Identification of new mechanistic biomarkers of adverse responses to acetaminophe

描述（由申请人提供）：对乙酰氨基酚（APAP）是当今世界上最常用的用于治疗疼痛和发烧的药物，也是美国急性肝衰竭的主要原因。APAP毒性的初始阶段已被很好地表征，涉及母体药物向化学反应性代谢物n -乙酰基-对苯醌亚胺（NAPQI）的生物转化，NAPQI与细胞蛋白共价结合。NAPQI通过与肝谷胱甘肽（GSH）上的半胱氨酸硫醇结合来解毒。在有毒的APAP暴露中，谷胱甘肽储备被耗尽，增加了与细胞蛋白上的半胱氨酸巯基结合的NAPQI的数量，产生各种APAP蛋白加合物。该提案的先导点率先测量了APAP-蛋白加合物总量（APAP- adducs）作为APAP毒性的临床标志物，并在急性APAP过量、APAP相关急性肝衰竭的儿童和成人以及最近接受推荐剂量APAP的患者中测试了该生物标志物。在接受推荐剂量APAP的成人中，检测到低水平的APAP- adducs，并发现这些患者中较高的APAP- adducs水平和较高的转氨酶值水平升高存在关联。根据我们最近的数据，我们将对以下假设进行检验：在接受APAP暴露的儿童和青少年中，(1)在接受治疗性暴露的住院儿童中检测到APAP加合物，(2)存在独特和特异性的APAP加合物蛋白，并且随着APAP暴露程度的不同而不同，(3)独特的蛋白质加合物将与APAP毒性的既定测量和协变量相对应。使用最先进的，以加合物为中心的蛋白质组学方法，以下建议将识别和评估接受治疗剂量APAP的儿童/青少年和接受过量APAP的儿童/青少年中APAP毒性的检查特异性“第二代”生物标志物。参与儿童药理学研究单位网络（PPRU；国家儿童健康与人类发展研究所）的儿科学术中心将协助临床样本收集和分析及药代动力学数据分析。鉴定特异性APAP蛋白加合物，并检查这些特异性加合物与新描述的APAP毒性代谢组学标记物和已建立的肝毒性指标的关系，将为今后更好地评估儿童和青少年使用APAP的风险和安全性奠定基础。公共卫生相关性：对乙酰氨基酚是当今世界上用于治疗疼痛和发烧的最常用药物。当使用非常大剂量的对乙酰氨基酚或发生毒性过量时，会导致严重的肝损伤。对乙酰氨基酚，在推荐剂量下，最近已被证明对一些患者造成轻度肝损伤。先前的研究表明，对乙酰氨基酚改变了肝脏中的蛋白质，这些蛋白质的变化可以作为对乙酰氨基酚损伤肝脏的标志。本方案将采用先进的蛋白质鉴定方法来鉴定对乙酰氨基酚改变的确切蛋白质。研究样本将从正在接受对乙酰氨基酚推荐剂量的儿童和青少年以及对乙酰氨基酚过量受害者的儿童和青少年中获取。将对这些样本进行复杂的蛋白质鉴定研究，并将这些信息与肝损伤的常见临床试验联系起来进行研究。这一新知识将增加我们对扑热息痛如何导致肝损伤的理解，以及为什么一些患者可能有更大的肝损伤风险。

DESCRIPTION (provided by applicant): Acetaminophen (APAP) is the most common drug used for the treatment of pain and fever in the world today and is also the leading cause of acute liver failure in the United States. The initial stages of APAP toxicity have been well-characterized and involve the biotransformation of the parent drug to a chemically reactive metabolite N-acetyl-p-benzoquinone imine (NAPQI), which binds covalently to cellular proteins. NAPQI is detoxified by binding to the cysteinyl thiol on hepatic glutathione (GSH). In toxic APAP exposures, GSH reserves are depleted, increasing the amount of NAPQI that binds to cysteinyl thiols on cellular proteins, producing a variety of APAP-protein adducts. The lead site for this proposal pioneered the measurement of total APAP-protein adducts (APAP-ADDUCTS) as clinical markers of APAP toxicity and tested this biomarker in children and adults with acute APAP overdose, APAP-related acute liver failure, and recently, in patients receiving recommended doses of APAP. In adults receiving recommended doses of APAP, low levels of APAP-ADDUCTS were detected and an association was found for higher APAP- ADDUCT levels and higher elevated transaminase values levels in these patients. Based on our recent data, the following hypotheses will be tested: In children and adolescents with APAP exposures, (1) APAP-ADDUCTS will be detected in hospitalized children receiving therapeutic exposures, (2) unique and specific APAP adduct proteins exist and differ as a function of the magnitude of APAP exposure and, (3) unique protein adducts will correspond with established measures and co- variates of APAP toxicity. Using state-of-the-art, adduct-focused proteomic approaches, the following proposal will identify and evaluate examine specific "second generation" biomarkers of APAP toxicity in children/adolescents receiving therapeutic doses of APAP and in children/adolescents that have received overdoses of APAP. Pediatric academic centers participating in the Network of Pediatric Pharmacology Research Units (PPRU; National Institutes of Child Health and Human Development) will assist with clinical sample collection and analytical and pharmacokinetic data analysis. Identification of specific APAP protein adducts and examination of these specific adducts relative to newly described metabolomic markers of APAP toxicity and established indices of liver toxicity will lay the foundation for improved future assessments of risk and safety for APAP in children and adolescents. PUBLIC HEALTH RELEVANCE: Acetaminophen is the most common drug used in the world today for the treatment of pain and fever. When very large doses of acetaminophen are used or when a toxic overdose occurs, severe liver injury results. Acetaminophen, at recommended doses, has recently been shown to cause mild liver injury in some patients. Previous studies have shown that proteins in the liver are changed by acetaminophen and these protein changes can serve as markers of liver injury from acetaminophen. This proposal will use advanced protein identification methods to identify the exact proteins that are changed by acetaminophen. Study samples will be obtained from children and adolescents that are receiving recommended doses of acetaminophen and from children and adolescents that are victims of acetaminophen overdoses. Sophisticated protein identification studies will be performed on these samples and this information will be studied in relationship to common clinical tests for liver injury. This new knowledge will increase our understanding of how acetaminophen may cause liver injury and how some patients may be at greater risk for liver injury.