Evaluation of Toxicity Score and Precise Snake Venom Analytics for Next Generation Antivenom Development

下一代抗蛇毒血清开发的毒性评分评估和精确蛇毒分析

• 批准号: 10394860
• 负责人: Jacob Anthony Galan
• 金额: $ 13.8万
• 依托单位: TEXAS A&M UNIVERSITY-KINGSVILLE
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-01 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10394860
• 关键词: Address; Antibody Therapy; Antidotes; Antivenins; Biochemistry; Biological; Biological Markers; Biomedical Research; Blood Coagulation Disorders; Blood coagulation; Cells; Cessation of life; Chemistry; Child; Collection; Complex; Crotalid; Data; Development; Dose; Enzymes; Escherichia coli; Escherichia coli Proteins; Evaluation; Faculty; Foundations; Functional disorder; Future; Geographic Locations; Goals; Health; Hemorrhage; Human; Immune response; Individual; Injury; Knowledge; Laboratories; Light; Mass Spectrum Analysis; Medical; Metabolic; Metalloproteases; Methods; Modeling; Molecular; Mus; Necrosis; Pathology; Peptides; Plasma; Plasmids; Play; Procedures; Prognostic Marker; Proteins; Proteomics; Recombinants; Reporting; Research; Research Project Grants; Research Training; Role; STEM program; Schools; Science; Snake Bites; Snake Venoms; Snakes; Structure; Students; Technology; Testing; Texas; Therapeutic; Time; Toxic effect; Toxin; Toxinology; Training; Universities; Venoms; Woman; acute toxicity; base; cDNA Library; career; design; diagnostic biomarker; exosome; experimental study; global health; graduate student; in vivo; innovation; insight; interest; member; men; method development; mouse model; neglected tropical diseases; new technology; next generation; novel; programs; protein expression; protein purification; statistics; stem; tool; tool development; treatment strategy; undergraduate student

ABSTRACT The overall goal of this project is to engage undergraduate and graduate students in innovative studies in molecular toxinology that have the potential to contribute to important advances in our understanding of snake venom pathology and the development of novel antibody-based therapeutics. According to WHO statistics, it is estimated that in 2020 there will be 125,000 new deaths world-wide reported due to snake envenomation. In spite of its importance for snake bite envenomation as a neglected tropical disease and global health issue, very little is known of the mechanism of action of snake venom and intrinsic toxic/lethal contribution. The scientific foundation for our study is that a specific combination of snake venom toxins plays a critical role in snakebites by causing necrosis, coagulopathy, severe hemorrhaging and eventually death. Inasmuch as individual toxins have been evaluated, synergistic mechanism for snake venom pathology are lacking and warranted for antidote development. We hypothesize that precise toxicity evaluation and snake venom analytics of crude/fractionated venom and recombinant toxins will provide a model in which priority can be given to the most toxic venom component(s). To test our hypothesis, we will address (3) Specific Aims: Specific Aim 1: To profile snake venom composition, combinational assessment and intrinsic toxicity. We will purify and characterize snake venom of Crotalid snake species. Characterization will include profiling snake venom to identify the toxins in each fraction and abundance. Students will investigate the mechanism of action different combinations of venom fractions. We will characterize the lethal concentrations/doses of each venom fraction on HUVEC cells, blood coagulation, and will validate the toxicity score for each fraction using an in vivo lethality mouse model. Specific Aim 2: To produce recombinant venom peptides. We will clone these toxins from our collection of cDNA libraries and will generate plasmids and transform competent E.coli cells for protein expression and purification. Development of methods to produce recombinant toxins is essential for antivenom development and investigating toxicity and targets for medically relevant venom peptide. These studies will also be essential to use toxins in solving structures to venom toxins or structures in complexes with known targets. Specific Aim 3: To identify snake venom biomarkers in plasma exosomes. We will explore the “venom-reactome” using novel Evtrap technology and proteomics in discovery-based analysis of snake venom biomarkers from mouse plasma exosomes. Comprehensive snake venom biomarker analysis will shed new light on the pathophysiology, metabolic, biological, cellular and immunological responses to snake venom.

抽象的 该项目的总体目标是让本科生和研究生参与创新研究 分子毒理学有可能为我们对蛇的理解做出重要贡献 根据世界卫生组织的统计数据，毒液病理学和新型抗体疗法的开发。 据估计，2020 年全球将有 125,000 人因蛇中毒而死亡。 尽管蛇咬伤作为一种被忽视的热带疾病和全球健康问题具有重要意义， 人们对蛇毒的作用机制和内在毒性/致死贡献知之甚少。 我们研究的科学基础是蛇毒毒素的特定组合在 蛇咬伤会导致坏死、凝血障碍、严重出血并最终导致死亡。 已对单个毒素进行了评估，但缺乏蛇毒病理学的协同机制 我们捕获了精确的毒性评估和蛇毒。 粗制/分馏毒液和重组毒素的分析将提供一个模型，其中优先考虑 为了检验我们的假设，我们将讨论 (3) 具体的毒液成分。 目标：具体目标 1：分析蛇毒成分、组合评估和内在特征 我们将纯化和表征响尾蛇种类的蛇毒。 分析蛇毒以确定每个组分和丰度的毒素。学生将调查蛇毒。 作用机制我们将表征毒液组分的不同组合的致命性。 每种毒液组分对 HUVEC 细胞、血液凝固的浓度/剂量，并将验证毒性 使用体内致死性小鼠模型对每个部分进行评分 具体目标 2：产生重组体。 我们将从我们的 cDNA 文库中克隆这些毒素并生成质粒。 并转化感受态大肠杆菌细胞以进行蛋白质表达和纯化的方法的开发。 生产重组毒素对于抗蛇毒血清的开发和研究毒性和目标至关重要 这些研究对于使用毒素来解决结构问题也至关重要。 具有已知目标的毒液毒素或复合物的结构。 具体目标 3：识别蛇毒。 我们将利用新型 Evtrap 技术探索“毒液反应组”。 基于发现的小鼠血浆外泌体蛇毒生物标志物分析中的蛋白质组学。 综合蛇毒生物标志物分析将为病理生理学、代谢、 对蛇毒的生物、细胞和免疫反应。

项目成果

Examination of the Efficacy and Cross-Reactivity of a Novel Polyclonal Antibody Targeting the Disintegrin Domain in SVMPs to Neutralize Snake Venom.

• DOI: 10.3390/toxins13040254
• 发表时间: 2021-03-31
• 期刊: Toxins
• 影响因子: 4.2
• 作者: Szteiter SS;Diego IN;Ortegon J;Salinas EM;Cirilo A;Reyes A;Sanchez O;Suntravat M;Salazar E;Sánchez EE;Galan JA
• 通讯作者: Galan JA