INTERPRETATION OF POSTMORTEM TOXICOLOGY RESULTS Pharmacogenetics and Drug-Alcohol Interaction Anna Koski Department of Forensic Medicine University of Helsinki Finland ACADEMIC DISSERTATION To be publicly discussed, with the permission of the Medical Faculty of the University of Helsinki, in the auditorium of the Department of Forensic Medicine on September 23rd 2005, at 12 noon. Helsinki 2005 SUPERVISORS Professor Antti Sajantila Department of Forensic Medicine University of Helsinki Helsinki, Finland Docent Ilkka Ojanperä Department of Forensic Medicine University of Helsinki Helsinki, Finland REVIEWERS Docent Eero Mervaala Institute of Biomedicine University of Helsinki Helsinki, Finland Docent Kari Poikolainen Finnish Foundation for Alcohol Studies National Research and Development Centre for Welfare and Health Helsinki, Finland OPPONENT Professor Jørg Mørland Division of Forensic Toxicology and Drug Abuse Norwegian Institute of Public Health Oslo, Norway ISBN 952-91-9214-2 (paperback) ISBN 952-10-2662-6 (pdf) http://ethesis.helsinki.fi Helsinki University Printing House Helsinki 2005 Obtaining a quantitative result is not the endpoint of the analytical process. Irving Sunshine CONTENTS ABBREVIATIONS......................................................................................................................... 6 LIST OF ORIGINAL PUBLICATIONS .............................................................................................. 7 ABSTRACT.................................................................................................................................. 8 INTRODUCTION........................................................................................................................... 9 REVIEW OF THE LITERATURE ................................................................................................... 10 1 Interpretation of Postmortem Forensic Toxicology Results................................................ 10 1.1 Blood Samples .............................................................................................................. 10 1.1.1 Toxicological Tables ............................................................................................ 11 1.2 Other Matrices .............................................................................................................. 12 1.3 Postmortem Changes .................................................................................................... 12 1.4 Metabolites.................................................................................................................... 13 1.5 Alcohol Toxicity........................................................................................................... 13 1.6 Drug Toxicity................................................................................................................ 13 1.6.1 Fatal Toxicity Index ............................................................................................. 14 1.6.2 Other Measures of Toxicity.................................................................................. 15 1.6.3 Sources of Bias in Toxicity Indices...................................................................... 15 2 Pharmacogenetics................................................................................................................ 16 2.1 Drug-Metabolizing Enzymes........................................................................................ 16 2.1.1 Cytochrome P450 System .................................................................................... 16 CYP2D6 ............................................................................................................... 17 CYP2C19 ............................................................................................................. 19 2.2 Studies on Drug Metabolism ........................................................................................ 19 2.2.1 Tramadol Metabolism .......................................................................................... 19 2.2.2 Amitriptyline Metabolism.................................................................................... 20 2.3 Clinical Pharmacogenetics............................................................................................ 21 2.4 Postmortem Pharmacogenetics..................................................................................... 22 3 Drug-Alcohol Interaction .................................................................................................... 23 3.1 Alcohol Effects and Anesthetic Action ........................................................................ 23 3.1.1 γ-Aminobutyric Acid Receptor Type A ............................................................... 24 3.2 Animal Studies.............................................................................................................. 24 3.3 Postmortem Studies ...................................................................................................... 25 AIMS OF THE STUDY................................................................................................................. 26 4 CONTENTS MATERIALS AND METHODS...................................................................................................... 27 1 Autopsy cases ...................................................................................................................... 27 1.1 Blood Samples .............................................................................................................. 27 1.2 Database........................................................................................................................ 27 2 Analysis of Drug Concentrations ........................................................................................ 27 2.1 Screening....................................................................................................................... 27 2.2 Metabolite Analysis ...................................................................................................... 28 3 Genotyping .......................................................................................................................... 28 3.1 Long Polymerase Chain Reaction................................................................................. 29 3.2 Restriction Fragment Length Polymorphism Analysis................................................. 29 3.3 Multiplex Single-Base Extension Reaction..................................................................... 29 4 Case Selection Criteria ........................................................................................................ 29 5 Statistical Methods .............................................................................................................. 30 RESULTS................................................................................................................................... 31 1 Pharmacogenetics ................................................................................................................ 31 1.1 CYP2D6 and Tramadol (I) ............................................................................................ 31 1.2 CYP2D6 and Amitriptyline (II)..................................................................................... 31 1.3 CYP2C19 and Amitriptyline (II)................................................................................... 32 1.4 Allele and Genotype Frequencies (I, II)........................................................................ 33 2 Fatal Toxicity Indices (IV, V) ............................................................................................. 33 3 Drug-Alcohol Interaction..................................................................................................... 34 3.1 Alcohol and Benzodiazepines (III) ............................................................................... 34 3.2 Alcohol and Other Common Drugs (IV-VI)..................................................................... 34 DISCUSSION.............................................................................................................................. 38 1 Methodological Considerations........................................................................................... 38 2 Pharmacogenetics ................................................................................................................ 39 3 Drug-Alcohol Interaction..................................................................................................... 41 3.1 Alcohol and Benzodiazepines....................................................................................... 41 3.2 Alcohol and Other Common Drugs .............................................................................. 41 4 Drug Safety.......................................................................................................................... 42 4.1 Newer Antidepressants ................................................................................................. 42 5 Implications for Interpretation............................................................................................. 43 CONCLUSIONS .......................................................................................................................... 45 ACKNOWLEDGMENTS ..............................................................................................................