Pharmaceutical Research

, Volume 21, Issue 8, pp 1382–1389

In Situ Transport of Vinblastine and Selected P-glycoprotein Substrates: Implications for Drug-Drug Interactions at the Mouse Blood-Brain Barrier

  • Salvatore Cisternino
  • Christophe Rousselle
  • Marcel Debray
  • Jean-Michel Scherrmann


Purpose. To study the intrinsic parameters of P-glycoprotein (P-gp) transport and drug-drug interactions at the blood-brain barrier (BBB), as few quantitative in vivo data are available. These parameters could be invaluable for comparing models and predicting the in vivo implications of in vitro studies.

Methods. The brains of P-gp-deficient mice mdr1a(-/-) and wild-type mice were perfused in situ using a wide range of colchicine, morphine, and vinblastine concentrations. The difference between the uptake by the wild-type and P-gp-deficient mice gave the P-gp-linked apparent transport at the BBB. Drug-drug interactions were examined using vinblastine and compounds that bind to P-gp sites (verapamil, progesterone, PSC833) other than the vinblastine site to take into account the multispecific drug P-gp recognition.

Results. P-gp limited the brain uptake of morphine and colchicine in a concentration-independent way up to 2 mM. In contrast, vinblastine inhibited its own P-gp transport with an IC50 of ∼56 μM and a Hill coefficient of ∼4. The vinblastine efflux by P-gp was described by a Km at 16 μM and a maximal efflux velocity, Jmax, of ∼8 pmol s−1 g−1 of brain. Similarly, vinblastine brain transport was increased by inhibiting P-gp as shown by the IC50 ranking, which was PSC833 < verapamil < vinblastine < progesterone.

Conclusions. P-gp is responsible for both capacity-limited and -unlimited transport of P-gp substrates at the mouse BBB. In situ perfusion of mdr1a(\-/\-) and wild-type mouse brains could be used to predict drug-drug interactions for P-gp at the mouse BBB.

