Genetic polymorphisms of flavin-containing monooxygenase (FMO)

Mammalian flavin-containing monooxygenase (FMO) exists as six gene families and metabolizes a plethora of drugs and xenobiotics. The major FMO in adult human liver, FMO3, is responsible for trimethylamine (TMA) N-oxygenation. A number of FMO3 mutant alleles have been described and associated with a disease termed trimethylaminuria (TMAU). The TMAU patient excretes large amounts of TMA in urine and sweat. A more recent ethnically related polymorphism in expression of the major FMO in lung, FMO2, has been described. All Caucasians and Asians genotyped to date are homozygous for a CAG --> TAG amber mutation resulting in a premature stop codon and a nonfunctional protein truncated at AA 472 (wildtype FMO2 is 535 AA). This allele has been designated hFMO2*2A. Twenty-six percent of individuals of African descent and 5% of Hispanics genotyped to date carry at least one allele coding for full-length FMO2 (hFMO2*1 allele). Preliminary evidence indicates that FMO2.1 is very active toward the S-oxygenation of low MW thioureas, including the lung toxicant ethylene thiourea. Polymorphic expression of functional FMO2 in the individuals of African and Hispanic descent may markedly influence drug metabolism and/or xenobiotic toxicity in the lung.     

Non-synonymous genetic variants of flavin-containing monooxygenase 3 (FMO3) in cynomolgus macaques

Polymorphic human flavin-containing monooxygenase (FMO) 3 is an important drug-metabolizing enzyme for nitrogen- or sulfur-containing compounds. Cynomolgus macaques, a non-human primate species widely used in drug metabolism studies, have corresponding FMO3 molecular and enzymatic similarities to humans; however, genetic polymorphisms have not been investigated in macaques. In this study, re-sequencing of FMO3 in 64 cynomolgus and 32 rhesus macaques found a total of 18 non-synonymous variants. Nine variants were unique to cynomolgus macaques, of which 4 (including Q506K) were found only in Indochinese, 4 (including V299I, E348H, and G530A) only in Indonesian lineages, and one was common. Other five variants (including S504T at >10% allele frequencies) were unique to rhesus macaques. By functional characterization using cynomolgus FMO3 proteins heterologously expressed in Escherichia coli, FMO3 R509H variant appeared to suppress methimazole and benzydamine S- or N-oxygenations. Seven variants showed substantially lower benzydamine N-oxygenation as compared with wild-type FMO3 protein. Further analysis indicated that two of these variants, FMO3 G530A and R417H, showed significantly lower benzydamine N-oxygenation in liver microsomes of the homozygotes as compared with wild-type animals. Therefore, inter-animal variability of FMO3-dependent drug metabolism is at least partly accounted for by genetic polymorphisms in cynomolgus and rhesus macaques, similar to humans.     

Differences in FMO2*1 allelic frequency between Hispanics of Puerto Rican and Mexican descent.

A polymorphism for the phase I drug-metabolizing enzyme, flavin-containing monooxygenase isoform 2 (FMO2), encoding either truncated inactive protein, FMO2X472 (FMO2.2A), or full-length active enzyme, FMO2Q472 (FMO2.1), is known and exhibits significant interethnic differences in allelic frequency. FMO2 is the major or sole FMO isoform expressed in the lung of most mammals, including nonhuman primates. To date, FMO2.1 has been found only in African-American and Hispanic populations, rendering individuals with this allele subject to drug metabolism that is potentially different from that of the general population. Approximately 26% of African-Americans (n = 180) possess the FMO2*1 allele. In preliminary studies, we initially estimated that 5% of Hispanics (n = 40) have the FMO2*1 allele, but access to large cohorts of individuals of defined national origin has allowed us to determine the occurrence among Mexican-American and Puerto Rican-American groups. We used allele-specific genotyping to detect FMO2*1 from 632 Hispanic individuals, including 280 individuals of Mexican origin and 327 individuals of Puerto Rican origin. Statistical analysis indicated that results from Mexican (five sample sources) and Puerto Rican (three sample sources) samples were consistent with the hypothesis of homogeneity within each group from different sources. Data were subsequently pooled across sources to test for evidence of a difference in occurrence of FMO2*1 between ethnic groups. There was strong evidence (p = 0.0066) that FMO2*1 is more common among Puerto Ricans (7%) than among individuals of Mexican descent (2%). The overall occurrence of FMO2*1 among Hispanics of all origins is estimated to be between 2 and 7%.     

