Pseudocholinesterase deficiency

The Great Paracelsus (Philipus Aureolus Theophrastus Bombastus von Hohenheim-Paracelsus, 1493-1541) is remembered by toxicologists for his famous quote: "All substances are poisons; there is none which is not a poison. The right dose differentiates a poison from a remedy." (1) Indeed, the basic principle from the quotation still applies today. The right dose of morphine that will produce a blood level of approximately 0.07 mg/L to 0.083 mg/L is essential for the relief of pain. An overdose that results in a markedly increased blood level (0.2 mg/L to 2.3 mg/L) can cause death due to central nervous system depression. (2) In general, the same applies to all chemical substances.

As an important exception to Paracelsus' statement, however, there are instances in which a drug should be avoided completely. An example is found in the administration of succinylcholine as a skeletal muscle relaxant in surgery patients. Most patients can convert succinylcholine into inert metabolites through an enzyme called pseudocholinesterase because most patients possess two alleles for the common gene that can produce active enzyme. A certain small percentage of patients, however, carries the genes that are aberrant or silent and, thus, either produce protein (enzyme) that is defective in its function or cannot produce the protein that is capable of inactivating administered succinylcholine at all. Thus, these patients expire or suffer severe sequelae from administration of the drug. (3)

Along the same line of thinking, the dose of a given drug for certain individuals may differ from that used for the majority of the population due to genetic differences between the individual and the general population (e.g., a reduced dose of 6-mercaptopurine in the presence of reduced thiopurine methyl transferase, or TPMT, activity due to a genetic deficiency in synthesis of the active enzyme). (4,5) Therefore, in the Third Millennium, an expansion of Paracelsus' statement might be that the "right dose" for one individual may not be the "right dose" or even the right drug for another, due to differences in each individual's genetic makeup. The determination of what is the "right dose" and, in some cases, what is even the right drug for a given individual constitutes the reason for the development of the sciences of pharmacogenetics and pharmacogenomics.

Although often used interchangeably, there are subtle differences between pharmacogenetics and pharmacogenomics. Pharmacogenetics focuses on individual traits with respect to one compound or drug. Thus, pharmacogenetics, which historically actually preceded pharmacogenomics, looks at the responses of different individuals to one drug, while pharmacogenomics studies the differences among several compounds with regard to a single genome. (6,7) Pharmacogenomics is concerned with the systematic assessment of how chemical compounds (e.g., drugs) modify the overall expression pattern in certain tissues. Pharmacogenomics is not focused on the differences between individuals. Rather, pharmacogenomics focuses on differences among several drugs or compounds with regard to a generic set of expressed or nonexpressed genes. The focus in pharmacogenomics is on compound variability. For the purposes of this brief introduction, the two terms will be used interchangeably unless stated otherwise.

[FIGURE 1 OMITTED]

Before embarking upon a cursory journey through pharmacogenomics, a very brief discussion of DNA (deoxyribonucleic acid) and its products is in order. DNA is the genetic code that determines all of an individual's characteristics, including the synthesis of the proper proteins essential for life. The so-called "central dogma" of molecular biology is outlined in Figure 1. (8) DNA is a long chain of chemically linked (phosphate bond), single nucleotides. Although there is a great deal more detail to the system due to phenomena such as splice variants, (9) basically DNA is transcribed in the cell nucleus by an enzyme called RNA polymerase to yield messenger RNA, or mRNA. DNA sequences called promoters and enhancers are also usually required to initiate RNA synthesis. Silencers balance the effects of promoters and enhancers when synthesis is not required. A stop codon (a series of the nucleotides that tell the polymerase to stop transcription) also is required so that only the required RNA, not an almost infinitely long RNA, is produced. Regions of the DNA that do not code for amino acids are called introns. The introns are spliced out before the completed mRNA is capped (vide infra) and exported from the nucleus for translation. The regions of the DNA that do code for protein are called exons. RNA that has had the introns removed is capped on the 5-prime end with a special nucleotide called 7-methylguanosine and has a poly-adenosine (poly-A) "tail" added to the 3-prime end. The capped mRNA with its poly-A "tail" is then translated into protein in a cellular apparatus called a ribosome. (9)

