Xenobiotica the fate of foreign compounds in biological systems
ISSN: 0049-8254 (Print) 1366-5928 (Online) Journal homepage: http://www.tandfonline.com/loi/ixen20
Twenty years on: a review of the current practice of drug metabolism and pharmacokinetic studies in the pharmaceutical industry J. A. Bell & M. H. Tarbit To cite this article: J. A. Bell & M. H. Tarbit (1992) Twenty years on: a review of the current practice of drug metabolism and pharmacokinetic studies in the pharmaceutical industry, Xenobiotica, 22:7, 735-742, DOI: 10.3109/00498259209053136 To link to this article: http://dx.doi.org/10.3109/00498259209053136
Published online: 22 Sep 2008.
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Date: 21 March 2016, At: 22:07
XENOBIOTICA,
1992, VOL. 22,
NO.
7, 735-742
Comment Twenty years on: a review of the current practice of drug metabolism and pharmacokinetic studies in the pharmaceutical industry
J. A. BELL and M . H. T A R B I T Drug Metabolism Division, Giaxo Group Research Limited, Park Road, Ware, Herts., UK
Downloaded by [ECU Libraries] at 22:07 21 March 2016
(Received 1 November 1991; accepted 20 December 1991)
Introduction T h e 10th anniversary of the Drug Metabolism Discussion Group* was marked by a special issue of Xenobiotica in which Dr David Case (1981) reviewed the then current state of the art of drug metabolism and pharmacokinetics within the pharmaceutical industry. Ten years on we have another celebratory issue of this Journal and a further opportunity to review our field. Rather than make any attempt to emulate Case’s excellent commentary, we will restrict our review to a consideration of the progress and changes that have occurred during the intervening years. Many of the papers presented in this issue provide a suitable barometer for assessing these changes. For simplicity and to avoid the semantic difficulties noted by Case, we will refer to drug metabolism and pharmacokinetics as DMPK.
Role of industrial drug metabolism and pharmacokinetics Although recognizing the potential contribution of D M P K in supporting compound selection and pharmacology studies in animals, Case’s commentary was very largely devoted to the role of D M P K in safety evaluation programmes. In 1981 this was an accurate reflection of the major focus of drug metabolism and pharmacokinetic effort within the industry. T h e last 10 years has seen a continuation in the application of D M P K to safety evaluation and, indeed, validation of toxicity studies has become the major raison d’2tre of pre-clinical drug metabolism. However, we believe that greater attention is now paid to the role of D M P K in drug discovery programmes than was the case in 1981. It is difficult to substantiate this assertion with hard facts: the commercial sensitivity of many drug discovery projects precludes early publication of D M P K input. Nevertheless, information imparted at informal meetings indicates that many companies have recognized that the skills-base residing in D M P K groups can be applied successfully to solve problems such as low potency and inappropriate duration that may be identified in secondary pharmacology screens. An extension of this’ application is that an early
*DMDG is an informal association of scientists engaged in the study of drug metabolism and pharmacokinetics in the pharmaceutical industry. 0049-8254/92 $3.00 0 1992 Taylor & Francis Ltd.
736
J . A. Bell and M . H . Tarbit
input from DMPK scientists can minimize the risk of selecting for development a drug candidate with inappropriate pharmacokinetics. This ‘added-value’ helps to reduce the timescale and hence cost of drug development programmes. These principles are exemplified in the review by Humphrey and Smith (1992). We perceive that an early input from D M P K scientists has also become the norm in clinical development. Currently it is now common for preliminary pharmacokinetic information to be obtained in the first study of safety and tolerability in healthy volunteers. Although definitive pharmacokinetic studies will follow, this early information can give valuable insights on how the new drug is handled in man. Further exploitation of D M P K science is likely to arise from the great explosion of knowledge in the area of drug metabolizing enzymes discussed below.
