Bioactivation of PBDEs by Human Cytochrome P-450

描述(申请人提供)：在过去十年中，动物和人体研究都支持多溴二苯醚(PBDE)阻燃剂与神经行为/神经发育障碍之间的联系，特别是在宫内和出生后暴露。最近，人们发现多溴二苯醚的羟化代谢物(OH-PBDEs)在人体血清中的累积水平与亲本多溴二苯醚的水平相似，有时甚至更高。机制研究表明，OH-PBDEs通常比母体化合物更有效，并通过直接神经毒性(涉及钙信号的失调)和/或间接通过改变甲状腺功能而导致神经发育障碍，从而加强了这一发现的意义。总之，这些和其他研究表明，氧化代谢的生物激活大大增加了多溴二苯醚的神经毒性潜力。因此，迫切需要进一步了解多溴二苯醚在人体内的代谢情况。这项应用的总体目标是表征多溴二苯醚在人类体内的酶和同系物特异性代谢，并调查代谢的定性和定量差异，这些差异与关键生物转化酶的遗传变异性有关。我们最近发现，2，2‘，4，4’-四-(BDE-47)是由人细胞色素P-450 2B6(CYP2B6)特异性代谢的，由于调节现象和常见的遗传多态性，已知的肝脏蛋白表达具有高达100倍的变异性。因此，假设除了可变暴露外，多溴二苯醚CYP特定代谢中的遗传可变性有助于多溴二苯醚身体负担的个体间差异和有毒代谢物的形成。以下目标将解决这一假设，并产生关于多溴二苯醚同种特定新陈代谢的关键人类数据。目的1对2，2‘，4，4’-四-(BDE-47)、2，2‘，4，5’-四-(BDE-49)、2，2‘，4，4’，5-五-(BDE-99)和2，2‘，4，4’，6-五-(BDE-100)在人体内的CYP特异性代谢进行定性和定量表征。多溴联苯醚体外代谢的个体间变异性将利用人类肝脏微生物体(其活性水平最高可达100倍)和人类CYP2B6的重组多态变体进行评估。目的2将鉴定和定量母乳和血清中的羟基多溴二苯醚，并评估CYP2B6基因对多溴二苯醚身体负荷的潜在影响，以前发现这些受试者的多溴二苯醚的身体负荷变化超过100倍。拟议的研究最终将更好地为未来的机械学和流行病学研究提供信息，调查多溴二苯醚及其代谢物产生神经发育障碍的可能性。此外，这些研究将有助于确定潜在的遗传生物标记物，这些标记物有助于多溴联苯醚生物激活的个体间差异，并最终导致相对的 个体对这些药物潜在不良影响的易感性。 公共卫生相关性：在过去的十年中，动物和人类研究都支持多溴二苯醚(PBDE)阻燃剂与神经行为/神经发育障碍之间的联系，最近的机制研究表明，氧化代谢的生物激活大大增加了PBDEs的神经毒性潜力。拟议的关于多溴二苯醚的人细胞色素P-450特异性代谢的研究最终将更好地为未来的机制和流行病学研究提供信息，调查多溴二苯醚及其代谢物产生神经发育障碍的可能性。此外，这些研究将有助于确定潜在的遗传生物标记物，这些标记物有助于多溴联苯醚生物激活的个体间差异，并最终提高个体对这些制剂潜在不利影响的相对易感性。

DESCRIPTION (provided by applicant): During the past decade both animal and human studies have supported an association between polybrominated diphenyl ether (PBDE) flame retardants and neurobehavioral / neurodevelopmental disorders, particularly following in utero and postnatal exposure. Recently, hydroxylated metabolites of PBDEs (OH- PBDEs) have been found to accumulate in human serum at levels similar to and in some cases greater than that of the parent PBDEs. The significance of this finding is heightened by mechanistic studies showing that OH-PBDEs are often more potent than parent compounds and contribute substantially to neurodevelopmental disorders via direct neurotoxicity involving dysregulation of Ca2+ signaling and/or indirectly through altered thyroid disruption. Together, these and other studies suggest that bioactivation by oxidative metabolism adds considerably to the neurotoxic potential of PBDEs. Thus, there is a critical need to further our understanding of PBDE metabolism in humans. The overall objectives of this application are to characterize the enzyme- and congener-specific metabolism of PBDEs in humans and investigate qualitative and quantitative differences in metabolism which are related to genetic variability in key biotransforming enzymes. We have recently found that 2,2',4,4'-tetra-(BDE-47) is metabolized specifically by human cytochrome P-450 2B6 (CYP2B6), which is known to exhibit up to 100-fold variability in hepatic protein expression, due to regulatory phenomena and common genetic polymorphisms. Thus, it is hypothesized that in addition to variable exposures, genetic variability in the CYP-specific metabolism of PBDEs contributes to interindividual variability in the body burden of PBDEs and the formation of toxic metabolites. The following aims will address this hypothesis and generate critical human data on the congener-specific metabolism of PBDEs. Aim 1 will conduct a qualitative and quantitative characterization of the human CYP-specific in vitro metabolism of 2,2',4,4'-tetra-(BDE-47), 2,2',4,5'-tetra-(BDE-49), 2,2',4,4',5-penta-(BDE-99), and 2,2',4,4',6- penta-(BDE-100), which are the most abundant congeners in humans, and are susceptible to metabolism that forms potentially toxicologically active metabolites. Interindividua variability in the in vitro metabolism of PBDEs will be assessed utilizing both human liver microsomes, with up to a 100-fold range in the level of CYP2B6 activity, and recombinant polymorphic variants of human CYP2B6. Aim 2 will identify and quantify OH-PBDEs in human milk and serum, and assess the potential impact of CYP2B6 genotype on the body burden of PBDEs which was previously found to vary by over a 100-fold in these subjects. The proposed studies will ultimately better inform future mechanistic and epidemiological studies investigating the potential of PBDEs and their metabolites to produce neurodevelopmental disorders. In addition, these studies will lead to the identification of potential genetic biomarkers that contribute to interindividual variability in the bioactivation of PBDEs and ultimately the relative susceptibility of individuals to potential adverse effects of these agents. PUBLIC HEALTH RELEVANCE: During the past decade both animal and human studies have supported an association between polybrominated diphenyl ether (PBDE) flame retardants and neurobehavioral / neurodevelopmental disorders, and recent mechanistic studies suggest that bioactivation by oxidative metabolism adds considerably to the neurotoxic potential of PBDEs. The proposed studies on the human cytochrome P-450-specific metabolism of PBDEs will ultimately better inform future mechanistic and epidemiological studies investigating the potential of PBDEs and their metabolites to produce neurodevelopmental disorders. In addition, these studies will lead to the identification of potential genetic biomarkers that contribute to interindividual variability in the bioactivation of PBDEs and ultimately the relative susceptibilit of individuals to potential adverse effects of these agents.