Also required before pharmacogenomics can be discussed is a basic knowledge of the proteins that are the end product of the transcription of DNA and the translation of RNA. Essentially, a protein is a long chain of chemically linked (amide or peptide bond) amino acids. The sequence in the chain of amino acids is called the primary structure. The chain may form loops and/or helices (secondary structure), and the loops and helices may fold to form the tertiary structure. Further, the protein may exist by itself, associate with other protein chains like itself (e.g., dimerize to form an aggregate of two like chains), or associate with dissimilar protein chains to form the final, active structure, which is known as the quarternary structure. Further, a protein may be chemically and functionally changed (post-translational modification) by the addition of phosphate groups (phosphorylation), glucuronic acid groups (glucuronidation), or other groups, or by the addition or removal of amino acids or short stretches of amino acids called polypeptides. The study of proteins, along with the analysis by 2D gel and mass spectrometry and data analysis by bioinformatics, constitute the emerging field of clinical proteomics. (10,11)

Proteins can act as hormones (e.g., insulin, glucagons, chorionic gonadotrophin); enzymes, which are biological catalysts (e.g., lactate dehydrogenase, alkaline phosphatase, creatine kinase); structural components (e.g., troponin, collagens); receptors (e.g., opiate receptors, cholesterol receptors); and a plethora of other functions essential for life. When a protein is formed from DNA that was in the correct sequence, the DNA is correctly transcribed, and the RNA is correctly translated; a fully functional protein usually results. If the DNA code is incorrect (although exceptions exist here also) or there is a defect in the mechanism that creates the protein from the original DNA code, however, a partially functional protein, a nonfunctional protein, or even a protein that is deleterious to cell function may be produced. Gene duplication, where the gene is functional and codes for the correct protein, can result in the overproduction of protein. (12)

Also before pharmacogenomics can be discussed, it is helpful to review a few pharmacologic fundamentals. First, although it sounds obvious, before a drug can have any effect on an individual, the drug must somehow enter the individual's body. Entry can be accomplished through inhalation (e.g., a bronchodilator used for the treatment of asthma), absorption through the skin (e.g., a topical anesthetic such as cocaine or lidocaine) or mucous membranes (e.g., pilocarpine eye drops), parenterally (any number of drugs that may be delivered intravenously, intramuscularly, or subcutaneously), or, most commonly, orally (a large number of drugs, including such common substances as aspirin and acetaminophen) where the drug is absorbed from the gastrointestinal tract. Once inside an individual, a drug needs to be transported to the site where it will have its effect. A drug may have no action whatsoever (e.g., insulin), may act directly on a receptor to produce the desired effect (e.g., morphine for analgesia), or may require activation (e.g., the production of morphine from codeine for analgesia). Both before and after the desired effect has been produced, drugs may be excreted (multiple routes such as urine and bile) unchanged (e.g., free morphine), excreted as metabolites (e.g., the excretion of morphine glucuronide, which is an inactive metabolite of morphine), or excreted as a combination. (14) Proteins are essential to carry out all of the aforementioned steps in drug metabolism.

Proteins play an active role in the disposition of drugs and their metabolites (vide infra). Proteins in the intestinal enterocytes, such as breast cancer resistance protein (BCRP), multidrug resistance-associated protein (MRP2), and multidrug resistance protein (MDR1), are involved in the transport of xenobiotics into the intestinal lumen. (15) A member of the peptide transporter family such as PepT1 facilitates absorption from the gut lumen and tubular reabsorption in the kidney. Drugs such as valaciclovir, valganciclovir, and captopril are known to be transported by PepT1. OCTN2, which is part of the organic cation transporter family, is involved in both the efflux and influx of drugs such as quinidine and verapamil. Although numerous polymorphisms exist among the transporter proteins, their influence on pharmacogenomics is unclear and still in its infancy. (16)

