cells. JCI Insight. 5(22):e141293
Last updated on April-4-2023