The role of metabonomics at the interface between drug metabolism and safety assessment

Safety assessment of candidate drugs is a key stage in drug development and one which represents a significant attritional hurdle. There is also a focused effort in drug metabolism studies to assess bioactivation potential based on the notion this could lead to the risk of macromolecule adduct formation and subsequent toxicological consequences. However, characterization of the molecular mechanisms of drug toxicity still remains an enormous challenge. Recent advancements in 'omics sciences, and in particular metabonomics, has enabled some elucidation or insights into toxicological sequelae. Metabonomics is a global metabolic profiling framework which utilizes high resolution analytics (typically NMR and mass spectrometry) together with chemometric statistical tools (such as principal component analysis and partial least squares) to derive an integrated picture of both endogenous and xenobiotic metabolism. This review details some of the current progress in the application of metabonomics in drug safety and metabolism.     

Identification of end points relevant to detection of potentially adverse drug reactions

Expectations are high that the use of proteomics, gene arrays and metabonomics will improve risk assessment and enable prediction of toxicity early in drug development. These molecular profiling techniques may be used to classify compounds and to identify predictive markers that can be used to screen large numbers of chemicals. One of the challenges for the scientific community is to discriminate between changes in gene/protein expression and metabolic profiles reflecting physiological/adaptive responses, and changes related to pathology and toxicology. In these proceedings we provide a brief overview of the technologies with focus on proteomics and the possible applications to mechanistic and predictive toxicology. The discussion also includes strengths and limitations of molecular profiling technologies.     

代谢组学技术在痛风类疾病研究中的应用进展

目的 对近年来代谢组学技术在痛风领域的相关研究进行综述,以期为痛风今后更深入的研究提供思路和方向.方法 通过PubMed检索相关文献,分析和总结代谢组学技术在嘌呤代谢、高尿酸血症和痛风相关疾病研究中的应用.结果 代谢组学技术被广泛应用于痛风类疾病的生物标志物发现、发病机制探索以及药物作用机制研究,同时代谢组学技术与其他...     

Metabonomics Techniques and Applications to Pharmaceutical Research & Development

In this review, the background to the approach known as metabonomics is provided, giving a brief historical perspective and summarizing the analytical and statistical techniques used. Some of the major applications of metabonomics relevant to pharmaceutical Research & Development are then reviewed including the study of various influences on metabolism, such as diet, lifestyle, and other environmental factors. The applications of metabonomics in drug safety studies are explained with special reference to the aims and achievements of the Consortium for Metabonomic Toxicology. Next, the role that metabonomics might have in disease diagnosis and therapy monitoring is provided with some examples, and the concept of pharmacometabonomics as a way of predicting an individual's response to treatment is highlighted. Some discussion is given on the strengths and weaknesses, opportunities of, and threats to metabonomics.     

Paradigm Shift in Toxicity Testing and Modeling

The limitations of traditional toxicity testing characterized by high-cost animal models with low-throughput readouts, inconsistent responses, ethical issues, and extrapolability to humans call for alternative strategies for chemical risk assessment. A new strategy using in vitro human cell-based assays has been designed to identify key toxicity pathways and molecular mechanisms leading to the prediction of an in vivo response. The emergence of quantitative high-throughput screening (qHTS) technology has proved to be an efficient way to decompose complex toxicological end points to specific pathways of targeted organs. In addition, qHTS has made a significant impact on computational toxicology in two aspects. First, the ease of mechanism of action identification brought about by in vitro assays has enhanced the simplicity and effectiveness of machine learning, and second, the high-throughput nature and high reproducibility of qHTS have greatly improved the data quality and increased the quantity of training datasets available for predictive model construction. In this review, the benefits of qHTS routinely used in the US Tox21 program will be highlighted. Quantitative structure-activity relationships models built on traditional in vivo data and new qHTS data will be compared and analyzed. In conjunction with the transition from the pilot phase to the production phase of the Tox21 program, more qHTS data will be made available that will enrich the data pool for predictive toxicology. It is perceivable that new in silico toxicity models based on high-quality qHTS data will achieve unprecedented reliability and robustness, thus becoming a valuable tool for risk assessment and drug discovery.     