A clear example of how a protein and a mutation of that protein can affect a drug's absorption, however, is found with the cardiac glycoside digoxin. P-glycoprotein, which is a membrane protein that functions as an exporter of xenobiotics from cells, is a product of the MDR1 gene. Although several models have been proposed for P-glycoprotein's action, basically P-glycoprotein acts to move xenobiotics from epithelial cells into the adjacent lumen. P-glycoprotein is found in numerous cells associated with excretory function. In the case of digoxin (and certain other drugs), reduced intestinal absorption of the drug can be associated with induction (increased amounts) of the enzyme or the C3435T (where the deoxyribonucleotide cytidine replaces the deoxyribonucleotide thymidine at position 3435 in the DNA sequence that codes P-glycoprotein) mutation of P-glycoprotein. Thus, the mutant form of the protein causes a lowered overall intestinal absorption of digoxin by excreting more back into the intestinal lumen than the wild-type protein. (17)

Once a drug has entered the bloodstream, it is transported to various parts of the body where the drug may be activated or inactivated by certain enzymes (vide infra) by a process commonly referred to as biotransformation--or metabolism; be excreted unchanged; interact with a receptor or other location where the desired (and, sometimes, undesired or side-effect) action(s) may take place; or be stored (e.g., the retention of [[DELTA].sup.9]-tetrahydrocannabinol or THC in body fat or lead in bone) for future uses such as those previously described. Many drugs and other xenobiotics express their pharmacodynamic action by interacting with a specific protein receptor. As an example, morphine acts at what are called [mu] receptors. Indeed, polymorphism is exhibited by the various opiate receptors. (18)

Perhaps the best-characterized and most extensively studied area in pharmacogenomics is biotransformation. Fundamentally, biotransformation can be divided into two areas--Phase I and Phase II. Both Phase I and Phase II are designed to make xenobiotics more polar and, thus, more water-soluble. By being more polar and more water-soluble, metabolites are more easily excreted into excretory fluids such as urine. Phase I reactions include hydrolysis, reduction, and oxidation. Phase I biotransformation may activate a drug (known in this case as a prodrug) into a biologically active form or may inactivate an active drug. An example of activation is seen with the conversion of Tegafur into the active anticancer agent 5-fluorouracil (5-FU). An example of deactivation is seen with the oxidation of ethanol into acetaldehyde by alcohol dehydrogenase and the further oxidation of acetaldehyde into acetate by aldehyde dehydrogenase. Phase II biotransformation may or may not be preceded by Phase I biotransformation. Phase II biotransformation reactions involve glucuronidation; sulfation; acetylation; methylation; conjugation with glutathione; and conjugation with amino acids such as glycine, taurine, and glutamic acid. (19)

Hydrolysis as a Phase I chemical breakdown pathway has been mentioned above in the case of pseudocholinesterase and its variants. Reduction of the drug 5-fluorouracil, which was discussed above under activation of Tegafur, shows polymorphism in the 5-FU reduction when 5-FU is metabolized (deactivated) to its reduction product, 5-fluorodihydrouracil, in rare individuals who are deficient in the enzyme dihydropyrimidine dehydrogenase (DPD). Individuals who are deficient in DPD show toxicity, which may be fatal, to bone marrow and intestines due to increased levels of 5-FU. (19)

[FIGURE 2 OMITTED]

Since a more polar (and, thus, more water-soluble) product usually results, oxidation is one of the most common metabolic pathways in mammals. Due to genetic polymorphism, the metabolism of the most routinely observed analyte in toxicology, ethanol or ethyl alcohol, actually is more complex than usually visualized by the simple pathway depicted in Figure 2.

The first enzyme in the oxidation of ethanol--alcohol dehydrogenase--is a zinc-containing dimer, which means that the functional enzyme consists of two protein chains and the element zinc. The subunits are designated [alpha], [beta], [gamma], [pi], [chi], or [sigma]. The subunits are encoded by six different genetic loci (ADH1A, ADH1B*1, ADH1C*1, and ADH4-ADH7--formerly ADH1 through ADH7, respectively). To further add to the complexity of the system, there are three allelic variants of the beta subunit designated [[beta].sub.1], [[beta].sub.2], and [[beta].sub.3] and two allelic variants of the gamma chain designated [[gamma].sub.1] and [[gamma].sub.2], giving rise to, respectively, ADH1B*1, ADH1B*2, ADH1B*3, ADH1C*1, and ADH1C*2. The nine subunits of ADH can combine to form homodimers (i.e., both chains are identical). Further, the [alpha], [beta], and [gamma] chains and their allelic variants can form heterodimers (i.e., the two chains are different) with each other, but not with the other types of chains.