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Regulatory requirements At first sight it would appear that there have been extensive changes in regulatory requirements during the last 10 years. Most obvious of these are in Europe where new guidelines covering pre-clinical and clinical D M P K prepared by the Committee for Proprietary Medicinal Products (CPMP) have been published by the Commission of the European Communities (Commission of the EC 1989a). On closer examination, however, these ‘multi-state’ guidelines differ little in content from their national predecessors, e.g. MAL-4 ( D H S S 1977). It is interesting to note that, with the exception of single dose studies, all the guidelines for toxicity tests refer to requirements for metabolic and kinetic data. This serves to illustrate the central role of D M P K in safety evaluation (Davies 1988) and has been further enhanced by the introduction of the ‘Expert Report’ system which requires applicants seeking marketing authority in the European Community to submit a critical evaluation of their product (Commission of the EC 1989b). There have been relatively few new guidelines from the FDA, although it is apparent from recent scientific meetings that this Agency is also now recognizing the importance of toxicokinetics in safety evaluation studies. In differentiating between toxicologists and kineticists Case (1981) noted that the latter
‘. . . responds to broad principles of regulatory concern, often wishes for more specific guidance but always wants to retain a large measure of flexibility in study design.’ As noted above, guidelines have not become more specific during the past 10 years and significant flexibility in the design of studies to support drug submissions remains. For example, many companies are now supplementing ‘classical’ drug metabolism studies by experiments carried out in vitro, although it is not clear whether the information obtained is used for regulatory purposes. In particular, the number of reports on the use of isolated hepatocytes (e.g. Lavrijsen et al. 1992) has increased dramatically over the past 10 years. Looking to the future, it appears that flexibility of study design will be enhanced further rather than reduced. A draft note for guidance from CPMP on non-clinical testing strategies (Commission of the EC 1990a) states:
‘. . . detailed, fixed experimental dosing programmes prescribing animal species, types of tests and duration of treatment, which would be valid for all types of medicinal products and all trial phases in humans (including early
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pharmacokinetic studies in humans) cannot be scientifically justified. In order to avoid this type of rigid, dogmatic approach, only general guidance for the decision making processes regarding the need for experimental studies prior to human studies (and therapeutic use) can be given.’ Adoption of this ‘product-tailored’ approach in submissions to regulatory authorities has many attractive features such as an anticipated reduction in the number of experimental animals and the facilitation of global harmony in regulatory requirements. However, there are potential problems. During a drug development programme of say 10 years duration, decisions may be taken on the basis of scientific knowledge that could change significantly by the time the submission is reviewed. It appears, therefore, that joint agreement of scientific justification is likely to require effective communication between the applicant and regulatory authority at key stages throughout drug development. One aspect of D M P K where we have seen marked changes in regulatory expections is in the area of bioequivalence where a current CPMP draft note for guidance (Commission of the EC 1990b) contains considerably more detail than its predecessor (Commission of the EC 1989a). T h e review of statistical aspects of bioequivalence (Pidgen 1992) is therefore timely and validation of analytical methodology (Arnoux and Morrison 1992) is also receiving increased attention. In addition, the relationship between pharmacokinetics and pharmacodynamics (e.g. Ings et al. 1992) has been the subject of growing regulatory interest, particularly in the US, and there is an indication (e.g. Collins et al. 1990) that this area of research may be applied to the selection of clinical doses. T h e past 10 years has also seen an increase in pharmacokinetic studies in the elderly and in patients (e.g. Hughes et al. 1992). For the D M P K scientist perhaps the most significant change in the regulatory scene has been in the area of stereochemical drugs. Although some countries, e.g. Sweden and Switzerland, have produced discussion documents, as yet no regulatory authority has produced guidelines in this area. However, the CPMP Notes to Applicants (Cbmmission of the EC 1989b) states inter alia: ‘Possible problems relating to stereoisomerism, which should be discussed in the appropriate Expert Report and cross referenced, should include: -
pharmacokinetics (including information on the relative metabolism of the stereoisomers)
-
extrapolation of the preclinical data (paying particular attention to possible problems relating to species differences in handling of the stereoisomers)’.
This development has had a significant impact in our field and has provided new challenges to the bioanalyst. I n fact it might be argued that the current interest in the pharmacokinetics of stereochemical drugs has arisen through the recent development of enantiospecific analytical methodology (Hutt 1990).
Tools of the trade Throughout the past 10 years the pharmaceutical industry has continued to develop increasingly potent drugs. In turn the D M P K scientist investigating the fate of these drugs and their metabolites in the body is faced with the need to develop increasingly sensitive bioanalytical techniques.
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Currently it appears that technology is keeping pace with demand and low concentration (nglml) assays are commonplace. For quantitative work h.p.1.c. remains the most widely used technique. A variety of detectors, e.g. UV, fluorescence and electrochemical (Hughes et al. 1992) are now used routinely in conjunction with diverse column packing materials. Although direct injection methods can often be employed (e.g. Doyle et al. 1992) the bioanalyst has at his disposal an extensive armoury of solid phases for the selective extraction of drugs from body fluids. Supercritical fluid chromatography (SFC) and capillary zone electrophoresis (CZE) both of which have been developed during the past decade are beginning to offer alternatives to h.p.1.c. Immunoassay has frequently been considered inferior to chromatography for the quantification of drugs in biological fluids, a reputation obtained largely through lack of specificity in the assay of steroidal drugs. However, the past 10 years have seen increased use of specific immunoassays in response to the problems of analysing very potent drugs. In addition, cross-reactivity between antisera and metabolites may be exploited in the development of selective extraction methods that rely on structural characteristics rather than physiochemical properties of the analyte. Cross-reactivity between analogues can also be exploited in drug discovery where antisera raised against a drug development candidate may often be useful in the assay of closelyrelated back-up compounds. T h e increased requirement for information on the pharmacokinetics of stereochemical drugs, discussed above, has led to considerable activity in the field of chiral analysis (e.g. Beresford et al. 1992, Tomlinson et al. 1992) and chiral disposition studies (Tomlinson et al. 1992). There have also been significant developments in the application of spcctrometric methods (Wiltshire et al. 1991). In the first of mass spectrometry rew sample introduction techniques have facilitated the interfacing of spectrometers with chromatographs. Of the ‘hyphenated’ techniques, only g.1.c.-mass spectrometry was widely used in D M P K laboratories 10 years ago. However, h.p.1.c.-mass spectrometry and more recently tandem mass spectrometery are now routinely applied to the identification of drug metabolites. Improvements in resolution, analytical power and sensitivity have also occurred in n.m.r., with its application to fluorinated compounds being very effective (e.g. Gilbert et al. 1992). Advances in multipulse sequences have enhanced the available information through the development of two-dimensional techniques, e.g. COSY, although it appears that the industrial drug metabolism scientist has been slow to exploit these major developments (Preece and Timbrel1 1990). T h e ‘microchip revolution’ has continued to provide opportunities for the automation of our function. Computers can now control information flow from robotic instrumentation through data acquisition and processing to submission of the dossier to regulatory authorities. Although this development enhances compliance with good laboratory practice (GLP) by reducing or removing error-prone manual data entry, further GLP problems are introduced by the requirement for validation of the systems. Progress towards the fully automated, GLP-compliant laboratory has therefore been slower than perhaps predicted 10 years ago. The methodology in routine use by industrial DMPK sections still requires us to analyse drug and metabolites in samples of biological fluids that have been removed from the body and hence the information is obtained some time after the event. However, the past 5 years have seen the emergence of dynamic techniques such as
Current practice of drug metabolism and pharmacokinetic studies
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continuous flow fast atom bombardment mass spectrometry (CF-FAB) (Caprioli 1990) which in combination with microdialysis probes enable the acquisition of pharmacokinetic data in ‘real-time’. Techniques are also now available that allow n.m.r. to be used in whole animals (Preece and Timbrel1 1990) and further intriguing possibilities are presented by the application of positron-emitting isotopes to the investigation of clinical pharmacokinetics (Cunningham et al. 1991). In the future these exciting developments may present a significant business opportunity for industrial DMPK.