NMR-based metabolomics

Similar to genomics and proteomics which yield vast amounts of data about the expression of genes and proteins, metabolomics refers to the whole metabolic profile of the cell. The focus of this report concerns the use of nuclear magnetic resonance (NMR) spectroscopy for metabolic analyses and, in particular, its use in toxicology for examining the metabolic profile of biofluids. Examples from the literature will demonstrate how 1H NMR and pattern recognition methods are used to obtain the urinary metabolic profile, and how this profile is affected by exposure to various toxicants. These particular studies which focus on the metabolic profiles of biofluids, specifically urine, are referred to as metabonomics. NMR-based metabonomics provides a means to categorize organ-specific toxicity, monitor the onset and progression of toxicological effects, and identify biomarkers of toxicity. A future challenge, however, is to describe the cellular metabolome for purposes of understanding cellular functions (i.e., metabolomics). Thus the capabilities and advantages of multinuclear NMR to provide metabolic information in cells and tissues will also be discussed. Such information is essential if metabolomics is to provide a complementary dataset which together with genomics and proteomics can be used to construct computer network models to describe cellular functions.     

基于血浆代谢组学评价不同神经保护剂联用方案对缺血性脑卒中生物标志物的影响

目的：通过代谢组学挖掘神经保护剂联用治疗缺血性脑卒中患者血浆的回调性生物标志物,从代谢路径角度评价神经保护剂联用的合理性。方法：通过高效液相色谱与飞行时间质谱（Q-TOF）联用的非靶向代谢组学平台,获取缺血性脑卒中患者的血浆代谢组学数据,用多元变量统计分析进行数据处理,筛选出药物干预后回调的生物标记物及其代谢路径。结果：代谢组学研究表明缺血性脑卒中患者体内主要呈氨基酸、脂类及能量代谢紊乱,给予不同神经保护剂联用方案后整体代谢轮廓回调,经数据库鉴定和匹配后得到两药联用-丁苯酞+依达拉奉组、两药联用-丁苯酞+单唾液酸组和三药联用-丁苯酞+依达拉奉+单唾液酸组显著变化的共性回调生物标记物及特征性回调生物标记物（溶血磷脂酰胆碱18∶0、色氨酸和苹果酸等）。结论：神经保护剂联合应用可使缺血性脑卒中患者的代谢紊乱恢复。调节能量代谢（三羧酸循环）、脂类代谢（甘油磷脂代谢、脂肪酸代谢）和氨基酸代谢（芳香族氨基酸代谢、鸟氨酸代谢）通路等可能是其治疗缺血性脑卒中的神经保护机制。     

High performance liquid chromatography/atmospheric pressure ionization/tandem mass spectrometry (HPLC/API/MS/MS) in drug metabolism and toxicology

Studies of the metabolic fate of drugs and other xenobiotics in living systems may be divided into three broad areas: (1) elucidation of biotransformation pathways through identification of circulatory and excretory metabolites (qualitative studies); (2) determination of pharmacokinetics of the parent drug and/or its primary metabolites (quantitative studies); and (3) identification of chemically-reactive metabolites, which play a key role as mediators of drug-induced toxicities (mechanistic studies). Mass spectrometry has been regarded as one of the most important analytical tools in studies of drug metabolism, pharmacokinetics and biochemical toxicology. With the commercial introduction of new ionization methods such as those based on atmospheric pressure ionization (API) techniques and the combination of liquid chromatography-mass spectrometry (LC-MS), it has now become a truly indispensable technique in pharmaceutical research. Triple stage quadrupole and ion trap mass spectrometers are presently used for this purpose, because of their sensitivity and selectivity. API-TOF mass spectrometry has also been very attractive due to its enhanced full-scan sensitivity, scan speed, improved resolution and ability to measure the accurate masses for protonated molecules and fragment ions. This review aims to survey the utility of mass spectrometry in drug metabolism and toxicology and to highlight novel applications and future trends in this field.     

代谢组学在肝肾损伤研究中的应用

代谢组学是近年来新发展起来的一门组学,已成为生物医学研究领域的新热点.由于其广泛的应用前景,目前已是系统生物学的重要组成部分.肝损伤及肾损伤是临床常见的危害人类健康的疾病,但目前的指标缺乏特异性,且灵敏度不高,造成临床上难以对肝肾损伤做出准确而及时的预警和监测.代谢组学能从代谢的角度整体分析疾病,快速、安全地对疾病的严重程度进行判别,在肝肾损伤的早期诊断等方面的研究已初显优势.本文主要综述了代谢组学技术在肝肾损伤研究中的应用现状.     