Ethnic differences in human flavin-containing monooxygenase 2 (FMO2) polymorphisms: detection of expressed protein in African-Americans

The flavin-containing monooxygenases (FMOs) are a family of xenobiotic-metabolizing enzymes that are expressed in a species- and tissue-specific manner. FMO2 expression has been observed in pulmonary tissue from several species, but not human. Two human FMO2 point mutations have been reported: a cytosine to thymidine transition at position 1414 resulting in a premature stop codon and a thymidine insertion at position 1589 resulting in a frameshift. To define the frequency of these sequence variations and explore their significance, unrelated African-American, Caucasian, and Korean individuals were genotyped. In the African-American population tested (n = 180), the 1414C allele occurred at a 13% frequency; however, all of the tested Caucasians (n = 52) and Koreans (n = 100) were homozygous for the 1414T allele. The T1589 allele occurred at frequencies of 6.9 and 13.0% in African-Americans (n = 175) and Caucasians (n = 23), respectively, and appears to segregate with the 1414T allele. Thus, it would have no further impact on FMO2 activity. Western blot analysis of pulmonary microsomes failed to detect immunoreactive protein in 1414T homozygotes. A heterozygotic individual did exhibit a single band of the expected size, but no detectable FMO activity in the corresponding lung microsomes. Sequence analysis, however, was consistent with the 1414C allele encoding an active FMO2 enzyme. FMO2 mRNA expression was observed in most individuals, but failed to correlate with genotype or protein expression. In summary, functional FMO2 is expressed in only a small percentage of the overall population. However, in certain ethnic groups, active pulmonary FMO2 enzyme will be present in a significant number of individuals.     

Benzydamine N-oxidation as an index reaction reflecting FMO activity in human liver microsomes and impact of FMO3 polymorphisms on enzyme activity

AIMS: The role of flavin containing monooxygenases (FMO) on the disposition of many drugs has been insufficiently explored. In vitro and in vivo tests are required to study FMO activity in humans. Benzydamine (BZD) N-oxidation was evaluated as an index reaction for FMO as was the impact of genetic polymorphisms of FMO3 on activity. METHODS: BZD was incubated with human liver microsomes (HLM) and recombinant enzymes. Human liver samples were genotyped using PCR-RFLP. RESULTS: BZD N-oxide formation rates in HLM followed Michaelis-Menten kinetics (mean Km = 64.0 microM, mean Vmax = 6.9 nmol mg-1 protein min-1; n = 35). N-benzylimidazole, a nonspecific CYP inhibitor, and various CYP isoform selective inhibitors did not affect BZD N-oxidation. In contrast, formation of BZD N-oxide was almost abolished by heat treatment of microsomes in the absence of NADPH and strongly inhibited by methimazole, a competitive FMO inhibitor. Recombinant FMO3 and FMO1 (which is not expressed in human liver), but not FMO5, showed BZD N-oxidase activity. Respective Km values for FMO3 and FMO1 were 40.4 microM and 23.6 microM, and respective Vmax values for FMO3 and FMO1 were 29.1 and 40.8 nmol mg-1 protein min-1. Human liver samples (n = 35) were analysed for six known FMO3 polymorphisms. The variants I66M, P135L and E305X were not detected. Samples homozygous for the K158 variant showed significantly reduced Vmax values (median 2.7 nmol mg-1 protein min-1) compared to the carriers of at least one wild type allele (median 6.2 nmol mg-1 protein min-1) (P < 0.05, Mann-Whitney-U-test). The V257M and E308G substitutions had no effect on enzyme activity. CONCLUSIONS: BZD N-oxidation in human liver is mainly catalysed by FMO3 and enzyme activity is affected by FMO3 genotype. BZD may be used as a model substrate for human liver FMO3 activity in vitro and may be further developed as an in vivo probe reflecting FMO3 activity.     

