Elsevier

Translational Research

Volume 160, Issue 4, October 2012, Pages 298-308
Translational Research

Original Article
Influence of hepatic and intestinal efflux transporters and their genetic variants on the pharmacokinetics and pharmacodynamics of raloxifene in osteoporosis treatment

https://doi.org/10.1016/j.trsl.2012.03.002Get rights and content

Raloxifene exhibits a large and unexplained interindividual variability in its pharmacokinetics and pharmacodynamics. The aim of our study was to identify transporters involved in the efflux of raloxifene and its glucuronide metabolites by various in vitro models and by an in vivo study to explore the possible involvement of P-glycoprotein (Pgp), multidrug resistance–associated protein (MRP)1, MRP2, MRP3, and breast cancer resistance protein in the observed high interindividual variability. Experiments with the parallel artificial membrane permeability assay showed the highest passive permeability for raloxifene, followed by raloxifene-6-β-glucuronide (M1), raloxifene-4'-β-glucuronide (M2), and raloxifene-6,4'-diglucuronide (M3). Caco-2 cell monolayer experiments indicated an interaction of raloxifene with Pgp. The ATPase assay confirmed the raloxifene interaction with Pgp and indicated interactions of all raloxifene species with MRP1, MRP2, MRP3, and breast cancer resistance protein, except for M1, which did not show any interactions with MRP2. Furthermore, the vesicular experiments confirmed the interaction of M2 and M3 with MRP2. Although the in vivo study on osteoporotic postmenopausal women on raloxifene could not confirm a significant influence of ABCB1 and ABCC2 genetic polymorphisms on its pharmacokinetics, a clear trend toward higher total raloxifene concentrations was observed in carriers of at least 1 ABCB1 c.3435T allele. Moreover, the same polymorphism effect was also observed as a significant increase in total hip bone mineral density after 1 year of treatment. The results of our study support the involvement of efflux transporters in disposition of raloxifene and its metabolites and may partially explain the observed raloxifene variability by the influence of the ABCB1 c.3435C>T polymorphism.

Section snippets

Materials

Raloxifene hydrochloride, fumitremorgin C (Fum C), MK571, verapamil, haloperidol, fluorescein (FLU), dimethylsulfoxide (DMSO), estradiol-17β-glucuronide (E17βG), and formic acid were from Sigma-Aldrich Chemie (Deisenhofen, Germany). Raloxifene metabolites M1, M2, and M3 were synthesized by incubating raloxifene with Streptomyces sp. American Tissue Culture Collection 5504320 followed by semipreparative chromatographic purification and lyophilization. The details of the synthesis and

Passive permeability measurements

The passive part of raloxifene species permeability was assessed with a PAMPA model. The rank order of the compound permeability through the PAMPA membrane shows that raloxifene has the highest passive permeability, followed by raloxifene monoglucuronides and M3, with the lowest passive permeability (Table I).

Transcellular transport assay

Caco-2 cell monolayers were used for the identification of transporters involved in raloxifene transport and for the determination of absorptive raloxifene permeability. The results

Discussion

The main goal of our study was to determine which excretory transporters may play a significant role in the disposition of raloxifene and may also be involved in the observed variability in raloxifene exposure and therapeutic effect. Complex in vitro and in vivo approaches have been used to comprehensively describe the excretory transport of raloxifene and its conjugates. We have shown a strong in vitro evidence of raloxifene species interaction with Pgp and MRP transporters, which encouraged

Conclusions

We discovered which transporters are most likely involved in the excretion of raloxifene species into bile, which is an important step in the enterohepatic recirculation of raloxifene and therefore an important determinant for the overall raloxifene exposure. The presented data implicate that at least a part of the observed variability could be explained by the genetic influence of ABCB1 c.3435C>T polymorphism. In this study, the synergistic use of in vitro models combined with a small in vivo

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    This work was supported by Grants J1-0957 and P3-0298 provided by the Slovenian Research Agency.

    Conflict of Interest: none.

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