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New business for an old trade In many ways D M P K has grown as a synthesizing science aiding interpretation of toxicity tests and relationships to clinical use. In some areas of our business, however, this synthesis has failed to move the goalposts. A good example of this is genetic toxicology, a science fundamentally integrated with drug metabolism. Thus tests in vitro used to evaluate the genotoxicity of compounds and their metabolites were established on the drug metabolism science of the 1970s (Ames et al. 1975). Despite the huge growth in the understanding of the biology of drug-metabolizing enzymes over the intervening years they have essentially stayed that way. In many respects this is because the increased understanding of the role of enzymes in activiation has occurred primarily in academia and has been of limited interest to D M P K scientists in industry. We have therefore not been positioned to advise on the design of genetic toxicity tests, and communication between genetic toxicologists and drug metabolism scientists within the industry has been poor. We also have a poor record over the past 10 years in understanding how our drugs interact with drug-metabolizing enzymes, the mechanism of inhibition and induction, or even which enzymes are responsible for their metabolism or activation. This seems set to change. Like many areas of biological science the coming of the molecular biology ‘revolution’ has been fundamental in its impact on the enzymology of drug metabolism. No one 10 years ago could have foreseen the phenomenal rate at which the knowledge and understanding of cytochrome P-450 has moved, e.g. in 1982 about half a dozen P-450s had been isolated and identified in the various species examined (Ryan et al. 1982) and the wide role of the enzyme system in the regulation of steroidogenesis was just emerging (Waterman, 1982). Recently over 100 genes of P-450 in the plant and animal kingdom were classified (Nebert et al. 1991) with the high probability of discovering many m0r.e. T h e enzyme is now recognized as being essentially ubiquitous with multiple endogenous roles in addition to that of drug metabolism. Relatively under-researched systems, such as glucuronyl transferases, glutathione transferases and flavin monooxygenase, are now also assuming markedly higher profiles within the industry than 10 years ago, as the techniques of molecular biology allow more facile identification, isolation and characterization of their role in drug metabolism and detoxication. T h e impact of molecular biology techniques allied to the availability of human tissue has led to several established dogmas on the role of various enzymes in metabolism and activation of xenobiotics in animals and man being challenged. Thus recent studies have highlighted evidence that activation of a significant number of carcinogens and hepatotoxins is mediated via constitutively expressed enzymes in animals and man (Shimada et al. 1989), some of which are suppressed by the induction regimens regularly adopted for the metabolic activation systems used in genetic toxicity testing.
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In addition, we now have for the first time, opportunities via cloning and expression systems to examine the role of individual animal and human enzymes in the metabolism of our drugs in vitro-long before the compound reaches man. Several of the expression techniques in cell lines offer the hope, if not the certainty, that in the future, species validation of metabolism in vitro may be better established than it has been in the past. These techniques also provide the means to address issues such as genetic polymorphism of our drugs using simple tests in vitro without the need for costly clinical studies. In this area we may also see the introduction of identification of ‘poor metabolisers’ in the population by simple kinetic blood tests (Heim and Meyer 1991, Wolf et al. 1990), thus avoiding the need for expensive phenotyping studies. Currently it is difficult to appreciate whether the pharmaceutical industry regards these developments as an opportunity or a threat. T h e positive view of this explosion of knowledge and technique is that it provides scope to understand significantly more about the mechanisms and in the inter-relationships concerned in the processing of our drug candidates and their effects on the functions and expression of enzymes. This is surely important if we are to put context to the questions that face us such as the significance of enzyme induction by drugs in their long-term clinical benefitlrisk assessment. T h e one ‘isle of doubt in the sea of euphoria’ surrounding the new science is the apparent conservatism of the industrial D M P K scientist. Much of this research and technology still lies in academia. We in industrial drug metabolism have been somewhat slow to seize these opportunities despite the fact that molecular science has been embraced avidly by our pharmacology and biochemist research colleagues. Many pharmaceutical companies have molecular biology departments: few are functioning on behalf of their drug metabolism needs. We believe that we must be prepared to be proactive in seeking to establish where these techniques will work for us if we are to avoid the risk of these elements becoming ‘requirements’ before their significance is placed in context, or taking the form of uncertainty factors in the development of drugs. In this respect the context and timing of these studies in the development process should be the subject of wide and continuing debate within the industrial environment. Overall, we should aim to use the opportunities presented by these techniques to enable us to answer regulatory questions which are increasingly likely to become focused on how new chemical entities interact with drug metabolizing enzymes.
