gms | German Medical Science

Introduction: Caffeine biotransformation has been extensively studied and inter-individual variation in caffeine metabolism can be explained by multiple environmental and genetic factors. Due to the excellent safety in low doses, coffee or caffeine have been extensively used as probe drugs particularly for the in vivo cytochrome P450 1A2 (CYP1A2) or N-acetyltransferase 2 (NAT2) activities. Heritability of NAT2 activity is well understood at the molecular level. But for CYP1A2, results concerning heritability in general and concerning the functional role of specific polymorphisms are controversial. Thus, our primary objective is to estimate the genetic influence on the in vivo activity of CYP1A2 using the caffeine plasma AUC as primary parameter. The analysis of the urinary ratio of 5-acetylamino-6-formylamino-3-methyluracil (AFMU) over 1-methylxanthine (1X) as parameters of NAT2 activity should serve as a positive control in our analysis of heritability. The additional use of 13C caffeine should particularly abolish any interferences with caffeine from coffee or other bevarages.

Methods: Unlabelled 12C Caffeine (50 mg) was orally administered to 44 monozygotic (MZ) and 14 dizygotic (DZ) twin pairs on up to three occasions. Additionally a subgroup of 29 monozygotic and 8 dizygotic twin pairs ingested once 50 mg 13C labeled caffeine. Caffeine (1,3,7 trimethylxanthine) and its metabolite paraxanthine (1,7 dimethylxanthine) were measured in plasma. AFMU and 1X were measured in urines collected between 4 - 7 h after intake of unlabeled caffeine and between 5 and 8 h after intake if 13C labeled caffeine. Heritability was calculated by structural equation modeling dividing variation into additive genetic (A), common environmental (C) and unique environmental (E) effects. Genotyping covered the CYP1A2*1F allele as well as the NAT2*4, *5A, *5B, *5C, *6A and *7B alleles. Smoking status was confirmed by inquiry on the day of the screening procedures.

Results and Discussion: Our calculations confirmed the well known high heritability of NAT2 activity (additive genetic effects (A) = 99%, unique environmental effects (E) = 1%) thereby confirming that the study design was appropriate to detect highly heritably traits and showing that there are apparently only very few environmental effects modulating in vivo NAT2 activity For both traits, NAT2 and CYP1A2, results were very similar for labeled and unlabeled caffeine. No heritability was found in the variation of the AUC0-infinity and AUC0-300 as indicators of CYP1A2 activity but a large proportion of 70% or more of the variation in CYP1A2 activity was due to common environmental effects (C). Five monozygotic and 2 dizygotic twin pairs from the total group of 58 twin pairs were discordant for smoking habits, but correlation coefficients were almost the same concerning just twin pairs with the same smoking habits (rMZ = 0.85, rDZ = 0.65) and all twin pairs together (rMZ = 0.83, rDz = 0.68) as well as concerning concordant smoking MZ twin pairs (r = 0.88) and concordant non-smoking twin pairs (r = 0.81). A fraction of over 50% in AFMU/1X variation was determined by the studied NAT2 genotypes whereas the one CYP1A2 genotype did not determine caffeine AUC. These results concerning CYP1A2 are in contrast to several earlier data showing or assuming high heritability of CYP1A2 activity. One reason for these differences may be the differences in phenotyping CYP1A2 activity because we assessed CYP1A2 activity from plasma AUC which we assume to reflect best total metabolic clearance whereas urinary metabolic ratios analyzed by others may also reflect the activities of renal anion transporters. The wide variation in CYP1A2 activity is well known and is known to be partially due to enzyme induction via the Ah receptor. According to our data, heritability of CYP1A2 activity and Ah receptor mediated gene regulation is apparently only small but there are apparently environmental factors acting in parallel in monozygotic and dizygotic twins which have a relevant impact on caffeine AUC.