摘要:

Four non-allelic genes, which encode four different aldehyde dehydrogenase (ALDH) isozymes, have been cloned and characterized at the present time. The coding nucleotide sequences, and organization of introns and exons of these genes have been elucidated. TheALDH1gene encodes the major cytosolic ALDH1 existing in the liver and other tissues. The genetic deficiency of this isozyme was found at a low frequency (≪10%) in both Caucasians and Orientals. The deficiency and alcohol sensitivity character are inherited together in one large Caucasian family examined. TheALDH1gene contains two hormone response elements in its upstream 5‘ region. TheALDH2gene encodes the major liver mitochondrial ALDH2 which has a very low Km for acetaldehyde. The atypicalALDH22allele is common (about 30%) in Orientals; and subjects withALDH22allele, both homozygous and heterozygous status, lack ALDH activity. These individuals are alcohol sensitive and have a markedly reduced risk in developing alcoholism and alcoholic liver diseases. TheALDH3gene produces a cytosolic ALDH3 isozyme existing in the stomach and liver carcinoma but hardly in normal liver. TheALDH3locus is polymorphic in Orientals and presumably other populations. TheALDH5gene, which is expressed in testes and liver, is highly polymorphic in both Caucasians and Orientals. The variation of these two loci may affect the development of alcohol-related problems.

ISSN:0960-314X    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

Thiopurine methyltransferase (TPMT) catalyses theS-methylation of thiopurine drugs such as 6-mercaptopurine (6-MP). TPMT activity in the human red blood cell (RBC) is controlled by a common genetic polymorphism. Gene frequencies for this polymorphism are such that approximately one in 300 subjects is homozygous for the allele for low activity and lacks RBC TPMT activity, 11% of subjects are heterozygous and have intermediate levels of enzyme activity and 89% are homozygous for the allele for high activity. Our experiments were performed to determine whether the properties of TPMT in an important human drug metabolizing organ, the liver, were similar to those of RBC TPMT and to test the hypothesis that the genetic polymorphism which controls TPMT activity in the human RBC might also regulate the level of this enzyme activity in hepatic tissue. Human liver TPMT is a cytoplasmic enzyme and the Kmvalues for 6-MP andS-adenosyl-L-methionine, cosubstrates for the reaction, were 580 μM and 2.7 μM, respectively. These properties, as well as the sensitivity of human liver TPMT to a panel of methyltransferase inhibitors, were similar to those of RBC TPMT. The enzyme activity was then measured in 119 surgical biopsy samples of hepatic tissue. Average hepatic TPMT activity was 13.6% higher in samples from male than in those from female patients. Frequency distribution histograms demonstrated the presence of a subgroup with intermediate enzyme activity that included 8.4% of samples. In addition, when TPMT activity was measured in both RBCs and hepatic tissue for 35 patients, those with inherited intermediate levels of RBC TPMT activity also had intermediate hepatic enzyme activity. Finally, ion exchange chromatography demonstrated the presence of two isozymes of TPMT in human hepatic tissue, but the isozymes did not appear to explain the molecular mechanism responsible for the genetic polymorphism. These results were compatible with the conclusion that the genetic polymorphism which controls TPMT activity in the RBC also controls levels of this important enzyme activity in a major human drug metabolizing organ, the liver.

ISSN:0960-314X    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

Male and female patas (Erythrocebus patas) and cynomolgus (Macaca fascicularis) monkeys were treated with phenobarbitone (PB) and examined for the induction of various cytochrome P450 (P450)-mediated drug metabolizing enzymes. Hydroxylation of testosterone at the 6β, 2β, and 15β positions, metabolites normally associated with CYP3A P450s increased 2− to 5-fold in PB-treated animals. Induction of this P450 family was confirmed by the use of polyclonal antisera directed against the human CYP3A enzymes which inhibited both induced and constitutive 6β – and 15β -hydroxylase activities in both species of monkeys. The enzymatic activities testosterone 16β -hydroxylation, pentoxyresorufinO-dealkylation [PROD], and benzyloxyresorufinO-dealkylation [BZROD] typically associated with the rodent CYP2B subfamily in rodents were also examined. Testosterone 16β -hydroxylation activity was highly induced up to 15-fold in both species of monkeys to maximal levels similar to those induced in PB-treated male rats. BZROD was similarly induced up to 10-fold in both species of monkeys, but the maximal levels of BZROD achieved were substantially lower than those obtained in PB-treated rats. Finally, PROD yielded an idiosyncratic response showing substantial induction (> 30-fold) in certain patas monkeys (4 out of 8) but minimal (< 5-fold) induction in the other patas monkeys (4 out of 8) or any of the cynomolgus monkeys (0 out of 8). Immunodetection of various cytochromes using polyclonal antisera directed against rat cytochromes confirmed the induction of CYP3A proteins as well as the induction of protein(s) immunologically cross-reactive with rat CYP2B P450s. BZROD, PROD, or testosterone 16β-hydroxylase activities, were not inhibited by antibody to CYP3A P450s. However, high concentrations of polyclonal antiserum directed against rat CYP2B inhibited all three activities in PB-induced patas monkeys. In contrast, this antiserum failed to inhibit the hydroxylation of testosterone at the 6β or 2β positions. Induction of the CYP2C subfamily was observed by immunochemical detection. Interestingly, induction of this subfamily appeared to be more pronounced in cynomolgus than in patas monkeys. Finally, we failed to observe significant sex-dependent differences in P450-mediated enzymatic activities in either control or induced monkeys. These results confirm the induction of a similar spectrum of P450 proteins by PB in non-human primates to that which has previously been observed in rodents.

ISSN:0960-314X    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

The molecular basis for the use of caffeine (CA; 1,3,7-trimethyIxanthine) as a probe for specific human cytochromes P450 has been investigated. The CA 1-, 3− and 7-demethylations (to form theobromine, paraxanthine and theophylline, respectively) all followed biphasic kinetics in human liver microsomes. Mean apparent Kmvalues for the high- and low-affinity components of the demethylations ranged from 0.13–0.31 mM and 19.2–30.0 mM, respectively. cDNA-expressed CYP1A2 catalysed all three CA demethylations, and the apparent Kmfor CA 3-demethylation (the major metabolic pathway in humans) by the expressed enzyme was similar to the Kmfor the high-affinity liver microsomal CA 3-demethylase. IC50values for inhibition of the CA demethylations by a-naphthoflavone were similar for both expressed CYP1A2 and the high-affinity microsomal demethylases. Moreover, CA was a competitive inhibitor of expressed CYP1A2 catalysed phenacetin 0-deethylation, with the apparent K1(0.080 mM) closely matching the apparent Km(0.082 mM) for CA 3-demethylation by the expressed enzyme. Expressed CYP1A1 was additionally shown to catalyse the 3-demethylation of CA, although activity was lower than that observed for CYP1A2. While these data indicate that CYP1A2 is responsible for the high-affinity component of human liver CA 3-demethylation, two limitations associated with the use of CA as anin vitroprobe for CYP1A2 activity have been identified: (i) CA 3-demethylation reflects hepatic CYP1A2 activity only at appropriately low substrate concentrations; and (ii) CA is a non-specific CYP1A substrate and CYP1A1 may therefore contribute to CA 3-demethylase activity in tissues in which it is expressed. An anti-CYP3A antibody essentially abolished the 8-hydroxylation of CA to form trimethyluric acid, suggesting formation of this metabolite may potentially serve as a marker of CYP3A isozyme(s) activity.

ISSN:0960-314X    

出版商:OVID    

年代:1992

数据来源: OVID