OPRM1基因A118G多态性与吗啡使用剂量的meta分析

摘要: 目的 系统评价OPRM1 基因 A118G多态性与吗啡使用剂量之间的相关性。方法 检索PubMed和EMBASE数据库以评估OPRM1基因 A118G多态性与吗啡使用剂量相关性的病例-对照研究。采用Stata 12.0软件进行系统性评价。结果16篇文献纳入研究,合计共有4115名病例。Meta分析结果显示：OPRM1基因 A118G多态性突变野生杂合子(AG)吗啡的需要量比野生纯合子(AA)高(P=0.009),突变纯合子(GG)吗啡的需要量较纯合子(AA)更高(P = 0.000)。显性遗传模型（AG+GG）吗啡的需要量较野生纯合子（AA）高(P=0.000)。亚组分析中,对于种族,突变纯合子(GG)、野生杂合子(AG),和野生纯合子(AA),白种人群吗啡需求量均高于亚洲人群,且患者年龄和疼痛评分也是吗啡需求量的重要预测因子。结论 当前证据显示,在OPRM1 基因A118G多态性突变中,G等位基因携带者均需要较高剂量的吗啡, 且白种人群吗啡需求量均高于亚洲人群。     

Metabolism of the anti-tuberculosis drug ethionamide by mouse and human FMO1, FMO2 and FMO3 and mouse and human lung microsomes

Tuberculosis (TB) results from infection with Mycobacterium tuberculosis and remains endemic throughout the world with one-third of the world's population infected. The prevalence of multi-drug resistant strains necessitates the use of more toxic second-line drugs such as ethionamide (ETA), a pro-drug requiring bioactivation to exert toxicity. M. tuberculosis possesses a flavin monooxygenase (EtaA) that oxygenates ETA first to the sulfoxide and then to 2-ethyl-4-amidopyridine, presumably through a second oxygenation involving sulfinic acid. ETA is also a substrate for mammalian flavin-containing monooxygenases (FMOs). We examined activity of expressed human and mouse FMOs toward ETA, as well as liver and lung microsomes. All FMOs converted ETA to the S-oxide (ETASO), the first step in bioactivation. Compared to M. tuberculosis, the second S-oxygenation to the sulfinic acid is slow. Mouse liver and lung microsomes, as well as human lung microsomes from an individual expressing active FMO, oxygenated ETA in the same manner as expressed FMOs, confirming this reaction functions in the major target organs for therapeutics (lung) and toxicity (liver). Inhibition by thiourea, and lack of inhibition by SKF-525A, confirm ETASO formation is primarily via FMO, particularly in lung. ETASO production was attenuated in a concentration-dependent manner by glutathione. FMO3 in human liver may contribute to the toxicity and/or affect efficacy of ETA administration. Additionally, there may be therapeutic implications of efficacy and toxicity in human lung based on the FMO2 genetic polymorphism, though further studies are needed to confirm that suggestion.     

Effects of genetic polymorphism of CYP1A2 inducibility on the steady-state plasma concentrations of trazodone and its active metabolite m-chlorophenylpiperazine in depressed Japanese patients

