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）是当今世界上用于治疗疼痛和发热的最常见药物，也是美国急性肝衰竭的主要原因。APAP毒性的初始阶段已得到充分表征，涉及母体药物生物转化为化学反应性代谢物N-乙酰基-对苯醌亚胺（NAPQI），其共价结合细胞蛋白。NAPQI通过与肝谷胱甘肽（GSH）上的半胱氨酰巯基结合而解毒。在毒性APAP暴露中，GSH储备耗尽，增加了与细胞蛋白上的半胱氨酰硫醇结合的NAPQI的量，产生各种APAP-蛋白加合物。该提案的牵头单位率先测量了总APAP-蛋白加合物（APAP-ADDUCTS）作为APAP毒性的临床标志物，并在急性APAP过量、APAP相关急性肝功能衰竭的儿童和成人中测试了该生物标志物，最近还在接受推荐剂量APAP的患者中进行了测试。在接受推荐剂量的APAP的成人中，检测到低水平的APAP-ADDUCTS，并且在这些患者中发现较高的APAP- ADDUCT水平与较高的转氨酶升高值水平相关。根据我们最近的数据，将检验以下假设：在APAP暴露的儿童和青少年中，（1）在接受治疗暴露的住院儿童中将检测到APAP-加合物，（2）存在独特和特异性APAP加合物蛋白，并且作为APAP暴露量的函数而不同，（3）独特的蛋白加合物将与APAP毒性的既定指标和协变量相对应。使用最先进的、以内收物为重点的蛋白质组学方法，以下建议将鉴定和评价检查接受治疗剂量APAP的儿童/青少年和接受过量APAP的儿童/青少年中APAP毒性的特定“第二代”生物标志物。参与儿科药理学研究单位网络（PPRU;美国国立儿童健康与人类发展研究院）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。特异性APAP蛋白加合物的鉴定和这些特异性加合物相对于新描述的APAP毒性代谢组学标志物和已建立的肝毒性指数的检查将为改善未来APAP在儿童和青少年中的风险和安全性评估奠定基础。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热的最常用药物。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，最近已被证明会导致一些患者轻度肝损伤。以前的研究表明，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肝损伤的标志。该提案将使用先进的蛋白质鉴定方法来鉴定被对乙酰氨基酚改变的确切蛋白质。研究样本将从接受推荐剂量对乙酰氨基酚的儿童和青少年以及对乙酰氨基酚过量的儿童和青少年中获得。将对这些样本进行复杂的蛋白质鉴定研究，并将研究这些信息与肝损伤的常见临床试验的关系。这些新知识将增加我们对乙酰氨基酚如何导致肝损伤以及一些患者如何可能面临更大的肝损伤风险的理解。

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.