[Objectives] The fetal-to-maternal concentration ratio (F/M ratio) of metformin, a substrate of organic cation transporters (OCT) and multidrug and toxin extrusion proteins (MATE), has been reported to be higher in pregnant humans than in pregnant rats. The objective of the present study was to identify transporters that cause species differences in metformin transfer to the fetus.

[Methods] Uncomplicated term human placental tissues were obtained with written informed consent and with the approval of the Institutional Ethics Committee of Keio University Faculty of Pharmacy (150421–2) and Keio University School of Medicine (20110250). The absolute protein expression levels of OCT3 and MATE1 in human, rat, and mouse placentas were quantified using LC-MS/MS. The mRNA expression levels were determined from RNA-seq data. Pregnant rats and mice were continuously infused with [¹⁴C]metformin via the jugular vein, and the F/M ratio was measured. Pyrimethamine was used as a potent inhibitor of MATE1. Statistical analyses were performed using an unpaired, two-tailed Student’s t-test for comparisons between the two groups.

[Results] OCT3 protein expression was detected in human, rat, and mouse placenta (n=3-4). However, MATE1 protein expression was selectively detected in rat placenta, but not in human and mouse placenta. These protein expression levels exhibited consistency with the mRNA expression levels: while no significant species differences were observed in OCT3 mRNA expression levels, MATE1 mRNA expression levels in the rat placental labyrinth were found to be considerably higher than those in human syncytiotrophoblasts and mouse labyrinth. The F/M ratio of [¹⁴C]metformin was significantly lower in rats (n=5) than in mice (n=7). The pre-administration of pyrimethamine at 2 μmol/kg was found to reach a plasma concentration capable of specific inhibition of Mate1, as indicated by the measured IC₅₀ of pyrimethamine to rat Mate1 (0.015 μM) and Oct3 (71.5 μM), and resulted in a significant increase in the F/M ratio of [¹⁴C]metformin in rats (n=5) but not in mice (n=7).

[Conclusions] The findings of this study suggest that the fetal transfer of metformin in rats is constrained by placental MATE1 expression. These findings indicate that nonclinical developmental and reproductive toxicity studies of MATE1 substrate drugs in rats may underestimate fetal exposure when extrapolated to humans. This information will be helpful in estimating fetal transfer and toxicity of drugs in human fetuses from rat data.