Arzu Gunes

Variation in CYP1A2 activity and its clinical implications: influence of environmental factors and genetic polymorphisms

CYP1A2 is involved in the metabolism of several widely used drugs and endogenous compounds, and in the activation of procarcinogens. Both genetic and environmental factors influence the activity of this enzyme. The current knowledge regarding factors influencing the activity of CYP1A2 is summarized in this review. Substrates, inhibitors and inducers of CYP1A2 activity, as well as phenotyping probes, are discussed. The functional significance and clinical importance of CYP1A2 gene polymorphisms are reviewed and interethnic differences in the distribution of CYP1A2 variant alleles and haplotypes are summarized. Finally, future perspectives for the possible clinical applications of CYP1A2 genotyping are discussed.

Influence of genetic polymorphisms, smoking, gender and age on CYP1A2 activity in a Turkish population

AIMS To study the variation in CYP1A2 activity in relation to smoking, gender, age and CYP1A2 polymorphisms. MATERIALS & METHODS CYP1A2 activity was determined by plasma paraxanthine:caffeine ratio (17X:137X) 4 h after the intake of a standardized cup of coffee in 146 Turkish healthy volunteers. Seven CYP1A2 polymorphisms (-3860G>A, -3113G>A, -2467del/T, -739T>G, -729C>T, -163C>A and 5347T>C) were analyzed. RESULTS The 17X:137X ratios were increased in smokers (p < 0.0001) and tended to be higher in men both among nonsmokers (p = 0.051) and smokers (p = 0.064). Age-related differences were observed only among nonsmoking women (p = 0.024). The -163C>A polymorphism correlated with 17X:137X ratios only in smokers (p = 0.006). Furthermore, increased 17X:137X ratios were observed in CYP1A2 haplotype H4 (-3860G, -3113G, -2467del, -739T, -729C, -163A and 5347T) carriers in the overall study population (p = 0.026). Multiple regression analyses including smoking, gender, -163C>A genotype and age revealed a significant influence of smoking (p < 0.0001) and gender (p = 0.002) in the overall study population. However, in nonsmokers only the influence of gender remained significant (p = 0.021), while in smokers the influence of the -163C>A genotype held the statistical significance (p = 0.019). The influence of haplotype H4 remained significant (p = 0.028) in the overall study population in similar analyses. CONCLUSION Smoking has the strongest impact on CYP1A2 activity, while gender and haplotype H4 showed marginal effects. The influence of the -163C>A polymorphism on CYP1A2 activity in smokers suggests an effect on the inducibility of the enzyme.

ABCB1 polymorphisms influence steady-state plasma levels of 9-hydroxyrisperidone and risperidone active moiety

Risperidone is metabolized to its active metabolite, 9-hydroxyrisperidone, mainly by the cytochrome P450 enzymes CYP2D6 and 3A4. Its antipsychotic effect is assumed to be related to the active moiety, that is, the sum of risperidone and 9-hydroxyrisperidone. Both risperidone and 9-hydroxyrisperidone are substrates of P-glycoprotein (P-gp), a transport protein involved in drug absorption, distribution, and elimination. The aim of the present study was to evaluate the influence of polymorphisms in genes encoding CYP3A5 and P-gp (ABCB1) on the steady-state plasma levels of risperidone, 9-hydroxyrisperidone, and the active moiety, taking CYP2D6 genotype status into account. Forty-six white patients with schizophrenia treated with risperidone (1-10 mg/d) in monotherapy for 4-6 weeks were genotyped, and their plasma concentrations of risperidone and 9-hydroxyrisperidone were measured. Dose-corrected plasma concentrations (C/D) of risperidone, 9-hydroxyrisperidone, and active moiety showed up to 68-, 9-, and 10-fold interindividual variation, respectively. Six patients carried 1 CYP3A5*1 allele and therefore were likely to express the CYP3A5 enzyme. The CYP3A5 genotype did not influence risperidone, 9-hydroxyrisperidone, or active moiety C/Ds. The CYP2D6 genotype in these 46 patients was again associated with risperidone C/D (P = 0.001) but not with 9-hydroxyrisperidone C/D or active moiety C/D, as previously shown by our group in 37 of these patients. Patients homozygous for the ABCB1 3435T/2677T/1236T haplotype had significantly lower C/Ds of 9-hydroxyrisperidone (P = 0.026) and active moiety (P = 0.028) than patients carrying other ABCB1 genotypes. In conclusion, our results confirmed the significant effect of CYP2D6 genotype on the steady-state plasma levels of risperidone and showed that ABCB1 polymorphisms have a moderate effect on those of 9-hydroxyrisperidone and the active moiety.

Flavin-containing monooxygenase 3 polymorphisms in 13 ethnic populations from Europe, East Asia and sub-Saharan Africa: frequency and linkage analysis

AIMS To investigate intra- and inter-ethnic differences in three widespread (E158K, V257M and E308G) and two African-specific (D132H and L360P) flavin-containing monooxygenase 3 (FMO3) polymorphisms. MATERIALS & METHODS Allele frequencies were determined by TaqMan allelic discrimination assay in 2152 healthy volunteers from Europe (Swedes, Italians and Turks), East Asia (Japanese) and sub-Saharan Africa (nine ethnic groups covering eastern, southern and western regions), followed by haplotype and linkage analysis. RESULTS Significant subpopulation differences (p < 0.001) in allele frequencies were found for E158K, V257M and E308G in Europeans and regional differences (p < 0.01) for D132H among Africans. No carrier of P360 was identified. Cis-linkage between G308 and K158 was confirmed with the compound variant (K158/G308) being found in a high proportion (12.0-38.3%) of non-African subjects, but rarely (1.3%) among Africans. CONCLUSIONS Distribution of functionally relevant FMO3 polymorphisms varies not only between ethnicities but also within. The K158/G308 variant may have potential clinical importance primarily in non-African populations due to its low prevalence in Africa.