blood-brain barrier in situ brain perfusion multidrug resistance P-glycoprotein vinblastine 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    H. Sun, H. Dai, N. Shaik, and W. F. Elmquist. Drug efflux transporters in the CNS. Adv. Drug Deliv. Rev. 55:83–105 (2003).Google Scholar
  2. 2.
    T. Eisenblatter and H. J. Galla. A new multidrug resistance protein at the blood-brain barrier. Biochem. Biophys. Res. Commun. 293:1273–1278 (2002).Google Scholar
  3. 3.
    S. V. Ambdukar, S. Dey, and C. A. Hrycyna, M. Ramachandra, I. Pastan, and M. M. Gottesman. Biochemical, cellular and pharmacological aspects of the multidrug transporter. Annu. Rev. Pharmacol. Toxicol. 39:361–398 (1999).Google Scholar
  4. 4.
    A. Seelig. and E. Landwojtowicz. Structure-activity relationship of P-glycoprotein substrates and modifiers. Eur. J. Pharm. Sci. 12:31–40 (2000).Google Scholar
  5. 5.
    A. H. Schinkel, E. Wagenaar, L. van Deemter, C. A. Mol, and P. Borst. Absence of the mdr1a P-Glycoprotein in mice affects tissue distribution and pharmacokinetics of dexamethasone, digoxin, and cyclosporin A. J. Clin. Invest. 96:1698–1705 (1995).Google Scholar
  6. 6.
    S. Cisternino. F. Bourasset, Y. Archimbaud, D. Semiond, G. Sanderink, and J. M. Scherrmann. Nonlinear accumulation in the brain of the new taxoid TXD258 following saturation of Pglycoprotein at the blood-brain barrier in mice and rats. Br. J. Pharmacol. 138:1367–1375 (2003).Google Scholar
  7. 7.
    M. F. Fromm, R. B. Kim, C. M. Stein, G. R. Wilkinson, and D. M. Roden. Inhibition of P-glycoprotein-mediated drug transport: a unifying mechanism to explain the interaction between digoxin and quinidine. Circulation 99:552–557 (1999).Google Scholar
  8. 8.
    D. K. Yu. The contribution of P-glycoprotein to pharmacokinetic drug-drug interactions. J. Clin. Pharmacol. 39:1203–1211 (1999).Google Scholar
  9. 9.
    Q. R. Smith. Brain perfusion systems for studies of drug uptake and metabolism in the central nervous system. In R. T. Borchardt, P. L. Smith, G. Wilson, (eds). Models for assessing drug absorption and metabolism,Vol. 8, New York, Plenum Press, pp. 285–307 (1996).Google Scholar
  10. 10.
    S. Cisternino, C. Rousselle, C. Dagenais, and J. M. Scherrmann. Screening of multidrug-resistance sensitive drugs by in situbrain perfusion in P-glycoprotein-deficient mice. Pharm. Res. 18:183–190 (2001).Google Scholar
  11. 11.
    M. Garrigos. L. M. Mir, and S. Orlowski. Competitive and noncompetitive inhibition of the multidrug-resistance-associated Pglycoprotein ATPase-further experimental evidence for a multisite model. Eur. J. Biochem. 244:664–673 (1997).Google Scholar
  12. 12.
    C. Martin, G. Berridge, C. F. Higgins, P. Mistry, P. Charlton, and R. Callaghan. Communication between multiple drug binding sites on P-glycoprotein. Mol. Pharmacol. 58:624–632 (2000).Google Scholar
  13. 13.
    A. B. Shapiro. K. Fox, P. Lam, and V. Ling. Stimulation of Pglycoprotein-mediated drug transport by prazosin and progesterone. Evidence for a third drug-binding site. Eur. J. Biochem. 259:841–850 (1999).Google Scholar
  14. 14.
    C. Dagenais. C. Rousselle, G. M. Pollack, and J. M. Scherrmann. Development of an in situmouse brain perfusion model and its application to mdr1a P-glycoprotein deficient mice. J. Cereb. Blood Flow Metab. 20:381–386 (2000).Google Scholar
  15. 15.
    W. D. Stein. Kinetics of the multidrug transporter (P-glycoprotein) and its reversal. Physiol. Rev. 77:545–590 (1997).Google Scholar
  16. 16.
    Y. Takasato, S. I. Rapoport, and Q. R. Smith. An in situbrain perfusion technique to study cerebrovascular transport in the rat. Am. J. Physiol. 247:H484–H493 (1984).Google Scholar
  17. 17.
    S. Cisternino, C. Rousselle, M. Debray, and J. M. Scherrmann. In vivosaturation of the transport of vinblastine and colchicine by Cisternino et al. 1388P-glycoprotein at the rat blood-brain barrier. Pharm. Res. 20: 1607–1611 (2003).Google Scholar
  18. 18.
    M. Ponec. J. Kempenaar, B. Shroot, and J. C. Caron. Glucocorticoids: binding affinity and lipophilicity. J. Pharm. Sci. 75:973–975 (1986).Google Scholar
  19. 19.
    G. Klopman, L. M. Shi, and A. Ramu. Quantitative structureactivity relationship of multidrug resistance reversal agents. Mol. Pharmacol. 52:323–334 (1997).Google Scholar
  20. 20.
    T. Litman, M. Brangi, E. Hudson, P. Fetsch, A. Abati, D. D. Ross, K. Miyake, J. H. Resau, and S. E. Bates. The multidrugresistant phenotype associated with overexpression of the new ABC half-transporter, MXR (ABCG2). J. Cell Sci. 113:2011–2021 (2000).Google Scholar
  21. 21.
    K. Tunblad, E. N. Jonsson, and M. Hammarlund-Udenaes. Morphine blood-brain barrier transport is influenced by probenecid co-administration. Pharm. Res. 20:618–623 (2003).Google Scholar
  22. 22.
    J. Van Asperen, O. Van Tellingen, A. H. Schinkel, and J. H. Beijnen. Comparative pharmacokinetics of vinblastine after a 96-hour continuous infusion in wild-type mice and mice lacking mdr1a P-glycoprotein. J. Pharmacol. Exp. Ther. 289:329–333 (1999).Google Scholar
  23. 23.
