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Atorvastatin Transport in the Caco-2 Cell Model: Contributions of P-Glycoprotein and the Proton-Monocarboxylic Acid Co-Transporter

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Abstract

Purpose. The purpose of this study was to elucidate the mechanismsby which an HMG-CoA reductase inhibitor, atorvastatin (an organicacid with a pKa of 4.46), was transported in the secretory and absorptivedirections across Caco-2 cell monolayers.

Methods. Caco-2 cells were grown on polycarbonate membrane insertsin 6-well Snapwell plates (Costar). The permeability of radiolabeledcompounds across Caco-2 cell monolayers was determined using aside-by-side diffusion apparatus (NaviCyte) and an automated liquidhandler (Hamilton Microlab 2200). The apical uptake of14C-atorvastatin was also determined in Caco-2 cells. Cyclosporin A (20 μM) waspresent in the uptake media to block potential P-glycoprotein-mediatedatorvastatin efflux.

Results. Polarized permeation of atorvastatin was observed with thebasolateral-to-apical (B-to-A) permeability being 7-fold greater thanthe A-to-B permeability (35.6 × 10−6 and 4.9 × 10−6 cm/s,respectively). The secretion of atorvastatin was a saturable process with anapparent Km of 115 μM. The B-to-A permeability of atorvastatin wassignificantly reduced by cyclosporin A (10 μM), verapamil (100 μM),and a P-glycoprotein specific monoclonal antibody, UIC2(10 μg/ml)(43%, 25%, and 13%, respectively). Furthermore, both CsA andverapamil significantly increased the A-to-B permeability of atorvastatinby 60% however, UIC2 did not affect the A-to-B permeability ofatorvastatin. CsA uncompetitively inhibited the B-to-A flux ofatorvastatin with a Ki of 5 μM. In addition, atorvastatin (100 μM) significantlyinhibited the B-to-A permeability of vinblastine by 61%. The apicaluptake of atorvastatin increased 10.5-fold when the apical pH decreasedfrom pH 7.4 to pH 5.5 while the pH in the basolateral side wasfixed at pH 7.4. A proton ionophore, carbonylcyanidep-trifluoro-methoxyphenylhydrazone (FCCP) significantly decreased atorvastatinuptake. In addition, atorvastatin uptake was significantly inhibited bybenzoic acid, nicotinic acid, and acetic acid each at 20 mM (65%,14%, and 40%, respectively). Benzoic acid competitively inhibitedatorvastatin uptake with a Ki of 14 mM. Similarly, benzoic acid,nicotinic acid, and acetic acid significantly, inhibited the A-to-Bpermeability of atorvastatin by 71%, 21%, and 66%, respectively.

Conclusion. This study demonstrated that atorvastatin was secretedacross the apical surface of Caco-2 cell monolayers viaP-glycoprotein-mediated efflux and transported across the apical membrane in theabsorptive direction via a H+-monocarboxylic acid cotransporter(MCT). In addition, this study provided the first evidence thatnegatively charged compounds, such as atorvastatin, can be a substrate forP-glycoprotein.

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REFERENCES

  1. D. D. Cilla, Jr., L. R. Whitfield, D. M. Gibson, A. J. Sedman, and E. L. Posvar. Multiple-dose pharmacokinetics, pharmacody-namics, and safety of atorvastatin, an inhibitor of HMG-CoA reductase, in healthy subjects. Clin. Pharmacol. Ther. 60, 687-695 (1996).

    Google Scholar 

  2. D. M. Gibson, R. H. Stern, R. B. Able, and L. R. Whitfield. Absolute bioavailability of atorvastatin in man, Pharm. Res. 14 (Suppl):S-253 (1997).

    Google Scholar 

  3. B. M. Michniewicz, A. E. Black, M. W. Sinz, and T. F. Woolf. In vitro and in vivo metabolism of atorvastatin (CI-981). ISSX Proceeding, Volume 6, p 93, Sixth North American ISSX Meeting, Raleigh, NC, Oct 23-27 (1994).

    Google Scholar 

  4. B. H. Stewart, E. L. Reyner, R. Stern, E. A. Zegarac, R. Boyd, and L. R. Whitfield, Atorvastatin inhibits the p-glycoprotein-mediated secretion of 3 H-digoxin in Caco-2 cell monolayers. Pharm. Res. 14 (Suppl):S-671 (1997).

    Google Scholar 

  5. J. Dimitroulakos and H. Yeger. HMG-CoA reductase mediates the biological effects of retinoic acid on human neuroblastoma cells. Nature Med. 2, 326-333 (1996).

    Google Scholar 

  6. A. Tsuji, H. Takanaga, I. Tamai, and T. Terasaki. Transcellular transport of benzoic acid across Caco-2 cells by a pH-dependent and carrier-mediated transport mechanism. Pharm. Res. 11:30-37 (1994).

    Google Scholar 

  7. I. Tamai, H. Takanaga, H. Maeda, T. Ogihara, M. Yoneda, and A. Tsuji. Proton-co-transport of pravastatin across intestinal brush-border membrane. Pharm. Res. 12:1727-1732 (1995).

    Google Scholar 

  8. M. Dixon and E. C. Webb, Chapter VIII: enzyme inhibition and activation in Enzyme. 3rd ed. pp332-467, Academic Press (1979).

  9. J. Hunter, M. A. Jepson, T. Tsuruo, N. L. Simmons, and B. H. Hirst. Functional expression of P-glycoprotein in apical membranes of human intestinal Caco-2 cells. J. Biol. Chem. 268:14991-14997 (1993).

    Google Scholar 

  10. E. B. Mechetner and I. B. Roninson. Efficient inhibition of P-glycoprotein-mediated multidrug resistance with a monoclonal antibody. Proc. Natl. Acad. Sci. USA 89:5824-5828 (1992).

    Google Scholar 

  11. H. Gutmann, G. Fricker, M. Török, S. Michael, C. Beglinger, and J. Drewe. Evidence for different ABC-transporters in Caco-2 cells modulating drug uptake. Pharm. Res. 16:402-407 (1999).

    Google Scholar 

  12. U. K. Walle, K. I. French, R. A. Walgren, and T. Walle. Transport of genitein-7-glucoside by human intestinal Caco-2 cells: potential role for MRP2. Res. Commun. Mol. Pathol. Pharmacol. 103:45-56 (1999).

    Google Scholar 

  13. I. Tamai, H. Takanaga, H. Maeda, Y. Sai, T. Ogihara, H. Higashida, and A. Tsuji. Participation of a proton-cotransporter, MCT1 in the intestinal transport mechanism for monocarboxylic acids. Biochem. Biophys. Res. Commun. 214:482-489 (1995).

    Google Scholar 

  14. A. Tsuji and I. Tamai. Carrier-mediated intestinal transport of drugs. Pharm Res. 13:963-977 (1996).

    Google Scholar 

  15. B.-B. Yang, J. A. Smithers, R. H. Stern, A. J. Sedman, and S. C. Olson, Pharmacokinetics and dose proportionality of atorvastatin and its active metabolites. Pharm. Res. 13 (Suppl):S-437 (1996).

    Google Scholar 

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Wu, X., Whitfield, L.R. & Stewart, B.H. Atorvastatin Transport in the Caco-2 Cell Model: Contributions of P-Glycoprotein and the Proton-Monocarboxylic Acid Co-Transporter. Pharm Res 17, 209–215 (2000). https://doi.org/10.1023/A:1007525616017

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