Rational Structure-Based Design of Broad Neutralizing Humanized svMP mAbs

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PROJECT SUMMARY In this R21 award, we aim to develop a novel and broad neutralizing human monoclonal antibody for treating snakebite envenoming by rational structure-based design in order to produce a more effective and safer next generation antivenom. Snake envenomation is a serious global public health concern and ranked on the Wor ld Health Organization’s list of neglected tropical diseases, killing on average 125,000 people per year and leaving another 400,000 permanently disabled. The majority of snake envenomation in the US, inflicted by members of the snake family Viperidae, causes local tissue damage (such as myonecrosis, blisters, and local inflammation and pain) and systemic effects, including hemorrhage and coagulopathies which can lead to shock, renal failure and death. Snake venom metalloproteinases are major causative agents for spontaneous systemic bleeding and coagulopathies. Current antivenoms, produced by immunization of domestic animals, have limited efficacy in the prevention of both local and systemic effects of Viperidae envenomation as well as an associated risk of hypersensitivity reactions. Our long-term goal is to develop novel, effective humanized antivenom therapeutics for Viperidae envenomation. The objective of this project is to test the hypothesis that camelid-inspired inhibitory paratope synthetic human antibodies targeted to the active site of medically-relevant viperid venom metalloproteinases (svMPs) can provide broad antivenom protection without cross-reaction with human metalloproteinases and without the risk of hypersensitivity. This objective will be addressed through our established collaboration of complementary expertise between the snake venom toxinology team at National Natural Toxins Research Center (NNTRC) and the antibody discovery team at University of California Riverside (UCR). To test our hypothesis, we will address the following three Specific Aims. Aim 1: Qualitative and Quantitative Characterization of the hemorrhagic activity of viperid svMPs (Galan), Aim 2: Discovery of Broadly Neutralizing svMP-Specific Human mAbs (Ge). Aim 3: Evaluation of the antivenom efficacy of svMP inhibitory mAbs in vitro (Ge) and in vivo (Sanchez). The proposed research is significant because it will advance our understanding of the hemorrhagic aspects caused by snake envenomation at biochemical/cellular levels and develop effective humanized mAb antivenoms, which will be directly translatable for therapeutic use. The novelties of our project are (1) development and application of a novel Hemorrhage Score system to characterize svMPs; (2) isolation of humanized svMP-specific antivenom mAbs from libraries carrying novel convex paratopes; (3) development groundbreaking functional (rather than binding-based) HTS for facile discovery of mAbs inhibiting hemorrhagic snake toxins; and (4) potentially shifting the conventional antivenom production into specific neutralizing humanized mAb therapeutics.