    S. Scala and N. Akhmed. U. S. Rao, K. Paull, L. B. Lan, B. Dickstein, J. S. Lee, G. H. Elgemeie, W. D. Stein, and S. E. Bates. P-glycoprotein substrates and antagonists cluster into two distinct groups. Mol. Pharmacol. 51:1024–1033 (1997).Google Scholar
  24. 24.
    A. Garrigues, N. Loiseau, M. Delaforge, J. Ferte, M. Garrigos, F. Andre, and S. Orlowski. Characterization of two pharmacophores on the multidrug transporter P-glycoprotein. Mol. Pharmacol. 62:1288–1298 (2002).Google Scholar
  25. 25.
    P. Gros, R. Dhir, J. Croop, and F. A. Talbot. Single amino acid substitution strongly modulates the activity and substrate specificity of the mouse mdr1 and mdr3 drug efflux pumps. Proc. Natl. Acad. Sci. USA 88:7289–7293 (1991).Google Scholar
  26. 26.
    F. J. Sharom, P. Lu, R. Liu, and X. Yu. Linear and cyclic peptides as substrates and modulators of P-glycoprotein: peptide binding and effects on drug transport and accumulation. Biochem. J. 333: 621–630 (1998).Google Scholar
  27. 27.
    R. Callaghan and J. R. Riordan. Synthetic and natural opiates interact with P-glycoprotein in multidrug-resistant cells. J. Biol. Chem. 268: 16059–16064 (1993).Google Scholar
  28. 28.
    S. Ekins, R. B. Kim, B. F. Leake, A. H. Dantzig, E. G. Schuetz, L. B. Lan, K. Yasuda, R. L. Shepard, M. A. Winter, J. D. Schuetz, J. H. Wikel, and S. A. Wrighton. Three-dimensional quantitative structure-activity relationships of inhibitors of P-glycoprotein. Mol. Pharmacol. 61:964–973 (2002).Google Scholar
  29. 29.
    S. Ekins, R. B. Kim, B. F. Leake, A. H. Dantzig, E. G. Schuetz, L. B. Lan, K. Yasuda, R. L. Shepard, M. A. Winter, J. D. Schuetz, J. H. Wikel, and S. A. Wrighton. Application of threedimensional quantitative structure-activity relationships of Pglycoprotein inhibitors and substrates. Mol. Pharmacol. 61:974–981 (2002).Google Scholar
  30. 30.
    R. H. Stephens, C. A. O'Neill, A. Warhurst, G. L. Carlson, M. Rowland, and G. Warhurst. Kinetic profiling of P-glycoproteinmediated drug efflux in rat and human intestinal epithelia. J. Pharmacol. Exp. Ther. 296:584–591 (2001).Google Scholar
  31. 31.
    S. V. Ambudkar, C. O. Cardarelli, I. Pashinsky, and W. D. Stein. Relation between the turnover number for vinblastine transport and for vinblastine-stimulated ATP hydrolysis by human Pglycoprotein. J. Biol. Chem. 272:21160–21166 (1997).Google Scholar
  32. 32.
    S. Orlowski, L. M. Mir, J. JR Belehradek, and M. Garrigos. Effects of steroids and verapamil on P-glycoprotein ATPase activity: progesterone, desoxycorticosterone, corticosterone and verapamil are mutually non-exclusive modulators. Biochem. J. 317: 515–522 (1996).Google Scholar
  33. 33.
    E. J. Wang, C. N. Casciano, R. P. Clement, and W. W. Johnson. Cooperativity in the inhibition of P-glycoprotein-mediated daunorubicin transport: evidence for half-of-the-sites reactivity. Arch. Biochem. Biophys. 383:91–98 (2000).Google Scholar
  34. 34.
    L. B. Lan, S. Ayesh, E. Lyubimov, I. Pashinsky, and W. D. Stein. Kinetic parameters for reversal of the multidrug pump as measured for drug accumulation and cell killing. Cancer Chemother. Pharmacol. 38:181–190 (1996).Google Scholar
  35. 35.
    M. Arboix, O. G. Paz, T. Colombo, and M. D'Incalci. Multidrug resistance-reversing agents increase vinblastine distribution in normal tissues expressing the P-glycoprotein but do not enhance drug penetration in brain and testis. J. Pharmacol. Exp. Ther. 281:1226–1230 (1997).Google Scholar
  36. 36.
    E. Lyubimov, L. B. Lan, I. Pashinsky, and W. D. Stein. Effect of modulators of the multidrug resistance pump on the distribution of vinblastine in tissues of the mouse. Anticancer Drugs 7:60–69 (1996).Google Scholar
  37. 37.
    M. Lemaire, A. Bruelisauer, P. Guntz, and H. Sato. Dosedependent brain penetration of SDZ PSC 833, a novel multidrug resistance-reversing cyclosporin, in rats. Cancer Chemother. Pharmacol. 38:481–486 (1996).Google Scholar
  38. 38.
    N. Simon, E. Dailly, O. Combes, E. Malaurie, M. Lemaire, J. P. Tillement, and S. Urien. Role of lipoproteins in the plasma binding of SDZ PSC 833, a novel multidrug resistance-reversing cyclosporin. Br. J. Clin. Pharmacol. 45:173–175 (1998).Google Scholar
  39. 39.
    W. M. Pardridge and E. M. Landaw. Tracer kinetic model of blood-brain barrier transport of plasma protein-bound ligands. Empiric testing of the free hormone hypothesis. J. Clin. Invest. 74:745–752 (1984).Google Scholar
  40. 40.
    H. Tanaka and K. Mizojiri. Drug-protein binding and blood-brain barrier permeability. J. Pharmacol. Exp. Ther. 288:912–918 (1999).Google Scholar
  41. 41.
    Y. Adachi, H. Suzuki, and Y. Sugiyama. Comparative studies on in vitromethods for evaluating in vivofunction of MDR1 Pglycoprotein. Pharm. Res. 18:1660–1668 (2001).Google Scholar

Copyright information

© Plenum Publishing Corporation 2004

Authors and Affiliations

  • Salvatore Cisternino
    • 1
  • Christophe Rousselle
    • 1
  • Marcel Debray
    • 2
  • Jean-Michel Scherrmann
    • 1
  1. 1.INSERM U26Hôpital Fernand WidalParis cedex 10France
  2. 2.Faculté de PharmacieDépartement de BiomathématiquesParisFrance

Personalised recommendations