The effects of a genetic polymorphism of inducibility of cytochrome P450 (CYP) 1A2 on the steady-state plasma concentrations of trazodone and its active metabolite, m-chlorophenylpiperazine, were studied in order to clarify if these steady-state plasma concentrations are dependent on the CYP1A2 polymorphism. Fifty-eight Japanese depressed patients received trazodone 150 mg/day at bedtime. The steady-state plasma concentrations of trazodone and m-chlorophenylpiperazine were measured in duplicate using high performance liquid chromatographic method, and were corrected to the mean body weight for analyses. A point mutation from guanine (wild type) to adenine (mutated type) at position -2964 in the 5'-flanking region of CYP1A2 gene was identified by polymerase chain reaction fragment length polymorphism method. The mean steady-state plasma concentration of trazodone, but not m-chlorophenylpiperazine was significantly (P<0.05) lower in smokers than in non-smokers. Twenty-two smokers had 16 homozygotes of the wild type allele, 5 heterozygotes of the wild type and mutated alleles, and one homozygote of the mutated allele. There was no significant difference in the mean steady-state plasma concentration of trazodone or m-chlorophenylpiperazine between smokers with no mutation and those with mutation, although one homozygote of the mutated allele had the highest steady-state plasma concentration of trazodone in smokers. The present study thus suggests that CYP1A2 polymorphism does not necessarily have predictive value of the steady-state plasma concentration of trazodone or m-chlorophenylpiperazine in most of the smokers treated with trazodone.     

Population-specific polymorphisms of the human FMO3 gene: significance for detoxication.

Flavin-containing monooxygenase form 3 (FMO3) is one of the major enzyme systems that protect humans from the potentially toxic properties of drugs and chemicals. FMO3 converts nucleophilic heteroatom-containing chemicals and endogenous materials to polar metabolites, which facilitates their elimination. For example, the tertiary amine trimethylamine is N-oxygenated by human FMO3 to trimethylamine N-oxide, and trimethylamine N-oxide is excreted in a detoxication and deoderation process. In normal humans, virtually all trimethylamine is metabolized to trimethylamine N-oxide. In a few humans, trimethylamine is not efficiently metabolized to trimethylamine N-oxide, and those individuals suffer from trimethylaminuria, or fishlike odor syndrome. Previously, we identified mutations of the FMO3 gene that cause trimethylaminuria. We now report two prevalent polymorphisms of this gene (K158E and V257M) that modulate the activity of human FMO3. These polymorphisms are widely distributed in Canadian and Australian white populations. In vitro analysis of wild-type and variant human FMO3 proteins expressed from the cDNA for the two naturally occurring polymorphisms showed differences in substrate affinities for nitrogen-containing substrates. Thus, for polymorphic forms of human FMO3, lower k(cat)/K(m) values for N-oxygenation of 10-(N, N-dimethylaminopentyl)-2-(trifluoromethyl) phenothiazine, trimethylamine, and tyramine were observed. On the basis of in vitro kinetic parameters, human FMO1 does not significantly contribute to human metabolism of trimethylamine or tyramine. The results imply that prevalent polymorphisms of the human FMO3 gene may contribute to low penetrance predispositions to diseases associated with adverse environmental exposures to heteroatom-containing chemicals, drugs, and endogenous amines.     

5-Lipoxygenase polymorphism and in-vivo response to leukotriene receptor antagonists

OBJECTIVE: To determine whether genetic polymorphisms of the core promoter region of the 5-lipoxygenase gene contribute to the clinical response to leukotriene receptor antagonists. METHODS: We retrospectively genotyped 52 asthmatics for mutations of this gene from four placebo-controlled studies measuring leukotriene receptor antagonist responses. All studies measured bronchodilator response, and bronchial hyperresponsiveness to adenosine monophosphate was measured in three studies ( n = 34). RESULTS: Of the 52 patients genotyped, 40 were homozygous wild type, 12 heterozygous, and none was homozygous mutant. There was no significant difference in any improvements conferred by leukotriene receptor antagonists versus placebo in the forced expiratory volume in 1 s (0.20 l for wild-type homozygotes and 0.01 l for heterozygotes), forced mid-expiratory flow rate (0.16 l/s and 0.14 l/s), peak expiratory flow rate (10 l/min and 29 l/min) and adenosine monophosphate 20% fall in forced expiratory volume in 1 s (2.8-fold shift and 2.3-fold shift) between the two genotypes. CONCLUSION: In our population, screening for this polymorphism as an aid to guiding treatment is probably not worthwhile. In addition, we found no difference between homozygous wild types and heterozygotes in terms of bronchodilator response or bronchial hyperresponsiveness with leukotriene receptor antagonists.     

Characterization of expressed full-length and truncated FMO2 from rhesus monkey.

