Pharmacogenetics of acenocoumarol: CYP2C9, CYP2C19, CYP1A2, CYP3A4, CYP3A5 and ABCB1 gene polymorphisms and dose requirements.

BACKGROUND AND OBJECTIVE: Acenocoumarol (AC) is a coumarin derivative, vitamin K antagonist anticoagulant drug. It has a narrow therapeutic index and shows large pharmacokinetic and pharmacodynamic interindividual variability. Our objective was to investigate the association between AC dose requirements to achieve a target level of anticoagulation and genetic polymorphisms of genes possibly associated with its metabolism (CYP1A2, CYP2C9, CYP2C19, CYP3A4, CYP3A5) and transport (ABCB1). METHODS: Ninety-six Bulgarian patients treated orally with AC for at least 3 months were included. They were separated into three groups according to their AC dose requirement, i.e. low, medium and high. RESULTS AND DISCUSSION: CYP2C9*1/*3 (associated with an intermediate CYP2C9 activity), CYP2C9*2/*2, and CYP2C9*2/*3 genotypes (associated with a low CYP2C9 activity) were more prevalent in the group with low dose requirement of AC compared with the other two groups (P = 0.003). The frequency of CYP2C9*1/*1 genotype, which is associated with an extensive CYP2C9 activity, was higher in the group of patients with high dose requirements (79%), compared with the groups of the medium and low dose requirements (67% and 21% respectively). In addition, the ABCB1 2677GG/3435CC haplotype was associated with use of lower AC dose, whereas the 2677TT/3435TT and 2677GT/3435TT haplotypes were associated with use of higher AC dose (P = 0.03). The distribution of polymorphisms of other genes did not show significant differences between the three groups. CONCLUSION: In vivo, cytochromes P450 isoforms other than CYP2C9 [DOSAGE ERROR CORRECTED] were not significantly associated with dose requirement of AC. In our Bulgarian patients, the presence of CYP2C9*2 or/and CYP2C9*3 alleles, as well as the ABCB1 2677GG/3435CC haplotype were associated with low dose requirement of AC.

Detection of apoB-100 R3500Q mutation by competitive allele-specific polymerase chain reaction.

The apolipoprotein B-100 mutation R3500Q is one of the most common inherited defects causing abnormality of the lipid metabolism. We describe a one-step, single-tube PCR technique for detection of the mutation based on competition between allele-specific primers. Three oligonucleotides are used: two allele-specific primers differing in their 3' nucleotide (for the wild-type and the mutant allele) together with a common primer, resulting in simultaneous amplification of both alleles. This provided internal control of successful amplification and is expected to result in increased specificity. The allele-specific primers differ also in length, allowing us to distinguish both alleles by their size in a single electrophoretic run. For optimization of the protocol, DNAs genotyped before by oligonucleotide ligation assay were used. The individual genotypes obtained by CAS-PCR coincided fully with the ones from a referent OLA test: seven heterozygous individuals were found, 4 of them among 150 unrelated hypercholesterolemic individuals studied and other three in the pedigrees of heterozygous carriers. On the overall 160 genotypes were determined, neither false-positive (0 out of 153 non-carriers) nor false-negative (0 out of 7 carriers) results were obtained. No homozygous mutant genotypes were identified in this sample.

Is the incidence of Balkan endemic nephropathy decreasing?

Balkan Endemic Nephropathy (BEN) is a non-inflammatory, slowly progressing, familial, primarily tubulo-interstitial, bilateral renal disease that affects rural populations in several Balkan countries. Our study describes a time trend of the incidence of BEN in eight villages of Vratza District, Bulgaria, for the period 1965-1987, based on three various data sets. The data suggest that after the initial peak between 1967 and 1970, the incidence remained quite stable for the period 1970-1984, and declined after 1984. However, the study also demonstrates under-recording of BEN cases and less complete case identification, especially after 1979. Migration of population might also have contributed to an apparent decline in registered cases. We detected cases of BEN in villages that previously were BEN-free. We recommended a rigorous monitoring of BEN in all afflicted countries, before concluding that the incidence of BEN is decreasing.

Ochratoxin A concentrations in food and feed from a region with Balkan Endemic Nephropathy.

Balkan Endemic Nephropathy (BEN), a chronic renal disease of unknown aetiology, is found in geographically close areas of Bulgaria, Romania, Serbia, Croatia, Bosnia and Herzegovina, Slovenia, and the former Yugoslav Republic of Macedonia. Ochratoxin A (OTA), a secondary metabolite of Aspergillus and Penicillium species and a natural contaminant of food and feed, is a putative cause of BEN. Some studies have found a geographic covariation between OTA content in food/feed and BEN manifestation; others have not. In May 2000, using a competitive direct ELISA assay for OTA (detection limit 1 microg kg(-1)), we investigated OTA contamination in 165 samples of home-produced food (beans, potatoes, corn, wheat, flour) and feed from households in villages from the BEN region (Vratza district) of north-western Bulgaria. Samples were collected from: (a) BEN villages (n = 8), and therein from BEN households (20), and BEN-free households (16) (within-village controls, WVC households); and (b) BEN-free villages (7) and therein BEN-free households (22) (between-village controls, BVC). BEN households consistently had a higher proportion of OTA-positive samples than WVC households, but similar (for some foods) or lower (for other foods) proportions to BVC households. The proportion of OTA-positive samples was also higher in BVC than in WVC households. Furthermore, BEN households had a similar proportion of OTA-positive samples to the pooled, WVC and BVC, group of households. OTA-exposure estimates, derived from our OTA-concentration findings and the reported average per capita monthly consumption of basic foods in rural Bulgaria, showed the highest OTA intake in BEN households (1.21 microg day(-1)), versus 1.03 microg day(-1) in BVC and 0.71 microg day(-1) in WVC households. These OTA intakes are higher than those in the EU, and are close to the upper limits acceptable to several food-safety organizations. The results indicate that OTA may not alone cause BEN; only synergistically with other environmental toxicants and/or predisposing genotypes may do so.