胆红素代谢酶UGT1A1介导的中药不良反应研究进展

尿苷二磷酸葡萄糖醛酸转移酶1A1（UGT1A1）是哺乳动物体内分布的一种重要的Ⅱ相代谢酶,其不仅介导了大量临床药物、毒物的代谢清除,还是机体参与内源性毒性物质胆红素代谢清除的唯一代谢酶。药物或食品中的化学成分对UGT1A1的强烈抑制会引发多种不良反应,如药物/草药-药物相互作用、高胆红素血症、肝功能异常及神经毒性等。近年来,研究发现许多草药提取物及其化学成分可通过强效抑制UGT1A1引发药物-草药相互作用等不良反应。结合药物代谢及药物毒理学相关领域的新近研究进展,系统总结了UGT1A1抑制剂筛选与评价常用方法,以及中草药化学成分对UGT1A1抑制作用的研究进展。上述信息对于临床安全合理的使用中草药,尤其是在中西药联用或中药方剂使用过程中尽可能避免因UGT1A1抑制引发的不良反应,具有重要指导意义。     

NMR-BASED METABOLOMICS

ABSTRACT

Similar to genomics and proteomics which yield vast amounts of data about the expression of genes and proteins, metabolomics refers to the whole metabolic profile of the cell. The focus of this report concerns the use of nuclear magnetic resonance (NMR) spectroscopy for metabolic analyses and, in particular, its use in toxicology for examining the metabolic profile of biofluids. Examples from the literature will demonstrate how ¹H NMR and pattern recognition methods are used to obtain the urinary metabolic profile, and how this profile is affected by exposure to various toxicants. These particular studies which focus on the metabolic profiles of biofluids, specifically urine, are referred to as metabonomics. NMR-based metabonomics provides a means to categorize organ-specific toxicity, monitor the onset and progression of toxicological effects, and identify biomarkers of toxicity. A future challenge, however, is to describe the cellular metabolome for purposes of understanding cellular functions (i.e., metabolomics). Thus the capabilities and advantages of multinuclear NMR to provide metabolic information in cells and tissues will also be discussed. Such information is essential if metabolomics is to provide a complementary dataset which together with genomics and proteomics can be used to construct computer network models to describe cellular functions.     

Biomarkers,metabonomics, and drug development: Can inborn errors of metabolism help in understanding drug toxicity?

Application of "omics" technology during drug discovery and development is rapidly evolving. This review evaluates the current status and future role of "metabonomics" as a tool in the drug development process to reduce the safety-related attrition rates and bridge the gaps between preclinical and clinical, and clinical and market. Particularly, the review looks at the knowledge gap between the pharmaceutical industry and pediatric hospitals, where metabonomics has been successfully applied to screen and treat newborn babies with inborn errors of metabolism. An attempt has been made to relate the clinical pathology associated with inborn errors of metabolism with those of drug-induced pathology. It is proposed that extending the metabonomic biomarkers used in pediatric hospitals, as "advanced clinical chemistry" for preclinical and clinical drug development, is immediately warranted for better safety assessment of drug candidates. The latest advances in mass spectrometry and nuclear magnetic resonance (NMR) spectroscopy should help replace the traditional approaches of laboratory clinical chemistry and move the safety evaluation of drug candidates into the new millennium.     

新药研发中毒理学研究方法的进展--非临床安全性评价的新工具

近年来在新药非临床安全性评价中越来越多采用新方法和新技术，以适应不同特点新药的开发和使安全性评价更具预测性。现综述国外近几年在非临床安全性评价中采用的新工具，包括生物标记物，以及毒理基因组学、毒理蛋白质组学、代谢组学和系统毒理学方面的新技术。     

An integrated metabonomic method for profiling of metabolic changes in carbon tetrachloride induced rat urine

Carbon tetrachloride (CCl₄) is a well-known model compound for inducing chemical hepatic injury. This work characterizes the metabolism disorders of hepatotoxicity induced by CCl₄ in a Wistar rat model with a single dosage of 1 ml/kg. A seven-day long continuous collection of urine was performed in male rats in this experiment. Blood biochemistry and histopathology were examined to identify specific changes of liver hepatotoxicity. At the same time, an integrated analytical approach based on liquid chromatography coupled with mass spectrometry (LC–MS) was developed to map the metabolic response in urine. The current metabonomic approach based on LC–MS indicated 23 endogenous metabolites as biomarkers in urine associated with the hepatotoxicity induced by CCl₄. The underlying regulations of CCl₄-perturbed metabolic pathways were discussed according to the identified metabolites. The present study proves the great potential of LC–MS based metabonomics in mapping metabolic response for toxicology.     