The different molecular forms of ADH are divided into four classes. Class 1 contains ADH1A, ADH1B*1, and ADH1C*1, which can be considered isozymes. ADH1A contains either two alpha subunits or one alpha plus one beta or gamma subunit. ADH1B*1 contains either two beta subunits, which could be [[beta].sub.1], [[beta].sub.2], or [[beta].sub.3] or a beta subunit plus a gamma subunit, which could be [[gamma].sub.1] or [[gamma].sub.2]. ADH1C*1 contains two gamma subunits that could be [[gamma].sub.1] or [[gamma].sub.2]. ADH1B enzymes that differ in the type of [beta] subunit are known as allelozymes, as are ADH1C enzymes that differ in the type of [gamma] subunit. Accordingly, ADH1B*1 is an allelozyme composed of [[beta].sub.1] subunits, ADH1B*2 is an allelozyme composed of [[beta].sub.2] subunits, and ADH1B*3 is an allelozyme composed of [[beta].sub.3] subunits. Class II contains ADH4, which is made up of two [pi] subunits. Class III contains ADH5, which is made up of two [chi] subunits. Class IV contains ADH7, which is made up of two [sigma] subunits.

It is the Class I ADH isozymes that are of the most interest to the practicing toxicologist, as it is these isozymes that are involved in the oxidation of ethanol and methanol. ADH1B*2 is an atypical isozyme that is responsible for an unusually rapid conversion of ethanol into acetaldehyde in 90% of the Pacific Rim Asian population, but is expressed to a lesser degree in Caucasians, Native Americans, and Asian Indians. Aldehyde dehydrogenase (ALDH) oxidizes aldehydes (like acetaldehyde) to the corresponding carboxylic acid. Twelve ALDH genes (ALDH1 through ALDH10, SSDH, and MMSDH) have been identified in humans. ALDH2 is primarily responsible for oxidizing simple aldehydes like acetaldehyde. A genetic polymorphism for ALDH2 has been identified in humans. A high percentage of Japanese, Chinese, Koreans, Taiwanese, and Vietnamese populations are deficient in ALDH2 due to a point mutation (Gl[u.sub.487] [right arrow] Ly[s.sub.487]). This inactive allelic variant of ALDH2, known as ALDH2*2, is found in the same population that has a high incidence of the atypical form of ADH--ADH2*2, which means that these individuals rapidly convert ethanol to acetaldehyde, but only slowly convert acetaldehyde to acetic acid. As a result, many Asians experience a flushing syndrome after consuming alcohol. Thus, what is considered to be very simple by most toxicologists--especially those who deal with driving-under-the-influence forensic cases--actually can be quite rich in detail for certain individuals within a diverse array of ethnic groups. (20,21)

In addition to the above example of the oxidation of ethanol, numerous other oxidative pathways for xenobiotics exist in humans and other animals. Many instances of drug oxidation are the result of a group of enzymes known as CYPs (from CYtochrome P450, the 450 being derived from the cytochrome's maximal absorbance of light at 450 nm). CYPs are categorized according to amino acid sequence homology. CYPs that have less than 40% homology are placed in a different family (e.g., 1, 2, 3, and so on). CYPs that are 40% to 55% identical are assigned to different subfamilies (e.g., 1A, 1B, 1C, and so on). P450 enzymes that are more than 55% identical are classified as members of the same subfamily (e.g., 2B1, 2B2, 2B3). The P450 enzymes are expressed in numerous tissues, but are especially prevalent in liver. CYPs, which exist in mammalian physiology, are so numerous that their complete description is beyond the scope of this basic introduction. According to a recent survey regarding the top 10 pharmacogenomics tests important to human drug metabolism and of interest for test development, five CYPs are CYP2D6, 2C9, 2C19, 3A5, and 2B6. (22)

The final step in drug metabolism is elimination. As stated earlier, elimination can occur with the unchanged drug, a drug that has been subjected to Phase I metabolism, a drug that has been subjected to Phase II metabolism, or a combination. Also as briefly discussed above, the proteins involved in elimination can be subject to polymorphism and, thus, are involved in pharmacogenomics.

