Laboratory Resource

20世纪70年代末，人们越来越认识到药代动力学在理解药物作用方面的重要性，因此需要分析化学来支持临床研究。Neal Benowitz发起了尼古丁和阿片类药物的药代动力学研究，Reese Jones发起了可卡因药代动力学和药效学研究，主要得到了P50奖的支持。当时，我们的分析实验室由一名化学家、两名技术员和两台气相色谱仪组成。 从那时起，许多需要分析化学支持的项目开始启动，我们的实验室工作人员已经 从3人增加到14人：2名博士研究化学家，9名研究助理和3名实验室助理。主要设备包括两台气相色谱仪、两台台式GC-MS系统、两台HPLC、三台三级四极杆LC-MS/MS系统和两台三级四极杆GC-MS/MS系统。 对实验室的支持来自P30中心、ROI和合同。 近年来，我们的小组广泛地使用了稳定同位素方法，我们认为这是创新性的。 稳定的同位素标记药物，不像放射性同位素标记的药物， 无标签药物稳定的同位素，如氢同位素氘，结合到药物分子中，允许标记的药物用作示踪剂。这是药代动力学和代谢研究中的有力工具，经常用于生物利用度研究。虽然天然药物是通过其通常的途径，如口服，透皮，或通过吸烟，标记的药物同时静脉内给药的药代动力学表征。我们使用该技术测定了吸烟（Benowitz et al. 1991 a）和无烟烟草（Jacob et al. 1999）的尼古丁摄入量、经皮尼古丁的生物利用度（Benowitz et al. 1991 b）和通过各种途径给药的可卡因的生物利用度。我们用稳定同位素方法研究了可卡因和乙醇的代谢处置，包括测定可卡因和乙醇的代谢产物。 可卡因部分转化为可卡因乙烯（Jacob等人，1997; Everhart等人，1998）。使用稳定同位素方法测定鼻内和烟熏甲基苯丙胺的生物利用度（Harris等人，2003）。我们对稳定同位素的使用正在继续和扩大。我们将利用标记的反式-3 '-羟基可替宁来进一步了解尼古丁的药物遗传学，并更好地了解尼古丁代谢中种族差异的机制。可替宁-d4和代谢物比率将用于研究青少年轻度吸烟者中代谢率与成瘾发展的相关性（Mark Rubinstein，MD，CA 140216）。 我们使用稳定同位素方法来解决药物滥用领域特有的问题。其中一个问题是确定静脉内尼古丁替代是否会抑制吸烟引起的尼古丁摄入（Benowitz和Jacob 1990）。由此得出结论，尼古丁替代药物如透皮贴剂（当时正在进行上市前临床试验）可以抑制吸烟，即使受试者无法完全戒烟。我们已经使用氘标记的可卡因管理，以研究可卡因及其代谢产物苯甲酰芽子碱到人的头发中的分布的时间过程。出于伦理原因，这些研究必须在可卡因滥用者中进行，并且使用标记药物可以保证随后的街头可卡因使用不会使结果无效（亨德森等人，1996年）。我们还使用标记的尼古丁来研究尼古丁在头发和指甲中积累的时间过程，这似乎是尼古丁暴露的良好长期生物标志物。 这些研究需要有合成和分析化学家的实验室以及现代分析仪器。当使用稳定同位素时，广泛使用质谱是必要的。适当标记的药物或代谢物可能无法在商业上获得，我们需要在合成有机化学方面的能力来制备稳定同位素标记的药物，代谢物和用于临床研究和测定的内标物。在此申请中，我们要求实验室和行政人员以及仪器的支持，以保持和提高我们的分析化学能力。

The need for analytical chemistry support for clinical studies arose because of increased appreciation of the importance of pharmacokinetics in understanding drug action in the late 1970s. Neal Benowitz had initiated pharmacokinetic studies of nicotine and opioids and Reese Jones initiated studies of cocaine pharmacokinetics and pharmacodynamics, largely supported by a P50 award. At that time, our analytical laboratory consisted of a chemist, two technicians and two gas chromatographs. Since, many projects requiring analytical chemistry support were initiated and our laboratory staff has grown from three to 14: Two PhD Research Chemists, nine Staff Research Associates and three Laboratory Assistants. Major equipment includes two gas chromatographs, two desktop GC-MS systems, two HPLCs, three triple-stage quadrupole LC-MS/MS systems, and two triple-stage quadrupole GC-MS/MS systems. Support for the laboratories comes from the P30 Center, ROIs and contracts. In recent years, our group has made extensive, and we believe innovative, use of stable isotope methodology. Stable isotope-labeled drugs, unlike those labeled with radioisotopes, are no more hazardous than unlabeled drugs. A stable isotope, such as the hydrogen isotope deuterium, incorporated into a drug molecule allows the labeled drug to be used as a tracer. This is a powerful tool in studies of pharmacokinetics and metabolism, frequently used in bioavailability studies. While the natural drug is administered by its usual route, such as oral, transdermal, or by smoking, the labeled drug is simultaneously administered intravenously for pharmacokinetic characterization. We have used this technique to determine nicotine intake from smoking (Benowitz et al. 1991a) and from smokeless tobacco (Jacob et al. 1999), bioavailability of transdermal nicotine (Benowitz et al. 1991b) and bioavailability of cocaine administered by various routes. We have used stable isotope methodology to study the metabolic disposition of cocaine and ethanol, including determination of the fractional conversion of cocaine to cocaethylene (Jacob et al. 1997; Everhart et al. 1998). Stable isotope methodology was used to determine the bioavailablity of intranasal and smoked methamphetamine (Harris et al. 2003). Our use of stable isotopes is continuing and expanding. We will be utilizing labeled frans-3'-hydroxycotinine to further our understanding of nicotine pharmacogenetics and to better understand the mechanism of racial differences in nicotine metabolism.. Cotinine-d4 and the metabolite ratio will be used to study the association of the rate of metabolism and development of addiction in adolescent light smokers (Mark Rubinstein, MD, CA140216). We have used stable isotope methodology to address questions unique to the drug abuse area. One such question was to determine whether intravenous nicotine replacement would suppress nicotine intake from smoking (Benowitz and Jacob 1990). This led to the conclusion that nicotine replacement medications such as transdermal patches (at that time undergoing premarketing clinical trials) would suppress smoking even if subjects were unable to quit entirely. We have used deuterium-labeled cocaine administration to study the time course of distribution of cocaine and its metabolite benzoylecgonine into human hair. For ethical reasons, these studies had to be carried out in cocaine abusers, and the use of labeled drug guaranteed that subsequent street cocaine use would not invalidate the results (Henderson et al. 1996). We are also using labeled nicotine to study the time course of accumulation of nicotine into hair and nails, which appear to be good long term biomarkers of nicotine exposure. Such studies require laboratories with synthetic and analytical chemists, and modern analytical instruments. Extensive use of mass spectrometry is necessary when using stable isotopes. Suitably labeled drugs or metabolites may not be commercially available, and our capability in synthetic organic chemistry has been needed to prepare stable-isotope labeled drugs, metabolites, and internal standards for clinical studies and assays. In this application, we are requesting support for laboratory, and administrative staff, and for instrumentation to maintain and enhance our analytical chemistry capabilities.