Circadian Variations in the Activities of 6‐Phosphogluconate Dehydrogenase and Glucose‐6‐Phosphate Dehydrogenase in the Liver of Conrol and Streptozotocin‐Induced Diabetic Rats

The aim of this study was to examine: the 24 h variation of 6‐phosphogluconate dehydrogenase and glucose‐6‐phosphate dehydrogenase activities, key enzymes for the maintenance of intracellular NADPH concentration, in rat liver in control and streptozotocin‐induced diabetic animals. Adult male rats were fed ad libitum and synchronized on a 12:12 h light‐dark cycle (lights on 08:00 h). One group of animals was treated with streptozotocin (STZ, 55 mg/kg, intraperitoneal) to induce experimental diabetes. Eight weeks after STZ injection, the animals were sacrificed at six different times of day—1, 5, 9, 13, 17 and 21 Hours After Lights On (HALO)—and livers were obtained. Enzyme activities were determined spectrophotometrically in triplicate in liver homogenates and expressed as units per mg protein. 6‐phosphogluconate dehydrogenase activity was measured by substituting 6‐phosphogluconate as substrate. Glucose‐6‐phosphate dehydrogenase activity was determined by monitoring NADPH production. Treatment, circadian time, and interaction between treatment and circadian time factors were tested by either one or two way analysis of variance (ANOVA). Two‐way ANOVA revealed that 6‐phosphogluconate dehydrogenase activity significantly depended on both the treatment and time of sacrifice. 6‐phosphogluconate dehydrogenase activity was higher in control than diabetic animals; whereas, glucose‐6‐phosphate dehydrogenase activity did not vary over the 24 h in animals made diabetic by STZ treatment. Circadian variation in the activity of 6‐phosphogluconate dehydrogenase was also detected in both the control and STZ treatment groups (one‐way ANOVA). Time‐dependent variation in glucose‐6‐phosphate dehydrogenase activity during the 24 h was detected in control but not in diabetic rats. No significant interaction was detected between STZ‐treatment and time of sacrifice for both hepatic enzyme activities. These results suggest that the activities of NADPH‐generating enzymes exhibit 24 h variation, which is not influenced by diabetes.

Implications of Inter-Individual Differences in Clopidogrel Metabolism, with Focus on Pharmacogenetics.

Increasing evidence for the role of pharmacogenetics in treatment resistance to the antiplatelet agent clopidogrel has been gained during the last years. Apart from CYP2C19 genetic polymorphisms, nongenetic factors, particularly drug-drug interactions, age and other clinical characteristics influence the interindividual variability in clopidogrel response to varying degrees. The present article reviews the so far accumulated evidence on the role of pharmacogenetic traits influencing CYP-activity as determinants of the antiplatelet response to clopidogrel, and its clinical implications. The genetic variation in CYP2C19 activity seems to influence short- and long-term antithrombotic effects of clopidogrel to a substantial extent. Prediction models for clopidogrel non-responsiveness that include CYP2C19 genotyping together with relevant non-genetic risk factors are needed to be verified for their potential benefit in individualization of antithrombotic therapy.

The Effect of Experimental Diabetes on the Twenty‐Four‐Hour Pattern of the Vasodilator Responses to Acetylcholine and Isoprenaline ın the Rat Aorta

The aim of this study was to investigate whether time‐dependent variations in the relaxant effect of acetylcholine, an endothelium‐dependent vasorelaxant via muscarinic receptors, and isoprenaline, a nonselective β‐adrenoceptor agonist in rat aorta, are influenced by streptozotocin (STZ)‐induced experimental diabetes. Adult male rats were divided randomly into two groups: control and STZ‐induced (STZ, 55 mg/kg, intraperitoneal) diabetes. The animals were synchronized to a 12∶12 h light‐dark cycle (lights on 08∶00 h) and sacrificed at six different times of day (1, 5, 9, 13, 17, and 21 hours after lights on; HALO) eight weeks after STZ injection. The in vitro responsiveness of thoracic aorta rings obtained from control and diabetic rats to acetylcholine (10−9–10−5 M) and isoprenaline (10−10–10−3 M) was determined in six different times. EC50 (the concentration inducing half of the maximum response) values and maximum responses were calculated from cumulative concentration‐response curves of the agonists and were analyzed with respect to time and STZ treatment. Treatment, time, and interactions between treatment and time were tested by two‐way analysis of variance (ANOVA). To analyze differences due to biological time, one‐way ANOVA was used. STZ treatment did not significantly change EC50 values or maximum responses for both agonists. There were statistically significant time‐dependent variations in the EC50 values for isoprenaline and maximum responses for both acetylcholine and isoprenaline in control groups by one‐way ANOVA, but significant time‐dependent variations disappeared in the aortas isolated from STZ‐induced diabetic rats. The vasodilator responses to acetylcholine and isoprenaline failed to show any significant interaction (treatment×time of study) between STZ treatment and time of sacrifice in both EC50 values and maximum responses by two‐way ANOVA. These results indicate there is a basic temporal pattern in the responses to acetylcholine and isoprenaline in rat aorta which continues in diabetes. It is shown for the first time that experimental diabetes does not change the 24 h pattern of responses to acetylcholine and isoprenaline, and that time‐dependent variations in the responses to these agonists disappear in diabetic animals. Although further studies are required to define the underlying mechanism(s) of these findings, results suggest that experimental diabetes can modify the time‐dependent vasorelaxant responses of rat aorta. This may help to understand the circadian rhythms in cardiovascular physiology and pathology or in drug effects in diabetes.