Flavin-containing monooxygenase (FMO) metabolizes a wide variety of nitrogen, sulfur, and phosphorous-containing xenobiotics. FMO2 is highly expressed in the lung of most mammals examined, but the protein has only recently been detected in humans, presumably due to a premature stop codon at AA472 in most individuals. In this study, full-length (mFMO2-535) and 3'-truncated (mFMO2-471) monkey FMO2 protein, produced by cDNA-mediated baculovirus expression, were characterized and compared with baculovirus-expressed rabbit FMO2 (rFMO2-535). Although baculovirus-expressed mFMO2-535 had properties similar to FMO in monkey lung microsomes and had catalytic properties similar to rFMO2-535, the expressed proteins differed in a number of properties in S-oxidation assays. Both enzymes had the same pH optima (pH 9.5); however, mFMO2-535 quickly lost activity at higher pH values whereas rFMO2-535 retained the majority of its activity. Also, mFMO2-535 was significantly less stable at elevated temperatures and in the presence of cholic acid but had greater activity in the presence of magnesium. mFMO2-535 had higher apparent K(m) and V(max)/K(m) values than rFMO2-535 did in N-oxygenation assays. mFMO2-471 was correctly targeted to the membrane fraction, but N- and S-oxygenation was not detected. Since the AA sequence identity of mFMO2 and human FMO2 is 97%, our results with mFMO2-535 suggest that individuals carrying the allele encoding full-length FMO2 are likely to have in vivo FMO2 activity. Such activity could result in marked differences in the metabolism, efficacy, and/or toxicity of drugs and xenobiotics for which lung is a portal of entry or target organ.     

S-oxygenation of the thioether organophosphate insecticides phorate and disulfoton by human lung flavin-containing monooxygenase 2

Phorate and disulfoton are organophosphate insecticides containing three oxidizable sulfurs, including a thioether. Previous studies have shown that only the thioether is oxygenated by flavin-containing monooxygenase (FMO) and the sole product is the sulfoxide with no oxygenation to the sulfone. The major FMO in lung of most mammals, including non-human primates, is FMO2. The FMO2*2 allele, found in all Caucasians and Asians genotyped to date, codes for a truncated, non-functional, protein (FMO2.2A). Twenty-six percent of individuals of African descent and 5% of Hispanics have the FMO2*1 allele, coding for full-length, functional protein (FMO2.1). We have here demonstrated that the thioether-containing organophosphate insecticides, phorate and disulfoton, are substrates for expressed human FMO2.1 with Km of 57 and 32 microM, respectively. LC/MS confirmed the addition of oxygen and formation of a single polar metabolite for each chemical. MS/MS analysis confirmed the metabolites to be the respective sulfoxides. Co-incubations with glutathione did not reduce yield, suggesting they are not highly electrophilic. As the sulfoxide of phorate is a markedly less effective acetylcholinesterase inhibitor than the cytochrome P450 metabolites (oxon, oxon sulfoxide or oxon sulfone), humans possessing the FMO2*1 allele may be more resistant to organophosphate-mediated toxicity when pulmonary metabolism is an important route of exposure or disposition.     

Cytochrome P-450 enzymes and FMO3 contribute to the disposition of the antipsychotic drug perazine in vitro