[FIGURE 3 OMITTED]

Glossary (11,23)

Allele. One or several forms of a gene of a single individual compared with other individuals. An allele is present on a specific site (genetic locus) of a chromosome controlling a particular characteristic and giving rise to noticeable hereditary difference. A single allele is inherited separately from maternal and paternal origin. Thus, with exception to unmatched sex chromosomes, every individual has two alleles for each gene.

Genotype. The precise genetic constitution (i.e., genomes, genes, or alleles at one locus or place) that determines the phenotype (observable characteristics) of an organism.

Heterozygote. An individual that carries a pair of different alleles of a particular gene (inherited from two parents) on each member of a pair of chromosomes.

Homozygote. An individual whose genotype has two identical alleles (each derived from one parent) at a given locus or place on a pair of homologous chromosomes.

Phenotype. The observable characteristics or physical appearance of an organism, resulting from and determined by its expressed genes.

Point mutation. A mutation in which one base in the DNA chain is substituted with another, resulting in a change in the resulting amino acid sequence (protein).

Polymorphism. "Many faces." The difference in genetic sequences among individuals, groups, or populations.

Wild-type. The allele, genotype, or phenotype that naturally occurs in the normal population or, in the case of microbes, the standard laboratory strain of a given organism.

Xenobiotic. A foreign chemical including drugs, industrial chemicals, pollutants, pyrolysis products in food, and toxins produced by plants and animals.

References and Suggested Further Reading

1. Gallo MA. History and Scope of Toxicology. In: Casarett & Doull's Toxicology. The Basic Science of Poisons, 6th ed. New York, NY: McGraw-Hill Medical Publishing Division; 2001.

2. Baselt RC, ed. Disposition of Toxic Drugs and Chemicals in Man, 7th ed. Foster City, CA: Biomedical Publications; 2004.

3. Moss DW, Henderson AR. Clinical Enzymology. In: Ashwood EA, Burtis CA, eds. Tietz Textbook of Clinical Chemistry, 3rd ed. Philadelphia, PA: W B Saunders Co.; 1999.

4. Innocenti F, Iyer L, Ratain MJ. Pharmacogenomics of Chemotherapeutic Agents in Cancer Treatment. In: Licinio J, Wong M-L, eds. Pharmacogenomics. The Search for Individualized Therapies. Weinheim (Germany): Wiley-VCH; 2002.

5. Weinshilboum R. Inheritance and Drug Response. N Engl J Med. 2003;348(6):529-537.

6. Lindpainter K. The Role of Pharmacogenomics in Drug Discovery and Therapeutics. In: Licinio J, Wong M-L, eds. Pharmacogenomics. The Search for Individualized Therapies. Weinheim (Germany): Wiley-VCH; 2002.

7. Ito RK, Demers LM. Pharmacogenomics and Pharmacogenetics: Future Role of Molecular Diagnostics in the Clinical Diagnostic Laboratory. Clin Chem. 2004;50(9):1526-1527.

8. Strachan T, Read AP. DNA Structure and Gene Expression. In: Human Molecular Genetics 3. London: Garland Science. Taylor & Francis Group; 2004.

9. Strachan T, Read AP. Human Gene Expression. In: Human Molecular Genetics 3, London: Garland Science. Taylor & Francis Group; 2004.

10. Proteins. In: Alberts B, Johnson A, Lewis J, Raff M, Roberts K, Walker P, eds. Molecular Biology of The Cell. London: Garland Science. Taylor & Francis Group; 2002.

11. Clarke W, Zhang Z, Chan DW. The Application of Clinical Proteomics to Cancer and other Diseases. Clin Chem Lab Med. 2003;41(12):1562-1570.

12. Dalen P, Dahl ML, Bernal-Ruiz ML, Nordin J, Bertilsson L. 10-Hydroxylation of Nortriptyline in White Persons with 0, 1, 2, 3, and 13 Functional CYP2D6 Genes. Clin Pharmacol Ther. 1998;63(4):444-452.

13. Steimer W, Xopf K, von Amelunxen S, et al. Allele-Specific Change of Concentration and Functional Gene Dose for the Prediction of Steady-State Serum Concentrations of Amitriptyline and Nortriptyline in CYP 2C19 and CYP2D6 Extensive and Intermediate Metabolizers. Clin Chem. 2004;0(9):1623-1633.

