An Enzyme-Based Antidote for Acute Nicotine Toxicity

一种基于酶的急性尼古丁中毒解毒剂

• 批准号: 10790758
• 负责人: Kim Janda
• 金额: $ 27.15万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-30 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10790758
• 关键词: 5 year old; Accident and Emergency department; Accidents; Acetylcholine; Activated Charcoal; Acute; Address; Adrenergic Antagonists; Adult; Affinity; Agonist; Animal Model; Antibodies; Antidotes; Attenuated; Autonomic ganglion; Benzodiazepines; Binding; Biodistribution; Biological; Biological Assay; Blood; Blood Pressure; Bradycardia; Brain; Brain region; Buffers; Candy; Carbon; Cessation of life; Child; Childhood; Cholinergic Receptors; Chronic; Clinical Research; Clinical Trials; Coma; Complex; Consumption; Directed Molecular Evolution; Dose; Drug Kinetics; Dyspnea; Electronic cigarette; Engineering; Enzymes; Evolution; Exposure to; Failure; Fatal Outcome; Fruit; Gastric Lavage; Gene Library; Generations; Genes; Genetic Recombination; Goals; Half-Life; Hand; Heart Rate; Hour; Hypotension; Immunologics; In Situ; Information Systems; Ingestion; Intervention; Libraries; Marketing; Measures; Mechanical ventilation; Menthol; Metabolism; Modeling; Monoclonal Antibodies; Muscle; Mutagenesis; Neuromuscular Junction; Neurons; Nicotine; Nicotinic Receptors; Nitrogen; Oral Ingestion; Overdose; Oxidoreductase; Patients; Peripheral; Pharmaceutical Preparations; Pharmacodynamics; Plethysmography; Poison; Poisoning; Population; Positioning Attribute; Pseudomonas putida; Publishing; Reporting; Respiratory Failure; Risk; Rodent; Rodent Model; Sales; Seizures; Serum; Signs and Symptoms; Site; Societies; Source; Structure; Supportive care; System; Therapeutic; Tobacco; Toxic effect; Treatment Protocols; Uncertainty; Vaccines; Variant; Vulnerable Populations; aspirate; cardiovascular collapse; cholinergic; combustible cigarette; efficacy evaluation; electronic cigarette use; electronic liquid; experimental study; improved; liquid nicotine; mouse model; mutant; nerve agent; neural; nicotine cessation; nicotine exposure; nicotine oxidase; nicotine use; nicotine vaccine; organophosphate poisoning; pharmacologic; placebo group; pre-clinical; receptor; respiratory; response; smoking abstinence; smoking cessation; therapeutic enzyme; thermostability; tobacco products

Project Summary/Abstract. Electronic cigarettes are gaining popularity as alternative to traditional cigarettes, with sales increasing from $283 million in 2012 to $2.5 billion in 2018 in the US. The global e-cigarette market is estimated to reach $24.2 billion by 2024. E-cigarette solutions, also known as e-liquids, are highly variable with enticing flavors, such as tobacco, menthol, fruit, candy, and dessert. Liquid nicotine concentrations vary in these products from 0 to 100 mg/ml according to an FDA study. Although nicotine toxicity in adults is rare with an estimated lethal dose between 60 and 500 mg (0.8-6.7 mg/kg), flavored e-liquids are increasingly being ingested orally, putting children at risk for exposure to high concentrations of nicotine. Indeed, nicotine toxicity in children under 5 years of age can occur with consumption of as little as a teaspoon of liquid nicotine. The unforeseen consequences of e-cigarettes, with respect to nicotine poisoning in children, presents an unmet need to counteract the harmful and potentially fatal outcomes that may occur among this vulnerable population. It is the pediatric population we plan to address in this proposal, as a high potential of accidental ingestion of liquid nicotine from e-cigarettes exists. Despite the increase in nicotine-related poisonings reported, there is no treatment for acute nicotine toxicity. Current treatment regimens for nicotine poisoning range from supportive care, to activated charcoal, to respiratory support with mechanical ventilation. An alternative means of altering the toxicity of nicotine poisoning could come via simple sequestering of the drug. Antibodies to nicotine have been prepared as a means to block the pharmacological effects of this drug. To date, vaccines for smoking cessation have shown promise in preclinical animal models; however, in clinical studies, these vaccines failed to measure significant differences in smoking abstinence between the intervention and placebo groups. Thus, the likelihood of an antibody attenuating an acute dose of nicotine is doubtful. What is needed is a sufficient pharmacokinetic (PK) biologic with the capacity to not just sequester nicotine but also increase its metabolism. We envision a biologic able to catabolize nicotine rather than simply sequestering the drug would have the potential to treat acute nicotine poisoning. The proposal at hand details a bacterial strain, Pseudomonas putida, which has evolved to use nicotine as its sole source of carbon and nitrogen. From this bacterial strain, we will examine a first-in-class enzyme, a nicotine oxidoreductase termed NicA2, as a means to treat nicotine poisoning. Our initial characterization of the enzyme indicates that it could be an excellent candidate for altering nicotine poisoning. However, the successful demonstration of this enzyme reversing nicotine poisoning will require several experimental undertakings including: (1) Evaluating the efficacy of NicA2 to attenuate acute nicotine toxicity in rodent models including plethysmography, blood/brain distribution and lethality. (2) Directed evolution of NicA2 to increase its catalytic capacity through gene recombination and random mutagenesis.

