Although subcellular mRNA trafficking has been demonstrated as a mechanism to control protein distribution, it is generally believed that most protein localization occurs subsequent to translation. To address this point, we developed and employed a high-resolution fluorescent in situ hybridization procedure to comprehensively evaluate mRNA localization dynamics during early Drosophila embryogenesis. Surprisingly, of the 3370 genes analyzed, 71% of those expressed encode subcellularly localized mRNAs. Dozens of new and striking localization patterns were observed, implying an equivalent variety of localization mechanisms. Tight correlations between mRNA distribution and subsequent protein localization and function, indicate major roles for mRNA localization in nucleating localized cellular machineries. A searchable web resource documenting mRNA expression and localization dynamics has been established and will serve as an invaluable tool for dissecting localization mechanisms and for predicting gene functions and interactions.

In a previous study we showed that the disposition of clozapine after a single oral dose is unrelated to either debrisoquine or S-mephenytoin hydroxylation polymorphism. The same 14 healthy subjects studied in that investigation were given 150 mg of caffeine. The reciprocal of plasma clozapine AUC (0,24), was correlated with an index of the N3-demethylation of caffeine (r, = 0.84; P = 0.0024), used as a measure of cytochrome P4501A2 (CYP1A2) activity. Ni-and N7-demethylation indices of caffeine also reflect CYP1A2 activity and were also correlated with clozapine clearance (rs = 0.89 and 0.85; P = 0.0013 and 0.0023; respectively). No significant relationships with xanthine oxidase and N-acetyl transferase activity, also assessed by a caffeine test, were found. This study suggests that clozapine is metabolised by CYP1A2 to a major extent.

Twenty-five healthy volunteers were given 100 mg caffeine orally and several estimates of cytochrome P450 1A2 (CYP1A2) activity were evaluated. The validation was performed by correlation of different parameters in plasma, saliva, and urine to two measures of caffeine clearance, CL(oral) and CL(137X-->17X) that served as standards of reference. Two subjects were excluded because of noncompliance with a caffeine-free diet. In the remaining 23 subjects, both plasma and saliva total clearances of caffeine were highly correlated with each other (r(s) = 0.97, p < 0.0001). The ratio 17X/137X restricted to one sampling point taken 4 hours after dose, showed a high correlation (r(s)) with CL(oral) and CL(137X-->17X) in plasma (0.84/0.83) and saliva (0.82/0.77) (p < 0.0001 for all the correlation values) where 17X is 1,7-dimethylxanthine (paraxanthine) and 137X is 1,3,7-trimethylxanthine (caffeine). Additionally, the ratio (AFMU + 1U + 1X + 17U + 17X)/137X in a 0-24 hours urine sampling showed the highest correlation with CL(137X-->17X) (r(s) = 0.85, p < 0.001) where AFMU is 5-acetylamino-6-formylamino-3-methyluracil, 1U is 1-methyluracil, 1X is 1-methylxanthine, and 17U is 1,7-dimethyluric acid. The major estimates of CYP1A2 activity were significantly less in nonsmoking females, and this probably was related to the use of oral contraceptives in this subpopulation. In summary, among caffeine-based approaches for CYP1A2, the authors recommend either plasma or saliva 17X/137X ratio and the urinary (AFMU + 1U + 1X + 17U + 17X)/137X ratio during a sampling interval of at least 8 hours, starting at time zero since caffeine intake. These indices are simple, reliable, and relatively inexpensive estimates of CYP1A2 activity to be used in the study of human populations.

Mephenytoin (100 mg) and debrisoquin (10 mg) were administered orally, both separately and together, to 41 healthy subjects. The ratios between the S and R enantiomers of mephenytoin and between debrisoquin and 4-OH-debrisoquin in urine were determined by use of GC. These ratios were used as measures of drug hydroxylation. There was no change in the phenotypic trait values of the two drugs when they were coadministered. Mephenytoin and debrisoquin then were coadministered to 253 healthy Swedish subjects, before bedtime, and urine samples were collected at periods of 0 to 8, 8 to 24, and 24 to 32 hours after drug administration. In the first sample, seven of the 253 subjects (2.8%, 95% confidence interval 0.8% to 4.8%) had an S/R ratio of greater than 0.8; this indicated that they were poor hydroxylators of S-mephenytoin. In the two consecutive samples, the S/R ratios of mephenytoin did not change in these seven persons, whereas it decreased to less than 0.2 in the third sample in the extensive hydroxylators. As was reported before, there was no relationship between the mephenytoin S/R ratio and the debrisoquin metabolic ratio (rs = 0.01). Coadministration of debrisoquin and mephenytoin before bedtime and urine collection during two consecutive nights allow for an accurate determination of both phenotypes in the population.