RATIONALE: Perazine (PER) is a phenothiazine antipsychotic drug frequently used in Germany that undergoes extensive metabolism. OBJECTIVES AND METHODS: To anticipate metabolic drug interactions and to explore the relevance of polymorphisms of metabolic enzymes, perazine-N-demethylation and perazine-N-oxidation were investigated in vitro using human liver microsomes and cDNA expressed enzymes. RESULTS: CYP3A4 and CYP2C9 were identified as the major enzymes mediating PER-N-demethylation. At 10 microM PER, a concentration consistent with anticipated in vivo liver concentrations, CYP3A4 and CYP2C9 contributed 50% and 35%, respectively, to PER-N-demethylation. With increasing PER concentrations, contribution of CYP2C9 decreased and CYP3A4 became more important. In human liver microsomes, PER-N-demethylation was inhibited by ketoconazole (>40%) and sulfaphenazole (16%). Allelic variants of recombinant CYP2C9 showed differences in PER-N-demethylase activity. The wild type allele CYP2C9*1 was the most active variant. Maximal activities of CYP2C9*2 and CYP2C9*3 were 88% and 18%, respectively, compared to the wild type activity. Perazine-N-oxidation was mainly mediated by FMO3. In the absence of NADPH, heat treatment of microsomes abolished PER-N-oxidase activity. Methimazole inhibited PER-N-oxidation, while CYP specific inhibitors had no inhibitory effect. Perazine is a potent inhibitor of dextromethorphan-O-demethylase, S-mephenytoin-hydroxylase, alprazolam-4-hydroxylase, phenacetin-O-deethylase and tolbutamide-hydroxylase activity in human liver microsomes. CONCLUSIONS: Alterations in the activity of CYP3A4, CYP2C9 and FMO3 through genetic polymorphisms, enzyme induction or inhibition bear the potential to cause clinically significant changes in perazine clearance. PER may alter the clearance of coadministered compounds metabolized by CYP2D6, CYP2C19, CYP2C9, CYP3A4 and CYP1A2.     

Effects of Genetic Polymorphism of CYP1A2 Inducibility on the Steady‐State Plasma Concentrations of Trazodone and its Active Metabolite m‐Chlorophenylpiperazine in Depressed Japanese Patients

Abstract: The effects of a genetic polymorphism of inducibility of cytochrome P450 (CYP) 1A2 on the steady‐state plasma concentrations of trazodone and its active metabolite, m‐chlorophenylpiperazine, were studied in order to clarify if these steady‐state plasma concentrations are dependent on the CYP1A2 polymorphism. Fifty‐eight Japanese depressed patients received trazodone 150 mg/day at bedtime. The steady‐state plasma concentrations of trazodone and m‐chlorophenylpiperazine were measured in duplicate using high performance liquid chromatographic method, and were corrected to the mean body weight for analyses. A point mutation from guanine (wild type) to adenine (mutated type) at position ?2964 in the 5′‐flanking region of CYP1A2 gene was identified by polymerase chain reaction fragment length polymorphism method. The mean steady‐state plasma concentration of trazodone, but not m‐chlorophenylpiperazine was significantly (P<0.05) lower in smokers than in non‐smokers. Twenty‐two smokers had 16 homozygotes of the wild type allele, 5 heterozygotes of the wild type and mutated alleles, and one homozygote of the mutated allele. There was no significant difference in the mean steady‐state plasma concentration of trazodone or m‐chlorophenylpiperazine between smokers with no mutation and those with mutation, although one homozygote of the mutated allele had the highest steady‐state plasma concentration of trazodone in smokers. The present study thus suggests that CYP1A2 polymorphism does not necessarily have predictive value of the steady‐state plasma concentration of trazodone or m‐chlorophenylpiperazine in most of the smokers treated with trazodone.     

Evaluation of xenobiotic N- and S-oxidation by variant flavin-containing monooxygenase 1 (FMO1) enzymes.

The flavin-containing monooxygenase gene family (FMO1-6) in humans encodes five functional isoforms that catalyze the monooxygenation of numerous N-, P- and S-containing drugs and toxicants. A previous single nucleotide polymorphism (SNP) analysis of FMO1 in African-Americans identified seven novel SNPs. To determine the functional relevance of the coding FMO1 variants (H97Q, I303V, I303T, R502X), they were heterologously expressed using a baculovirus system. Catalytic efficiency and stereoselectivity of N- and S-oxygenation was determined in the FMO1 variants using several substrates. The I303V variant showed catalytic constants equal to wild-type FMO1 for methimazole and methyl p-tolyl sulfide. Catalytic efficiency (V(max)/K(m)) of methyl p-tolyl sulfide oxidation by R502X was unaltered. In contrast, methimazole oxidation by R502X was not detected. Both H97Q and I303T had elevated catalytic efficiency with regards to methyl p-tolyl sulfide (162% and 212%, respectively), but slightly reduced efficiency with regards to methimazole (81% and 78%). All the variants demonstrated the same stereoselectivity for methyl p-tolyl sulfide oxidation as wild-type FMO1. FMO1 also metabolized the commonly used insecticide fenthion to its (+)-sulfoxide, with relatively high catalytic efficiency. FMO3 metabolized fenthion to its sulfoxide at a lower catalytic efficiency than FMO1 (27%) and with less stereoselectivity (74% (+)-sulfoxide). Racemic fenthion sulfoxide was a weaker inhibitor of acetylcholinesterase than its parent compound (IC(50) 0.26 and 0.015 mM, respectively). The (+)- and (-)-sulfoxides were equally potent inhibitors of acetylcholinesterase. These data indicate that all the currently known FMO1 variants are catalytically active, but alterations in kinetic parameters were observed.     