Conclusions Many of the aspects of our work discussed by Case (1981) have remained essentially unchanged and the past 10 years have represented a consolidation of the business of industrial drug metabolism and pharmacokinetics. Nevertheless, there have been considerable advances in technology to which D M P K scientists have responded rapidly and which have been exploited in the application of ‘classical studies’ to increasingly potent drugs. Response from industrial scientists to the huge growth in our understanding of drug metabolising enzymes has been much less obvious. Whilst the ‘core business’ of industrial drug metabolism and pharmacokinetics is unlikely to change, we may now be at the dawn of a new era in exploiting the ‘new knowledge’. Only the next 10 years will tell.
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References AMES,B. N., MCCANN, J., and YAMASKI, E., 1975, Methods for detecting carcinogens and mutagens with the Salmonella/mammalian microsome mutagenicity test. Mutation Research, 31, 347-364. R., 1992, Drug analysis in biological samples. A survey on validation ARNOUX, P., and MORRISON, approaches in chromatography. Xenobiotica (in press). BERESFORD, A. P., CASWELL, K., CHAMBERS, R., and KIRK,1. P., 1992, Advantages of a chiral H P L C as a preparative step for chiral analysis in biological samples: assays for GR92549 and GR50360 enantiomers using protein stationary phases. Xenobiotica (in press). CAPRIOLI, R. M., 1990, In Continuous Flow Fast Atom Bombardment Mass Spectrometry,edited by R. M . Caprioli (Chichester: John Wiley & Sons Ltd.), pp. 63-92. CASE,D. E., 1981, The State of the Art: a commentary on the current practice of metabolism and pharmacokinetic studies in the pharmaceutical industry. Xenobiotica, 11, 803-814. C. K., and CHABNER, B. A,, 1990, Pharmacology guided Phase I clinical COLLINS, J. M., GRIESHABER, trials based upon preclinical drug development. Journal ofthe National Cancer Institute, 82, 1321. OF THE EUROPEAN COMMUNITIES, 1989a, Guidelines on the Quality, Safety and Efficacy of COMMISSION Medicinal Products for Human Use. COMMISSION OF THE EUROPEAN, COMMUNITIES, 1989b, Notice to Applicants for Marketing Authorities for Proprietary Medicinal Products for Human Use in the Member States of the European Community. OF THE EUROPEAN COMMUNITIES, 1990a, CPMP Working Party on Safety of Medicinal COMMISSION Products: Note for Guidance. Recommendations for the Development of Non-clincal Testing Strategies (Draft No. 7). COMMISSION OF THE EUROPEAN COMMUNITIES, 1990b, CPMP Working Party in Efficacy of Medicinal Products: Note for Guidance. Investigation of Bioavailability and Bioequivalence (Draft). V. J., PIKE,V. W., BAILEY,D., FREEMANTLE, C. A. J., PAGE,B. C., JONES,A. K. P., CUNNINGHAM, D., LUTHRA, S. K., and JONES,T., 1991, A method for studying KENSETT,M. J., BATEMAN, pharmacokinetics in man at picomolar drug concentrations. British Journal of Clinical Pharmacology, 32, 167-1 72. DAVIFS,D . S., 1988, An introduction to metabolic and kinetic aspects of toxicological studies. Xenobiotic, 18 (Suppl. l ) , 3-7. DOYLE, E., HUBER,R., and PICOT, V. S., 1992, Direct injection/HPLC methods for the analysis of drugs in biological samples and their application to pharmacokinetic studies in the rat. Xenobiotica, 22, 765-774. DHSS, 1977, MAL-4: Notes on Application for Clinical Trial Certificates (Medicines for Human Use). GILBERT, P. J., HARTLEY, T . E., TROKE, J. A., TURCAN, R. G., VOSE,C. W., and WATSON, K . W., 1992. T h e application of "F-NMR spectroscopy to the identification of dog urinary metabolites of imirestat a spirohydantoin aldose reductase inhibitor. Xenobiotica, 22, 775-788. HEIM,M., and MEYER,U. A., 1991, Predicting debrisoquine phenotype. Lancet, 337, 363. S. L., MALONE-LEE, J., and HUGHES,K. M., LANG,J. C. T . , LAZARE, R., GORDON, D., STANTON, GERAINT, M., 1992, T h e measurement of oxybutynin and its N-desethyl metabolite in plasma and its application to pharmacokinetic studies in young volunteers and elderly and frail elderly patient volunteers. Xenobiotica, 22, 859-870. D. A., 1992, The role of metabolism and pharmacokinetic studies in the HUMPHREY, M., and SMITH, discovery of new drugs and their development-present and future perspectives. Xenobiotica, 22,743-756. HUTT,A. J., 1990, In Progress in Drug Metabolism, Vol. 12, edited by G . G. Gibson (London: Taylor and Francis), pp. 147-204. INGS,R. M. J., LUCAS,C., LIEVRE, E., ARELIET, C., CLAVEL, M., LEVYRAZ, S., MINAIDIS, D., LOKIEC, F., and