项目概要/摘要。电子香烟作为传统香烟的替代品越来越受欢迎， 在美国的销售额从2012年的2.83亿美元增加到2018年的25亿美元。全球电子烟市场 预计到2024年将达到242亿美元。电子烟解决方案，也称为电子液体，具有高度可变性， 诱人的口味，如烟草、薄荷、水果、糖果和甜点。液体尼古丁浓度在这些 根据FDA的研究，产品从0到100 mg/ml。虽然成人的尼古丁毒性是罕见的， 估计致死剂量在60至500毫克（0.8-6.7毫克/千克）之间，调味电子液体越来越多地被摄入 口服，使儿童处于暴露于高浓度尼古丁的风险中。事实上，儿童的尼古丁毒性 5岁以下的儿童只要摄入一茶匙的液体尼古丁就可能发生这种情况。不可预见的 电子烟对儿童尼古丁中毒的后果，提出了一个未得到满足的需求， 消除这一弱势群体中可能出现的有害和潜在致命后果。是 我们计划在本提案中针对儿科人群，因为意外摄入液体的可能性很高 电子烟中存在尼古丁。尽管报告的与尼古丁有关的中毒事件有所增加， 治疗急性尼古丁中毒。目前尼古丁中毒的治疗方案从支持性的 护理，活性炭，呼吸支持与机械通气。另一种改变的方法 尼古丁中毒的毒性可能来自于药物的简单隔离。尼古丁的抗体 作为一种手段来阻止这种药物的药理作用。到目前为止， 在临床前动物模型中显示出戒烟前景;然而，在临床研究中，这些疫苗失败了， 测量干预组和安慰剂组之间戒烟的显著差异。因此，在本发明中， 抗体减弱尼古丁急性剂量的可能性值得怀疑。我们需要的是一个足够的 这是一种药代动力学（PK）生物制剂，不仅能够螯合尼古丁，还能增加其代谢。 我们设想一种能够分解尼古丁而不是简单地隔离药物的生物制剂， 治疗急性尼古丁中毒的潜力。手头的提案详细介绍了一种细菌菌株，恶臭假单胞菌， 它已经进化到使用尼古丁作为其唯一的碳和氮来源。从这个菌株中，我们将 研究一种一流的酶，一种名为NicA 2的尼古丁氧化还原酶，作为治疗尼古丁中毒的一种手段。 我们对这种酶的初步鉴定表明，它可能是改变尼古丁的一个很好的候选者。 中毒然而，成功证明这种酶逆转尼古丁中毒将需要 几项实验工作包括：（1）评估NicA 2减弱急性尼古丁的功效 啮齿动物模型中的毒性，包括体积描记法、血液/脑分布和致死率。(2)定向进化 通过基因重组和随机诱变提高其催化能力。