Novel Therapeutic Approaches for Narcotic Overdose

麻醉药物过量的新治疗方法

• 批准号: 7479832
• 负责人: PHILIP M POTTER
• 金额: $ 27.39万
• 依托单位: ST. JUDE CHILDREN'S RESEARCH HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-09-30 至 2010-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7479832
• 关键词: 6-O-monoacetylmorphine; Accident and Emergency department; Acids; Active Sites; Acute; Alcohols; Amino Acid Sequence; Amino Acids; Animal Model; Animals; Area; Arrhythmia; Biochemical; Blood; Carboxylic Ester Hydrolases; Cocaine; Cocaine Abuse; Complex; Development; Engineering; Enzymes; Esters; Ethanol; Exposure to; Goals; Heroin; Hospitals; Human; Human Activities; Hydrolase; Hydrolysis; Injection of therapeutic agent; Liver; Measures; Metabolism; Methods; Morphine; Mus; Mutation; Naloxone; Narcotics; Numbers; Overdose; Patients; Pharmaceutical Preparations; Plasma; Principal Investigator; Proteins; Reaction; Reporting; Seizures; Site; Stream; Structure; Symptoms; Testing; United States; analog; base; carboxylesterase; cocaethylene; design; ecgonine; esterase; homatropine; human carboxylesterase 1; improved; methyl group; mutant; novel; novel therapeutics; programs; purge; urinary

DESCRIPTION (provided by applicant): Cocaine and heroin are metabolized by carboxylesterases (CE) present in the liver. With cocaine, the methyl group is removed by human carboxylesterase 1 (hCE1) to produce benzyl ecgonine, the major inactive metabolite of the drug. With heroin, both methyl esters are sequentially hydrolyzed by the same enzyme to yield 6-monoacetylmorphine and subsequently, morphine. We have recently determined the x-ray crystal structure of hCE1, in complex with the cocaine and heroin analogs, homatropine and naloxone, respectively. Using this information, we will engineer mutations within the hCE1 amino acid sequence to improve its ability to hydrolyze cocaine. These enzymes will be tested in an esterase-deficient animal model to determine whether they can be used to purge cocaine from the blood stream. Additionally, we will determine the ability of these mutants to transesterifiy cocaine in the presence of ethanol to form cocaethylene, and measure the hydrolysis of this compound by different hCE1 proteins. The Specific Aims of this application are therefore to: 1) generate hCE1 mutants with improved catalytic efficiency for cocaine hydrolysis; 2) determine the amino acid residues involved in the transesterification reaction of cocaine to cocaethylene by hCE1; 3) elucidate crystal structures of engineered forms of hCE1 in complexes with cocaine and cocaine analogues; and 4) assess the impact of mutant forms of hCE1 on plasma esterase-deficient mice exposed to cocaine. Overall, these studies should allow development of novel therapeutic approaches for the treatment of cocaine overdose.

描述（由申请人提供）：可卡因和海洛因被肝脏中存在的羧酸酯酶（CE）代谢。使用可卡因，通过人类羧酸酯酶1（HCE1）去除甲基，以产生苯甲烷基，这是该药物的主要非活性代谢产物。使用海洛因，两种甲基酯都被相同的酶顺序水解，以产生6-单乙酰乙酰氨基苯甲吗啡，随后是吗啡。我们最近分别确定了HCE1的X射线晶体结构，分别与可卡因和海洛因类似物，霍莫酮和纳洛酮分别确定了X射线晶体结构。使用此信息，我们将在HCE1氨基酸序列中设计突变，以提高其水解可卡因的能力。这些酶将在缺乏酯酶的动物模型中进行测试，以确定它们是否可以从血液中清除可卡因。此外，我们将确定这些突变体在乙醇存在下形成可可乙烯的能力，并测量不同HCE1蛋白对该化合物的水解。 因此，本应用的具体目的是：1）生成可卡因水解催化效率提高的HCE1突变体； 2）确定HCE1与可卡因对可卡因的酯交换反应有关的氨基酸残基； 3）在与可卡因和可卡因类似物的复合物中阐明HCE1工程形式的晶体结构； 4）评估HCE1突变形式对暴露于可卡因的血浆酯酶缺陷小鼠的影响。总体而言，这些研究应允许开发新的治疗方法来治疗可卡因过量。