TURPIN,F., 1992, The pharmacokinetics and pharmacodynamics of a new vinca alkaloid, S 12363, using data from a Phase I tolerance study in man. Xenobiotica, 22, 871-880. LAVRIJSEN, K., VAN HOUDT, J., VAN DYCK, D., HENDRICKX, J., BOCKX,M., HURKMANS, R., MEULDERMANS, W., LE JEUNE,L . LAUWERS, W., and HEYKANTS, J., 1992, Compariative metabolism of flunarizine in rats, dogs and man. An in vitro study with subcellular liver fractions and isolated hepatocytes. Xenobiotica, 22, 815-836. NEBERT, D. W., NELSON,D. R., COON,M. J., ESTABROOK, R. W., FEYEREISEN, R., FUJIIKURIYAMA, Y., GONZALEZ, F. J., GUENGERICH, F. P., GUNSALUS, I. C., JOHNSON, E. F., LOPER,V. C., SATO, R., WATERMAN, M. R., and WAXMAN, D. J., 1991, T h e P450 superfamily: update on new sequences, gene mapping and recommended nomenclature. DNA and Cell Biology, 10, 1-14. PIDGEN, A., 1992, Statistical aspects of bioequivalance-a review. Xenobiotica (in press). PREECE, N. E., and TIMBRELL, J. A,, 1990, In ProgressinDrug Metabolism, Vol. 12, edited by G. G. Gibson (London: Taylor and Francis), pp. 147-204. P. E., REIK, L. M., and LEVIN,W., 1982, Purification, characterisation and RYAN,D. E., THOMAS, regulation of five rat hepatic microsmal cytochrome P-450 isoenzymes. Xenobiotica, 12,727-744. F. P., 1989, Human liver microsomal SHIMADA, T., IWASAKA, M., MARTIN, M. V., and GUENGERICH, cytochrome P-450 enzymes involved in the bioactivation of procarcinogens detected by umu gene response in Salmonella typhimurium, TA1535/pSK1002. Cancer Research, 49, 3218-3228.
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TOMLINSON, P. W., RAMJI,J. V., and FILER,C. W., 1992, The disposition of the enantiomers of chromakalim in rat and cynmolgus monkey. Xenobiotica (in press). WATERMAN, M. R., 1982, ACTH-mediated induction of synthesis and activity of cytochrome P-450 and related enzymes in cultivated bovine adrenocortical cells. Xenobiotica, 12, 773-786. WILTSHIRE, H. R., HARRIS, S. R., PRIOR, K. J., KOZLOWSKI, U. M . and WORTH, E., 1992, The metabolism of the calcium antagonist Ro 40-5967:a case history of the use of diode array (UV spectroscopy and thermospray mass spectrometry in the elucidation of a complex metabolic pathway. Xenobiotica (in press).
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WOLF,C. R., Moss, J. E., MILES, J. S., GOUGH, A. C., and SPURR, N. K., 1990, Detection of debrisoquine phenotype. Lancet, 336, 1452-1453.
ISSN: 0049-8254 (Print) 1366-5928 (Online) Journal homepage: http://www.tandfonline.com/loi/ixen20
Twenty years on: a review of the current practice of drug metabolism and pharmacokinetic studies in the pharmaceutical industry J. A. Bell & M. H. Tarbit To cite this article: J. A. Bell & M. H. Tarbit (1992) Twenty years on: a review of the current practice of drug metabolism and pharmacokinetic studies in the pharmaceutical industry, Xenobiotica, 22:7, 735-742, DOI: 10.3109/00498259209053136 To link to this article: http://dx.doi.org/10.3109/00498259209053136
Published online: 22 Sep 2008.
Submit your article to this journal
Article views: 6
View related articles
Citing articles: 1 View citing articles
Full Terms & Conditions of access and use can be found at http://www.tandfonline.com/action/journalInformation?journalCode=ixen20 Download by: [ECU Libraries]
Date: 21 March 2016, At: 22:07
XENOBIOTICA,
1992, VOL. 22,
NO.
7, 735-742
Comment Twenty years on: a review of the current practice of drug metabolism and pharmacokinetic studies in the pharmaceutical industry
J. A. BELL and M . H. T A R B I T Drug Metabolism Division, Giaxo Group Research Limited, Park Road, Ware, Herts., UK
Downloaded by [ECU Libraries] at 22:07 21 March 2016
(Received 1 November 1991; accepted 20 December 1991)
Introduction T h e 10th anniversary of the Drug Metabolism Discussion Group* was marked by a special issue of Xenobiotica in which Dr David Case (1981) reviewed the then current state of the art of drug metabolism and pharmacokinetics within the pharmaceutical industry. Ten years on we have another celebratory issue of this Journal and a further opportunity to review our field. Rather than make any attempt to emulate Case’s excellent commentary, we will restrict our review to a consideration of the progress and changes that have occurred during the intervening years. Many of the papers presented in this issue provide a suitable barometer for assessing these changes. For simplicity and to avoid the semantic difficulties noted by Case, we will refer to drug metabolism and pharmacokinetics as DMPK.