14. Rozman KK, Klaassen CD. Absorption, Distribution, and Excretion of Toxicants. In: Casarett & Doull's Toxicology. The Basic Science of Poisons. 6th ed. New York, NY: McGraw-Hill Medical Publishing Division; 2001.

15. Evans WE, McLeod HL. Pharmacogenomics--Drug Disposition, Drug Targets, and Side Effects. N Engl J Med. 2003;348(6):539-49.

16. Tirona RG, Kim RB. Pharmacogenomics of Drug Transporters. In: Licinio J, Wong M-L, eds. Pharmacogenomics. The Search for Individualized Therapies. Weinheim (Germany): Wiley-VCH; 2002.

17. Fromm MF, Eichelbaum M. The Pharmacogenomics of Human P-Glycoprotein. In: Licinio J, Wong M-L, eds. Pharmacogenomics. The Search for Individualized Therapies. Weinheim (Germany): Wiley-VCH; 2002.

18. Resine T. Pharmacogenomics of Opioid Systems. In: Licinio J, Wong M-L, eds. Pharmacogenomics. The Search for Individualized Therapies. Weinheim (Germany): Wiley-VCH; 2002.

19. Parkinson A. Biotransformation of Xenobiotics, Absorption, Distribution, and Excretion of Toxicants. In: Casarett & Doull's Toxicology. The Basic Science of Poisons. 6th ed. New York, NY: McGraw-Hill Medical Publishing Division; 2001.

20. Jones AW. Disposition and Fate of Ethanol in the Body. In: Garriott JC, ed. Medical-Legal Aspects of Alcohol, 4th ed. Tuscon, AZ: Lawyers & Judges Publishing Co.; 2003.

21. Hurley TD, Edenberg HJ, Li T-K. Pharmacogenomics of Alcoholism. In: Licinio J, Wong M-L, eds. Pharmacogenomics. The Search for Individualized Therapies. Weinheim (Germany): Wiley-VCH; 2002.

22. Auxter-Parham S. Bringing Pharmacogenomic Assays to Market: How Will the FDA Regulate These New Tests? Clin Lab News. 2004;30:1-7.

23. Ashwood EA, Burtis CA, eds. Tietz Textbook of Clinical Chemistry, 3rd ed. Philadelphia, PA: W B Saunders Co.; 1999.

24. Smeraldi E, Zanardi R, Benedetti F, Di Bella D, Perez J, Catalona M. Polymorphism with the Promoter of the Serotonin Transporter Gene and Antidepressant Efficacy of Fluvoxamine. Mol Psychiatry. 1998;3:508-511.

25. Kim DK, Lim S-W, Lee S, Sohn SE, Kim S, Hahn CG, Carroll BJ. Serotonin Transporter Gene Polymorphism and Antidepressant Response. NeuroReport. 2000;11(1):215-219.

26. Salee FR, DeVane CL, Ferrell RE. Fluoxetine-related Death in a Child with Cytochrome P450 2D6 Genetic Deficiency. J Child Adolesc Psychopharmacol. 2000;27:27-34.

27. Wong SHY, Wagner MA, Jentzen JM, Schur C, Bjerke J, Gock SB, Chang CJ. Pharmacogenomics as an Aspect of Molecular Autopsy for Forensic Pathology/Toxicology: Does Genotyping CYP 2D6 Serve as an Adjunct for Certifying Methadone Toxicity? J For Sci. 2003;48:1406-1415.

28. Zhang Y-H, Zhang M. A Dictionary of Gene Technology Terms New York, NY: Parthenon Publishing, 2001.

29. Jannetto PJ, Wong SHY, Gock S, Sahin E, Jentzen JM. Pharmacogenomics as an adjunct to forensic toxicology: Genotyping Oxycodone Cases for Cytochrome P450 (CYP) 2D6. J Anal Tox. 2002;26:438-477.

