4-Methylpyrazole partially ameliorated the teratogenicity of retinol and reduced the metabolic formation of all-trans-retinoic acid in the mouse.

Oral administration of retinol (50 mg/kg) to NMRI mice on day 11 of gestation (vaginal plug = day 0) led to the metabolic formation of high quantities of all-trans retinoic acid and all-trans-4-oxoretinoic acid, both known as potent teratogenic agents in the mouse. A 96% reduction of the area under the concentration-versus-time-curve (AUC) of metabolically generated all-trans retinoic acid in maternal plasma, and an 84% decrease in the embryonic AUC were observed when mice had been pretreated with the alcohol dehydrogenase inhibitor 4-methylpyrazole. A similar reduction was observed for the major metabolite of all-trans retinoic acid in the mouse, all-trans-4-oxoretinoic acid. However, 4-methylpyrazole pretreatment decreased the AUC of retinol by 10% in maternal plasma and 15% in embryo. Treatment with retinol alone resulted in 55.6%, 43.9% and 56.0% skeletal anomalies of the forelimbs, hindlimbs and craniofacial structures, respectively. Pretreatment with 4-methylpyrazole lowered the retinol induced skeletal defects to 31.3%, 24.0% and 31.3%, respectively, in the forelimb, hindlimb and craniofacial region. Typical retinoid-induced malformations for gestational day 11, e.g. bent or reduced zeugopod or stylopod elements, or cleft palate, were significantly reduced by 4-methylpyrazole pretreatment but were still detected in significantly higher prevalence than in control mice. These data suggest that the teratogenic activity of a single high dose of vitamin A in mouse is partially but not exclusively dependent on the metabolic activation of retinol to all-trans retinoic acid. Thus it could be hypothesized that retinol is either a proximate teratogen or a coteratogen with all-trans retinoic acid.

Teratogenicity and placental transfer of all-trans-, 13-cis-, 4-oxo-all-trans-, and 4-oxo-13-cis-retinoic acid after administration of a low oral dose during organogenesis in mice.

13-cis-Retinoic acid (isotretinoin) is teratogenic in humans at therapeutic doses (0.5-1.5 mg/kg) but only marginally teratogenic in the mouse at a high dose of 100 mg/kg. Previous results explained why the cis isomer of retinoic acid was much less teratogenic than the trans isomer in mice. It was found that the placental transfer of all-trans retinoic acid to the mouse embryo was far greater than that of the 13-cis isomer. Since our previous study had been performed with exceedingly high doses (100 mg/kg) of 13-cis-retinoic acid and all-trans-retinoic acid, we have now performed additional experiments with 10-fold lower doses. Studies were also done with the main metabolites of the two retinoids (the 4-oxo-derivatives) to elucidate the metabolism, pharmacokinetics, and teratogenicity of each single compound. It was shown that all-trans-retinoic acid and 4-oxo-all-trans-retinoic acid were extremely teratogenic, whereas their corresponding cis isomers caused only 2% cleft palate. Embryonic exposure to the trans isomers was likewise higher than that to the cis isomers, as shown by the far higher embryonic peak concentrations and by the 30-fold higher areas under the concentration-time curve values reached for the trans isomers compared with the cis isomers. At 8 hr, embryo/maternal plasma ratios were higher than 1 after administration of the all-trans compounds. Concentrations found in the placenta and yolk sac were higher for the trans forms than for the cis forms. We propose a model for a facilitated transport of the all-trans forms to the developing embryo and suggest that the conversion to the trans isomer and trans metabolite could play a major role in the teratogenicity of 13-cis-retinoic acid in humans.