In vitro approaches to studying the metabolism of new psychoactive compounds

In the last two decades, a large number of new drugs from several drug classes have appeared on the illicit drug market. While some of these compounds have meanwhile been scheduled as controlled substances, the majority of them are (still) sold as so-called 'legal highs', mostly via the Internet. At the time they appear on the market the metabolism of these drugs is generally unknown. Therefore, it must be studied in order to obtain data necessary for analytical method development as well as toxicological risk assessment. In vitro metabolism studies of new designer drugs can be performed for identification and structure elucidation of new designer drug metabolites or to assess the qualitative and quantitative involvement of certain enzymes in the metabolism of a particular drug. In this review, the value of the following enzyme preparations for in vitro metabolism studies of new designer drugs will be discussed: liver microsomes, recombinant cDNA-expressed enzymes, liver cytosol, S9 mix, and hepatocytes. This will cover the major metabolic enzymes: cytochrome P450 monooxygenases, flavin-monooxygenases, monoamine oxidases, UDP-glucuronyltransferases, sulfotransferases, and catechol-O-methyltransferases. Important analytical aspects such as the value of mass spectrometric techniques will also be covered.     

Metabolic profiling in disease diagnosis, toxicology and personalized healthcare

Metabolic profiling employs a combination of sophisticated analytical tools to obtain global "untargeted" metabolic profiles from tissues, cells or biofluids. The resulting complex multivariate data are then modeled statistically to reveal differences between classes (e.g. dosed vs. control) and identify discriminatory metabolites. Metabolic profiling has a wide range of applications, encompassing nutrition, disease diagnosis, epidemiology and toxicology, providing insights into altered biological pathways and offering fresh mechanistic perspectives. Further, the untargeted nature of metabolic profiling can allow for new biomarkers of disease or toxic effect to be uncovered. In this review, key metabolic profiling technologies will be introduced and data analysis approaches described briefly. The role of metabolic profiling in disease diagnosis, toxicology and personalized healthcare will be discussed.     

原代肝细胞培养及其在药物代谢和毒理学研究中的应用进展

原代肝细胞培养技术已有四十几年的历史,随着技术水平的不断提高,其培养越来越成熟,并且已广泛用于研究药物对细胞色素P450酶的作用及其机制,药物相互作用,药物毒理学等多方面。因为其很好的维持和保留了肝脏的代谢能力,具有与体内一致的细胞色素P450酶的活性,成为日益重要和可信赖的评价药物和新化学实体的安全与有效的体外模型。该文对原代肝细胞培养技术及其在药物代谢和毒理学研究中的应用进展进行了综述。     

Spectroscopic and statistical methods in metabonomics

Metabonomics is rapidly evolving through advances in analytical technologies together with the development of new hyphenated approaches that are increasingly being applied to analyze complex biological systems. Improvements in analytical performance, such as increased sensitivity and selectivity, are providing greater resolution to analytical datasets and the rich potential of metabonomics as a systems biology tool of choice is becoming clear. However, such improvements are resulting in datasets becoming increasingly demanding in terms of data handling and interpretation, and the degree to which metabonomics continues to develop will be dependent on how chemometrics and data-handling approaches keep pace with continually improving analytical technologies. This review provides an overview of the field of metabonomics, with a particular focus on the analytical techniques that are chiefly employed and the chemometric methods that have found most use. However, in addition, we mention less widely used analytical methods and suggest that advanced statistical methods will play a larger role in the future.     

Application of metabonomics in drug development

Metabolic profiling (metabonomics/metabolomics) is the untargeted analysis of metabolic composition in a biological sample, and is principally aimed at biomarker discovery. The frequent use of noninvasive biofluid analysis in metabonomics is suited to the clinic and facilitates dynamic monitoring. Analytical protocols for metabolic biomarkers are potentially robust because a metabolite is the same chemical entity irrespective of its origin, facilitating 'bench-to-bedside' translational research. Metabonomics can make an impact at several points in the drug-development process: target identification; lead discovery and optimization; preclinical efficacy and safety assessment; mode-of-action and mechanistic toxicology; patient stratification; and clinical pharmacological monitoring. This review describes and exemplifies the latest developments in each of these areas, including the impact of new data and chemical analytical techniques. The future goals for metabonomics are the validation of existing biomarkers, in terms of mechanism and translation to man, together with a focus on characterizing the individual ('personalized healthcare').