RELATED ARTICLE: Clinical application: Depression

In order to provide therapeutic efficacy, the tricyclic antidepressant nortriptyline must achieve serum levels of 5 ng/mL to 150 ng/mL. (23) Usually, a therapeutic level is achieved by adjustment of a standard dose. Nortriptyline is metabolized to its hydroxy metabolite, 10-hydroxynortriptyline, by CYP2D6. Individuals with different genotypes and multiple copies of CYP2D6 metabolize nortriptyline at markedly different rates. When poor metabolizers of debrisoquin (a drug used to determine an individual's CYP2D6 status) with no functional CYP2D6 gene, intermediate metabolizers with one functional CYP2D6 gene, extensive metabolizers with two functional CYP2D6 genes, ultra-rapid metabolizers with duplicated CYP2D6*2 genes, and one ultra-rapid metabolizer with 13 copies of the CYP2D6*2 gene are compared, the results are quite striking. On one end of the spectrum, the individuals with no functional copy of CYP2D6 have maximal levels of serum nortriptyline of 51 ng/mL to 71 ng/mL. On the other end of the spectrum, an individual with 13 copies of the CYP2D6*2 gene had a maximal serum level of only 13 ng/mL. Needless to say, this represents an outstanding example where a "standard dose" will not necessarily achieve the required serum level of the active drug and, thus, the desired results. (12) A recent study further substantiated the prediction of amitriptyline and nortriptyline concentrations based on CYP2D6*4, *10, and *41 genotyping. (13)

Selective serotonin reuptake inhibitors, such as fluvoxetine, paroxetine and fluoxetine, probably exert their action through the serotonin transporter protein. The serotonin transporter gene (5-HTT) shows several polymorphisms. One polymorphism is in the transcriptional control region upstream of the 5-HTT coding sequence. It is either a 44-base pair insertion (long variant) or deletion (short sequence). One group has reported that individuals homozygous for the long variant and heterozygous individuals respond better to fluvoxamine than do individuals who are homozygous for the short variant. (24) Interestingly, another group reported just the opposite. (25) The disparity between the studies is a reminder that other factors that may never be separated out affect genomic expression and phenotypic response.

RELATED ARTICLE: Clinical application: Forensics

A 9-year-old born with probable fetal alcohol syndrome is treated with a combination of methylphenidate, clonidine, and fluoxetine for multiple behavioral problems. Over a period of time, the individual is hospitalized in status epilepticus, followed by cardiac arrest, and expires. Based on the levels of fluoxetine and its metabolite norfluoxetine in the deceased's post-mortem blood, the adoptive parents are suspected of homicide. The remainder of the adopted children are removed from the household. Due to the vociferous claims of the adoptive parents that there was no foul play involved, the deceased individual is tested for genetic polymorphism. Indeed, a polymorphism in the CYP2D6, which resulted in the poor metabolism of fluoxetine, was discovered. Based on the results of the post-mortem genetic testing, the adoptive parents of the deceased were exonerated and reunited with the remainder of the children. (26)

From a recently published study of assessing pharmacogenomics as an adjunct of molecular autopsy for forensic pathology/toxicology, CYP2D6 was genotyped for methadone deaths. (27) One of the three methadone deaths with poor phenotype based on genotyping results was a 41-year-old female who was six months pregnant. She was diagnosed with heart murmur and rheumatoid arthritis and treated with methadone. In addition, tricyclic antidepressants were prescribed for her depression. After celebrating New Year's Eve with her husband, she was found unresponsive in the living room the following morning. Toxicology analysis showed: methadone, 0.7 mg/L, amitriptyline, 1.5 mg/L, and nortriptyline, 2.2 mg/L. Pharmacogenomics tests showed CYP2D6*4 homozgyous, corresponding to poor metabolizer as a result of deficient CYP2D6 enzyme. The lack of enzyme predisposed her to the inability to hydroxylate methadone, amitriptyline, and nortriptyline, resulting in an overall accumulation and drug toxicity. Death certification showed the cause of death to be mixed drug overdose, and manner of death, accident. (27)

By Robert M. White, Sr., PhD, and Steven H.Y. Wong, PhD

Robert M. White, Sr., PhD, is technical director at DSI Laboratories in Fort Myers, FL, and Steven H.Y. Wong, PhD, is professor of pathology and director of clinical chemistry/toxicology, TDM, pharmacogenomics, and proteomics at the Medical College of Wisconsin and scientific director of the Department of Toxicology, Milwaukee County Medical Examiners' Office, Milwaukee, WI.

COPYRIGHT 2005 Nelson Publishing
COPYRIGHT 2005 Gale Group