Non-Redundant Roles of T Cell Costimulation Pathways in Inflammatory Arthritis Revealed by Dual Blockade of ICOS and CD28 with Acazicolcept (ALPN-101).

Objective: CD28 and inducible T cell costimulator (ICOS) appear to have nonredundant roles in T cell activation and adaptive immunity. We undertook this study to characterize in vitro and in vivo the therapeutic potential of acazicolcept (ALPN-101), an Fc fusion protein of a human variant ICOS ligand (ICOSL) domain designed to inhibit both CD28 and ICOS costimulation, in inflammatory arthritis.

Methods: Acazicolcept was compared in vitro with inhibitors of either the CD28 or ICOS pathways (abatacept and belatacept [CTLA-4Ig], prezalumab [anti-ICOSL monoclonal antibody]) in receptor binding and signaling assays, and in a collagen-induced arthritis (CIA) model.

Povetacicept, an Enhanced Dual APRIL/BAFF Antagonist That Modulates B Lymphocytes and Pathogenic Autoantibodies for the Treatment of Lupus and Other B Cell-Related Autoimmune Diseases.

Objective: Dysregulated APRIL/BAFF signaling is implicated in the pathogenesis of multiple autoimmune diseases, including systemic lupus erythematosus and lupus nephritis. We undertook this study to develop and evaluate a high-affinity APRIL/BAFF antagonist to overcome the clinical limitations of existing B cell inhibitors.

Methods: A variant of TACI-Fc generated by directed evolution showed enhanced binding for both APRIL and BAFF and was designated povetacicept (ALPN-303).

The engineered CD80 variant fusion therapeutic davoceticept combines checkpoint antagonism with conditional CD28 costimulation for anti-tumor immunity.

Despite the recent clinical success of T cell checkpoint inhibition targeting the CTLA-4 and PD-1 pathways, many patients either fail to achieve objective responses or they develop resistance to therapy. In some cases, poor responses to checkpoint blockade have been linked to suboptimal CD28 costimulation and the inability to generate and maintain a productive adaptive anti-tumor immune response. To address this, here we utilize directed evolution to engineer a CD80 IgV domain with increased PD-L1 affinity and fuse this to an immunoglobulin Fc domain, creating a therapeutic (ALPN-202, davoceticept) capable of providing CD28 costimulation in a PD-L1-dependent fashion while also antagonizing PD-1 - PD-L1 and CTLA-4-CD80/CD86 interactions.

Novel Immunomodulatory Proteins Generated via Directed Evolution of Variant IgSF Domains.

Immunoglobulin superfamily member (IgSF) proteins play a significant role in regulating immune responses with surface expression on all immune cell subsets, making the IgSF an attractive family of proteins for therapeutic targeting in human diseases. We have developed a directed evolution platform capable of engineering IgSF domains to increase affinities for cognate ligands and/or introduce binding to non-cognate ligands. Using this scientific platform, ICOSL domains have been derived with enhanced binding to ICOS and with additional high-affinity binding to the non-cognate receptor, CD28.

Structure of FcγRI in complex with Fc reveals the importance of glycan recognition for high-affinity IgG binding.

Fc gamma receptor I (FcγRI) contributes to protective immunity against bacterial infections, but exacerbates certain autoimmune diseases. The sole high-affinity IgG receptor, FcγRI plays a significant role in immunotherapy. To elucidate the molecular mechanism of its high-affinity IgG binding, we determined the crystal structure of the extracellular domains of human FcγRI in complex with the Fc domain of human IgG1.

Generation and characterization of human anti-human IL-21 neutralizing monoclonal antibodies.

Interleukin-21 (IL-21) is a type I four-helical bundle cytokine that exerts a variety of significant effects on many hematopoietic cells, including T and B lymphocytes and natural killer cells. IL-21 is produced predominantly by CD4+ T cells and natural killer T cells and, when aberrantly overexpressed, appears to play important roles in a wide variety of autoimmune disorders. To generate potential therapeutic reagents capable of inhibiting IL-21 for clinical use, we immunized human immunoglobulin transgenic mice with IL-21 and then identified and cloned a panel of human anti-human IL-21 binding monoclonal antibodies.

Vstm3 is a member of the CD28 family and an important modulator of T-cell function.

Members of the CD28 family play important roles in regulating T-cell functions and share a common gene structure profile. We have identified VSTM3 as a protein whose gene structure matches that of the other CD28 family members. This protein (also known as TIGIT and WUCAM) has been previously shown to affect immune responses and is expressed on NK cells, activated and memory T cells, and Tregs.

Generation of a high-affinity Fcgamma receptor by Ig-domain swapping between human CD64A and CD16A.

A recombinant soluble version of the human high-affinity receptor for IgG, rh-FcgammaRIA or CD64A, was expressed in mammalian cells and purified from their conditioned media. As assessed by circular dichroism, size exclusion chromatography and dynamic light scattering, incubation of rh-FcgammaRIA at 37 degrees C resulted in time-dependent formation of soluble aggregates caused by protein unfolding and loss of native structure. Aggregate formation was irreversible, temperature-dependent and was independent of rh-FcgammaRIA concentration.

Recombinant soluble human FcgammaR1A (CD64A) reduces inflammation in murine collagen-induced arthritis.

Binding of immune complexes to cellular FcgammaRs can promote cell activation and inflammation. In previous studies, a recombinant human (rh) soluble FcgammaR, rh-FcgammaRIA (CD64A), was shown to block inflammation in passive transfer models of immune complex-mediated disease. To assess whether rh-FcgammaRIA could block inflammation in a T cell- and B cell-dependent model of immune complex-mediated disease, the efficacy of rh-FcgammaRIA in collagen-induced arthritis was evaluated.