细胞色素P450 CYP2C9基因多态性对甲苯磺丁脲代谢动力学的影响

目的研究细胞色素P450 CYP2C9基因多态性对甲苯磺丁脲代谢动力学的影响。方法用基因芯片对137名健康志愿者进行CYP2C9基因多态性检测，将受试者分为CYP2C9野生型、杂合突变型和纯合突变型3组，用高效液相色谱法检测甲苯磺丁脲在受试者体内的药物代谢动力学参数，统计分析各组间药代动力学性质差异。结果在137名受试者中发现了9个CYP2C9*1/*3杂合型突变体和1个CYP2C9*3/*3纯合型突变体，其余为野生型个体。将9名CYP2C9*1/*3，1名CYP2C9*3/*3以及随机抽取的10名野生型个体分组，以甲苯磺丁脲为探药进行药物代谢动力学研究。结果在杂合型突变个体组以及纯合型突变个体组中，甲苯磺丁脲的代谢率显著低于对照的野生型个体组。结论CYP2C9基因多态性对甲苯磺丁脲代谢具有显著影响并呈基因剂量效应，检测突变型个体对指导临床合理用药和个体化医疗具有重要意义。     

The frequency and distribution of thiopurine methyltransferase alleles in Caucasian and Asian populations

Thiopurine methyltransferase metabolizes 6-mercaptopurine, thioguanine and azathioprine, thereby regulating cytotoxicity and clinical response to these thiopurine drugs. In healthy Caucasian populations, 89-94% of individuals have high thiopurine methyltransferase activity, 6-11% intermediate and 0.3% low, resulting from genetic polymorphism. Four variant thiopurine methyltransferase alleles were detected in over 80% of individuals with low or intermediate thiopurine methyltransferase activity. The wild-type allele is defined as TPMT*1 and the mutant alleles are TPMT*2 (G238C), TPMT*3A (G460A and A719G), TPMT*3B (G460A) and TPMT*3B (A719G). The frequency of these alleles in different ethnic groups is not well defined. In this study, DNA from 199 British Caucasian, 99 British South West Asian and 192 Chinese individuals was analysed for the presence of these variant alleles using polymerase chain reaction-restriction fragment length polymorphism and allele-specific polymerase chain reaction based assays. The frequency of individuals with a variant thiopurine methyltransferase genotype was: Caucasians 10.1% (20/199), South West Asians 2.0% (2/99) and Chinese 4.7% (9/192). Two TPMT*2 heterozygotes were identified in the Caucasian population, but this allele was not found in the two Asian populations. TPMT*3A was the only mutant allele found in the South West Asians (two heterozygotes). This was also the most common mutant allele in the Caucasians (16 heterozygotes and one homozygote) but was not found in the Chinese. All mutant alleles identified in the Chinese population were TPMT*3C (nine heterozygotes). This allele was found at a low frequency in the Caucasians (one heterozygote). This suggests that A719G is the oldest mutation, with G460A being acquired later to form the TPMT*3A allele in the Caucasian and South West Asian populations. TPMT*2 appears to be a more recent allele, which has only been detected in Caucasians to date. These ethnic differences may be important in the clinical use of thiopurine drugs.