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Impaired Clearance of Methotrexate in Organic Anion Transporter 3 (Slc22a8) Knockout Mice: A Gender Specific Impact of Reduced Folates

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Abstract

Purpose

To elucidate the role of the renal basolateral transporter, Oat3, in the disposition of methotrexate.

Materials and Methods

Chinese hamster ovary cells expressing mouse Oat3 were used to determine kinetics and specificity of inhibition of methotrexate transport. Methotrexate clearance was then examined in vivo in wildtype and Oat3 knockout mice.

Results

NSAIDs, ß-lactams, and uremic toxins inhibited mOat3-mediated methotrexate uptake by 70–100%, while folate, leucovorin, and 5-methyltetrahydrofolate inhibited transport by 25–50%. A K m of 60.6 ± 9.3 μM for methotrexate transport was determined. Oat3 knockout mice exhibited reduced methotrexate-to-inulin clearance ratios versus wildtype. Male wildtype mice, but not knockouts or females, demonstrated significantly accelerated methotrexate clearance in response to reduced folates. Reduced folates also markedly inhibited hepatic methotrexate accumulation in males, but not females, and the response was independent of Oat3 function.

Conclusions

Oat3 contributes to methotrexate clearance, but represents only one component responsible for methotrexate’s elimination. Therefore, in patients, dysfunctional hOAT3 polymorphisms or drug competition for hOAT3 transport may severely impact methotrexate elimination only when redundant means of methotrexate removal are also compromised. Furthermore, the present findings suggest that reduced-folate administration only influences methotrexate disposition in males, with the renal reduced-folate response influenced by OAT3 function.

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Abbreviations

5-CH3-THF:

5-methyltetrahydrofolate

5-CHO-THF:

5-formyltetrahydrofolate (a.k.a. folinic acid or leucovorin)

5-HIAA:

5-hydroxyindoleacetic acid

CHO-FRT:

Chinese hamster ovary cells transfected with empty vector

CHO-mOat3:

Chinese hamster ovary cells transfected with murine organic anion transporter 3

DOPAC:

dihydroxyphenylacetic acid

h:

human

K m :

the Michaelis–Menten constant

m:

mouse

MRP:

multidrug resistance-associated protein

MTX:

methotrexate

NSAIDs:

non-steroidal anti-inflammatory drugs

OAT or Oat:

human or non-human organic anion transporter, respectively

r:

rat

RFC-1:

reduced folate carrier 1

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Acknowledgements

This work was supported by National Institute of Diabetes and Digestive and Kidney Diseases Grant R01-DK-067216 (D.H.S.), by fellowship provisions from the American Foundation for Pharmaceutical Education (A.L.V.), and by the National Institutes of Health Grant Number C06 RR015455 from the Extramural Research Facilities Program of the National Center for Research Resources.

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  1. Department of Pharmaceutical Sciences, Medical University of South Carolina, 280 Calhoun Street (Rm. QE218), P.O. Box 250140, Charleston, South Carolina, 29425, USA

    Adam L. VanWert & Douglas H. Sweet

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  1. Adam L. VanWert
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Correspondence to Douglas H. Sweet.

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VanWert, A.L., Sweet, D.H. Impaired Clearance of Methotrexate in Organic Anion Transporter 3 (Slc22a8) Knockout Mice: A Gender Specific Impact of Reduced Folates. Pharm Res 25, 453–462 (2008). https://doi.org/10.1007/s11095-007-9407-0

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  • DOI: https://doi.org/10.1007/s11095-007-9407-0

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