Role of industrial drug metabolism and pharmacokinetics Although recognizing the potential contribution of D M P K in supporting compound selection and pharmacology studies in animals, Case’s commentary was very largely devoted to the role of D M P K in safety evaluation programmes. In 1981 this was an accurate reflection of the major focus of drug metabolism and pharmacokinetic effort within the industry. T h e last 10 years has seen a continuation in the application of D M P K to safety evaluation and, indeed, validation of toxicity studies has become the major raison d’2tre of pre-clinical drug metabolism. However, we believe that greater attention is now paid to the role of D M P K in drug discovery programmes than was the case in 1981. It is difficult to substantiate this assertion with hard facts: the commercial sensitivity of many drug discovery projects precludes early publication of D M P K input. Nevertheless, information imparted at informal meetings indicates that many companies have recognized that the skills-base residing in D M P K groups can be applied successfully to solve problems such as low potency and inappropriate duration that may be identified in secondary pharmacology screens. An extension of this’ application is that an early
*DMDG is an informal association of scientists engaged in the study of drug metabolism and pharmacokinetics in the pharmaceutical industry. 0049-8254/92 $3.00 0 1992 Taylor & Francis Ltd.
736
J . A. Bell and M . H . Tarbit
input from DMPK scientists can minimize the risk of selecting for development a drug candidate with inappropriate pharmacokinetics. This ‘added-value’ helps to reduce the timescale and hence cost of drug development programmes. These principles are exemplified in the review by Humphrey and Smith (1992). We perceive that an early input from D M P K scientists has also become the norm in clinical development. Currently it is now common for preliminary pharmacokinetic information to be obtained in the first study of safety and tolerability in healthy volunteers. Although definitive pharmacokinetic studies will follow, this early information can give valuable insights on how the new drug is handled in man. Further exploitation of D M P K science is likely to arise from the great explosion of knowledge in the area of drug metabolizing enzymes discussed below.
Downloaded by [ECU Libraries] at 22:07 21 March 2016
Regulatory requirements At first sight it would appear that there have been extensive changes in regulatory requirements during the last 10 years. Most obvious of these are in Europe where new guidelines covering pre-clinical and clinical D M P K prepared by the Committee for Proprietary Medicinal Products (CPMP) have been published by the Commission of the European Communities (Commission of the EC 1989a). On closer examination, however, these ‘multi-state’ guidelines differ little in content from their national predecessors, e.g. MAL-4 ( D H S S 1977). It is interesting to note that, with the exception of single dose studies, all the guidelines for toxicity tests refer to requirements for metabolic and kinetic data. This serves to illustrate the central role of D M P K in safety evaluation (Davies 1988) and has been further enhanced by the introduction of the ‘Expert Report’ system which requires applicants seeking marketing authority in the European Community to submit a critical evaluation of their product (Commission of the EC 1989b). There have been relatively few new guidelines from the FDA, although it is apparent from recent scientific meetings that this Agency is also now recognizing the importance of toxicokinetics in safety evaluation studies. In differentiating between toxicologists and kineticists Case (1981) noted that the latter
‘. . . responds to broad principles of regulatory concern, often wishes for more specific guidance but always wants to retain a large measure of flexibility in study design.’ As noted above, guidelines have not become more specific during the past 10 years and significant flexibility in the design of studies to support drug submissions remains. For example, many companies are now supplementing ‘classical’ drug metabolism studies by experiments carried out in vitro, although it is not clear whether the information obtained is used for regulatory purposes. In particular, the number of reports on the use of isolated hepatocytes (e.g. Lavrijsen et al. 1992) has increased dramatically over the past 10 years. Looking to the future, it appears that flexibility of study design will be enhanced further rather than reduced. A draft note for guidance from CPMP on non-clinical testing strategies (Commission of the EC 1990a) states:
‘. . . detailed, fixed experimental dosing programmes prescribing animal species, types of tests and duration of treatment, which would be valid for all types of medicinal products and all trial phases in humans (including early
Current practice of drug metabolism and pharmacokinetic studies
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pharmacokinetic studies in humans) cannot be scientifically justified. In order to avoid this type of rigid, dogmatic approach, only general guidance for the decision making processes regarding the need for experimental studies prior to human studies (and therapeutic use) can be given.’ Adoption of this ‘product-tailored’ approach in submissions to regulatory authorities has many attractive features such as an anticipated reduction in the number of experimental animals and the facilitation of global harmony in regulatory requirements. However, there are potential problems. During a drug development programme of say 10 years duration, decisions may be taken on the basis of scientific knowledge that could change significantly by the time the submission is reviewed. It appears, therefore, that joint agreement of scientific justification is likely to require effective communication between the applicant and regulatory authority at key stages throughout drug development. One aspect of D M P K where we have seen marked changes in regulatory expections is in the area of bioequivalence where a current CPMP draft note for guidance (Commission of the EC 1990b) contains considerably more detail than its predecessor (Commission of the EC 1989a). T h e review of statistical aspects of bioequivalence (Pidgen 1992) is therefore timely and validation of analytical methodology (Arnoux and Morrison 1992) is also receiving increased attention. In addition, the relationship between pharmacokinetics and pharmacodynamics (e.g. Ings et al. 1992) has been the subject of growing regulatory interest, particularly in the US, and there is an indication (e.g. Collins et al. 1990) that this area of research may be applied to the selection of clinical doses. T h e past 10 years has also seen an increase in pharmacokinetic studies in the elderly and in patients (e.g. Hughes et al. 1992). For the D M P K scientist perhaps the most significant change in the regulatory scene has been in the area of stereochemical drugs. Although some countries, e.g. Sweden and Switzerland, have produced discussion documents, as yet no regulatory authority has produced guidelines in this area. However, the CPMP Notes to Applicants (Cbmmission of the EC 1989b) states inter alia: ‘Possible problems relating to stereoisomerism, which should be discussed in the appropriate Expert Report and cross referenced, should include: -
pharmacokinetics (including information on the relative metabolism of the stereoisomers)
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extrapolation of the preclinical data (paying particular attention to possible problems relating to species differences in handling of the stereoisomers)’.
