TNT003, a Novel Anti-C1s Monoclonal Antibody, Inhibits Anti-HLA Antibody Induced Complement Deposition.

Abstract

PURPOSE: To determine whether the anti-C1s antibody, TNT003, inhibits complement deposition induced by anti-HLA Class I and II antibodies (Ab). METHODS: HLA reference and transplant patient sera were used as sources of anti-HLA Ab. Complement deposition was evaluated using C1qScreen (OneLambda) and C3d solid phase assays (Immucor/Lifecodes). Briefly, single antigen beads were coated with anti HLA Ab, incubated with normal human serum complement, and C1q or C3d production/deposition were detected by monoclonal PE conjugates and quantified by Luminex analysis. For in vitro studies, either HLA-typed primary human aortic endothelial cells (HAECs) or EBV-immortalized B cells (B cells) were incubated with sera containing cognate anti-HLA Ab and normal human complement serum, and deposition of complement proteins was detected by flow cytometry using monoclonal antibodies. Complement split products produced during deposition reactions were measured by ELISA. TNT003 (True North Therapeutics Inc, CA) was titrated into these experiments in a concentration-dependent manner (0-50 μg/mL). RESULTS: The anti-C1s monoclonal Ab, TNT003, inhibited complement deposition in both solid phase and in vitro assays, and complement split product formation during in vitro assays. TNT003 reduced both anti-HLA Class I and II Ab induced C3d but not C1q deposition on single antigen beads in a concentration-dependent manner. In vitro, increasing concentrations of TNT003 were inversely proportional to levels of complement deposition on ECs that had been treated with anti-HLA Ab; complement split product production was also diminished with TNT003 treatment. Similar results were seen using B cells as a source of HLA, as TNT003 addition to the complement deposition reaction decreased levels of complement proteins on the cell surface, as well as split products in the supernatant of these reactions. CONCLUSIONS: We demonstrate here that TNT003, a novel anti-C1s monoclonal Ab, is capable of inhibiting complement activation, deposition, and split product formation induced by anti-HLA Class I and II Ab in both solid phase and cell based in vitro assays. These data suggest TNT003 may be useful in reducing DSA-induced complement-mediated damage in transplant patients experiencing AMR. DISCLOSURES:Thomas, K.: Grant/Research Support, True North Therapeutics Inc. Valenzuela, N.: Grant/Research Support, True North Therapeutics Inc. Parry, G.: Employee, True North Therapeutics Inc. Panicker, S.: Employee, True North Therapeutics Inc. Reed, E.: Grant/Research Support, Pfizer, True North Therapeutics Inc, Novartis, Immucor.