Rapid and large scale plant-based production of catalytic nerve agent bioscavenge

催化神经毒剂生物清除剂的快速大规模植物基生产

• 批准号: 7689885
• 负责人: Tsafrir Shlomo Mor
• 金额: $ 51.35万
• 依托单位: U.S. ARMY MEDICAL RESEARCH INST CHEM DEF
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7689885
• 关键词: Acetylcholine; Address; Animal Experimentation; Animals; Arylesterase; Binding; Biochemical; Biological Assay; Biomanufacturing; Biomass; Blood Banks; Butyrylcholinesterase; Chemical Warfare; Cholinesterases; Clinical; Clinical Trials; Complex; Containment; Crystallography; Data; Data Set; Department of Defense; Development; Drug Kinetics; Endoplasmic Reticulum; Enzyme-Linked Immunosorbent Assay; Enzymes; Evaluation; Evolution; Fertility; Generations; Genes; Goals; Health; Human; Hydrolysis; In Vitro; Individual; Intervention; Lead; Mammals; Measures; Medical; Military Personnel; Modeling; Molecular; Mutagenesis; None or Not Applicable; Organophosphates; Paraoxonase 1; Pathway interactions; Performance; Pharmacologic Substance; Plant Leaves; Plant Viruses; Plants; Plasma; Polysaccharides; Production; Prophylactic treatment; Protein Engineering; Proteins; Protocols documentation; Recombinant Proteins; Recombinants; Research; Scheme; Security; Seeds; Services; Source; System; Technology; Testing; Tobacco; Toxin; Transfection; Transgenic Organisms; Transgenic Plants; Translating; Variant; Viral; Western Blotting; Work; base; bioscavenger; cost; day; efficacy evaluation; esterase; gene synthesis; improved; milligram; nerve agent; novel; pre-clinical; pre-clinical research; protein purification; research study; scale up; success; systems research; trait; vector

Bioscavenging of organophosphate (OP) by human proteins is emerging as a promising medical intervention for prophylaxis and post-exposure treatment against chemical warfare nerve agents. The best-studied bioscavengers (BSCs) to date, meeting considerable success in pre-clinical research, are human cholinesterases (ChEs). However, ChEs, which are highly efficient in binding and sequestering OPs, are also inactivated by the toxins and therefore administration of large amounts of protein is necessary for full protection, raising the question of the practicality of this approach. However the development of a new generation of BSCs that can catalytically degrade OPs may address this concern. The proposed effort offers a novel means to biomanufacture recombinant catalytic BSCs based on the human proteins butyrylcholinesterase and paraoxonase 1. Through efforts of other projects participating in the Center, the capacity of these proteins to hydrolyze OPs will be improved by subjecting their genes to either random in vitro evolution or rational mutagenesis. Concomitantly to the protein engineering research, the ASU team in this Project 5 will utilize tobacco plants to first produce research-scale quantities of BSC enzyme, but also provide a sustainable large-scale production platform. At present, purification of BChE from outdated blood-banked human plasma enables research on how bioscavenger therapy can be used. But this stop-gap measure cannot be practically implemented to allow for a sustained supply of that enzyme, and is not applicable for the new recombinant catalytic BSCs that will be developed by the Center. If it will be decided that such scavengers should be a component of the medical arsenal of the Departments of Defense, Homeland Security and Health and Human Services, it is vital that a reliable, safe, non supply-limited and inexpensive source of such enzymes be identified and developed. The primary significance of the proposed work is that it translates basic studies on the first generation ChE-based BSCs into novel biomanufacturing technology leading to clinical product development of the second generation catalytic BSCs.

利用人体蛋白质清除有机磷农药是一种很有前途的医学干预手段 用于预防和接触化学战神经毒剂后的治疗。研究最多的 迄今为止，在临床前研究中取得了相当大的成功的生物清除剂（BSC）是人类的 胆碱酯酶（ChE）。然而，在结合和螯合OP方面非常有效的ChE也是 被毒素灭活，因此需要给予大量的蛋白质， 保护，提出了这种做法的实用性问题。然而，新的发展 可以催化降解OP的BSC的产生可以解决这个问题。 所提出的努力提供了一种新的手段，以生物制造重组催化BSC的基础上， 人蛋白丁酰胆碱酯酶和对氧磷酶1。通过参与的其他项目的努力， 在该中心，这些蛋白质水解OP的能力将通过使它们的基因受到 随机的体外进化或合理的诱变。伴随着蛋白质工程研究， 亚利桑那州立大学的研究小组将利用烟草植物首先生产研究规模的BSC 酶，而且还提供了一个可持续的大规模生产平台。目前，BChE的纯化， 过时的血库人类血浆使得研究如何使用生物清除剂疗法成为可能。但这 实际上不能实施权宜之计以允许酶持续供应， 不适用于本中心拟开发的新型重组催化生物活性干细胞。这是否是 决定这种食腐动物应该成为国防部医疗武器库的一个组成部分， 国土安全部和卫生与公众服务部，提供可靠、安全、不受供应限制且 鉴定和开发这种酶的廉价来源。建议的主要意义 这项工作是将第一代基于胆碱酯酶的生物干细胞的基础研究转化为新型生物制造 该技术可用于第二代催化BSC的临床产品开发。