This development has had a significant impact in our field and has provided new challenges to the bioanalyst. I n fact it might be argued that the current interest in the pharmacokinetics of stereochemical drugs has arisen through the recent development of enantiospecific analytical methodology (Hutt 1990).
Tools of the trade Throughout the past 10 years the pharmaceutical industry has continued to develop increasingly potent drugs. In turn the D M P K scientist investigating the fate of these drugs and their metabolites in the body is faced with the need to develop increasingly sensitive bioanalytical techniques.
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Currently it appears that technology is keeping pace with demand and low concentration (nglml) assays are commonplace. For quantitative work h.p.1.c. remains the most widely used technique. A variety of detectors, e.g. UV, fluorescence and electrochemical (Hughes et al. 1992) are now used routinely in conjunction with diverse column packing materials. Although direct injection methods can often be employed (e.g. Doyle et al. 1992) the bioanalyst has at his disposal an extensive armoury of solid phases for the selective extraction of drugs from body fluids. Supercritical fluid chromatography (SFC) and capillary zone electrophoresis (CZE) both of which have been developed during the past decade are beginning to offer alternatives to h.p.1.c. Immunoassay has frequently been considered inferior to chromatography for the quantification of drugs in biological fluids, a reputation obtained largely through lack of specificity in the assay of steroidal drugs. However, the past 10 years have seen increased use of specific immunoassays in response to the problems of analysing very potent drugs. In addition, cross-reactivity between antisera and metabolites may be exploited in the development of selective extraction methods that rely on structural characteristics rather than physiochemical properties of the analyte. Cross-reactivity between analogues can also be exploited in drug discovery where antisera raised against a drug development candidate may often be useful in the assay of closelyrelated back-up compounds. T h e increased requirement for information on the pharmacokinetics of stereochemical drugs, discussed above, has led to considerable activity in the field of chiral analysis (e.g. Beresford et al. 1992, Tomlinson et al. 1992) and chiral disposition studies (Tomlinson et al. 1992). There have also been significant developments in the application of spcctrometric methods (Wiltshire et al. 1991). In the first of mass spectrometry rew sample introduction techniques have facilitated the interfacing of spectrometers with chromatographs. Of the ‘hyphenated’ techniques, only g.1.c.-mass spectrometry was widely used in D M P K laboratories 10 years ago. However, h.p.1.c.-mass spectrometry and more recently tandem mass spectrometery are now routinely applied to the identification of drug metabolites. Improvements in resolution, analytical power and sensitivity have also occurred in n.m.r., with its application to fluorinated compounds being very effective (e.g. Gilbert et al. 1992). Advances in multipulse sequences have enhanced the available information through the development of two-dimensional techniques, e.g. COSY, although it appears that the industrial drug metabolism scientist has been slow to exploit these major developments (Preece and Timbrel1 1990). T h e ‘microchip revolution’ has continued to provide opportunities for the automation of our function. Computers can now control information flow from robotic instrumentation through data acquisition and processing to submission of the dossier to regulatory authorities. Although this development enhances compliance with good laboratory practice (GLP) by reducing or removing error-prone manual data entry, further GLP problems are introduced by the requirement for validation of the systems. Progress towards the fully automated, GLP-compliant laboratory has therefore been slower than perhaps predicted 10 years ago. The methodology in routine use by industrial DMPK sections still requires us to analyse drug and metabolites in samples of biological fluids that have been removed from the body and hence the information is obtained some time after the event. However, the past 5 years have seen the emergence of dynamic techniques such as
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continuous flow fast atom bombardment mass spectrometry (CF-FAB) (Caprioli 1990) which in combination with microdialysis probes enable the acquisition of pharmacokinetic data in ‘real-time’. Techniques are also now available that allow n.m.r. to be used in whole animals (Preece and Timbrel1 1990) and further intriguing possibilities are presented by the application of positron-emitting isotopes to the investigation of clinical pharmacokinetics (Cunningham et al. 1991). In the future these exciting developments may present a significant business opportunity for industrial DMPK.
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New business for an old trade In many ways D M P K has grown as a synthesizing science aiding interpretation of toxicity tests and relationships to clinical use. In some areas of our business, however, this synthesis has failed to move the goalposts. A good example of this is genetic toxicology, a science fundamentally integrated with drug metabolism. Thus tests in vitro used to evaluate the genotoxicity of compounds and their metabolites were established on the drug metabolism science of the 1970s (Ames et al. 1975). Despite the huge growth in the understanding of the biology of drug-metabolizing enzymes over the intervening years they have essentially stayed that way. In many respects this is because the increased understanding of the role of enzymes in activiation has occurred primarily in academia and has been of limited interest to D M P K scientists in industry. We have therefore not been positioned to advise on the design of genetic toxicity tests, and communication between genetic toxicologists and drug metabolism scientists within the industry has been poor. We also have a poor record over the past 10 years in understanding how our drugs interact with drug-metabolizing enzymes, the mechanism of inhibition and induction, or even which enzymes are responsible for their metabolism or activation. This seems set to change. Like many areas of biological science the coming of the molecular biology ‘revolution’ has been fundamental in its impact on the enzymology of drug metabolism. No one 10 years ago could have foreseen the phenomenal rate at which the knowledge and understanding of cytochrome P-450 has moved, e.g. in 1982 about half a dozen P-450s had been isolated and identified in the various species examined (Ryan et al. 1982) and the wide role of the enzyme system in the regulation of steroidogenesis was just emerging (Waterman, 1982). Recently over 100 genes of P-450 in the plant and animal kingdom were classified (Nebert et al. 1991) with the high probability of discovering many m0r.e. T h e enzyme is now recognized as being essentially ubiquitous with multiple endogenous roles in addition to that of drug metabolism. Relatively under-researched systems, such as glucuronyl transferases, glutathione transferases and flavin monooxygenase, are now also assuming markedly higher profiles within the industry than 10 years ago, as the techniques of molecular biology allow more facile identification, isolation and characterization of their role in drug metabolism and detoxication. T h e impact of molecular biology techniques allied to the availability of human tissue has led to several established dogmas on the role of various enzymes in metabolism and activation of xenobiotics in animals and man being challenged. Thus recent studies have highlighted evidence that activation of a significant number of carcinogens and hepatotoxins is mediated via constitutively expressed enzymes in animals and man (Shimada et al. 1989), some of which are suppressed by the induction regimens regularly adopted for the metabolic activation systems used in genetic toxicity testing.
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In addition, we now have for the first time, opportunities via cloning and expression systems to examine the role of individual animal and human enzymes in the metabolism of our drugs in vitro-long before the compound reaches man. Several of the expression techniques in cell lines offer the hope, if not the certainty, that in the future, species validation of metabolism in vitro may be better established than it has been in the past. These techniques also provide the means to address issues such as genetic polymorphism of our drugs using simple tests in vitro without the need for costly clinical studies. In this area we may also see the introduction of identification of ‘poor metabolisers’ in the population by simple kinetic blood tests (Heim and Meyer 1991, Wolf et al. 1990), thus avoiding the need for expensive phenotyping studies. Currently it is difficult to appreciate whether the pharmaceutical industry regards these developments as an opportunity or a threat. T h e positive view of this explosion of knowledge and technique is that it provides scope to understand significantly more about the mechanisms and in the inter-relationships concerned in the processing of our drug candidates and their effects on the functions and expression of enzymes. This is surely important if we are to put context to the questions that face us such as the significance of enzyme induction by drugs in their long-term clinical benefitlrisk assessment. T h e one ‘isle of doubt in the sea of euphoria’ surrounding the new science is the apparent conservatism of the industrial D M P K scientist. Much of this research and technology still lies in academia. We in industrial drug metabolism have been somewhat slow to seize these opportunities despite the fact that molecular science has been embraced avidly by our pharmacology and biochemist research colleagues. Many pharmaceutical companies have molecular biology departments: few are functioning on behalf of their drug metabolism needs. We believe that we must be prepared to be proactive in seeking to establish where these techniques will work for us if we are to avoid the risk of these elements becoming ‘requirements’ before their significance is placed in context, or taking the form of uncertainty factors in the development of drugs. In this respect the context and timing of these studies in the development process should be the subject of wide and continuing debate within the industrial environment. Overall, we should aim to use the opportunities presented by these techniques to enable us to answer regulatory questions which are increasingly likely to become focused on how new chemical entities interact with drug metabolizing enzymes.
Conclusions Many of the aspects of our work discussed by Case (1981) have remained essentially unchanged and the past 10 years have represented a consolidation of the business of industrial drug metabolism and pharmacokinetics. Nevertheless, there have been considerable advances in technology to which D M P K scientists have responded rapidly and which have been exploited in the application of ‘classical studies’ to increasingly potent drugs. Response from industrial scientists to the huge growth in our understanding of drug metabolising enzymes has been much less obvious. Whilst the ‘core business’ of industrial drug metabolism and pharmacokinetics is unlikely to change, we may now be at the dawn of a new era in exploiting the ‘new knowledge’. Only the next 10 years will tell.
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