The Use of Surfactants to Enhance the Permeability of Peptides Through Caco-2 Cells by Inhibition of an Apically Polarized Efflux System
- 470 Downloads
Purpose. It has recently been reported that the permeability of peptides across Caco-2 cells, an in vitro model of the intestinal mucosa, was limited by an apically polarized efflux mechanism. Since surfactants (e.g. Cremophor EL, Polysorbate 80) have been reported to inhibit similar efflux systems in tumor cells, we determined whether they could enhance the permeability of peptides across monolayers of Caco-2 cells.
Methods. The transport studies of [3H]-mannitol and [14C]-model peptides were carried out across the Caco-2 cell monolayers. TEER values were determined using Voltohmmeter with STX-2 electrode and the equilibrium dialysis studies were conducted using side-by-side dialysis apparatus with cellulose ester membranes.
Results. Initially, [3H]-mannitol flux studies were conducted to find concentrations of the surfactants that did not cause damage to the cell monolayer. Based on these studies, Polysorbate 80 and Cremophor EL were selected for further study. The fluxes of [l4C]-AcfNH2 (a nonsubstrate for this efflux system) and [14C]-Acf(N-Mef)2NH2 (a substrate for this efflux system) were then measured in the absence and presence of the two surfactants. The permeability of [14C]-AcfNH2 was not affected by the surfactants, while that of [14C]-Acf(N-Mef)2NH2 increased with increasing concentrations of surfactants and then decreased. For example, the Pe values for [14C]-Acf(N-Mef)2NH2 were 3.75 × 10−6, 8.58 × 10−6, 10.29 × 10−6, 7.48 × 10−6, and 1.46 × 10−6 cm/sec with Cremophor EL concentrations of 0, 0.01, 0.1, 1 and 10% w/v, respectively. This bimodal effect of surfactants on the Caco-2 cell permeability of this peptide was shown to be due to the interactions between the peptide and micelles at higher concentrations of surfactants, which were demonstrated by the equilibrium dialysis experiments.
Conclusions. These results suggest that surfactants, which are commonly added to pharmaceutical formulations, may enhance the intestinal absorption of some drugs by inhibiting this apically polarized efflux system.
Unable to display preview. Download preview PDF.
- 1.P. S. Burton, R. A. Conradi, and N. F. H. Ho. Evidence for a polarized efflux system for peptides in the apical membrane of Caco-2 cells. Biochem. Biophys. Res. Commun. 190:760–766 (1993).Google Scholar
- 2.S. E. Kane, I. Pastan, and M. M. Gottesman. Genetic basis of multidrug resistance of tumor cells. J. Bioenerg. Biomembr. 22:593–618 (1990).Google Scholar
- 3.J. A. Endicott and V. Ling. The biochemistry of P-glycoprotein mediated multidrug resistance. Annu. Rev. Biochem. 58:137–171 (1989).Google Scholar
- 4.W. H. M. Peters, C. E. W. Boon, H. M. J. Roelofs, T. Wobbes, F. M. Nagengast, and P. G. Kremers. Expression of drug metabolising enzymes and P-170 glycoprotein in colorectal carcinoma and normal mucosa. Gastroenterology 103:448–455 (1992).Google Scholar
- 5.C. Cordon-Cardo, J. P. O'Brien, D. Casals, L. Rittman-Grauer, J. L. Biedler, M. R. Melamed, and J. R. Bertino. Multidrug resistance gene (P-glycoprotein) is expressed by endothelial cells at bloodbrain barrier sites. Proc. Natl. Acad. Sci. USA. 86:695–698 (1989).Google Scholar
- 6.J. Hunter, M. A. Jepson, T. Tsuruo, N. L. Simmons, and B. H. Hirst. Functional expression of P-glycoprotein in apical membrane of human intestinal Caco-2 cells. J. Biol. Chem. 268:14991–14997 (1993).Google Scholar
- 7.J. Hunter, B. H. Hirst, and N. L. Simmons. Drug absorption limited by P-glycoprotein mediated secretory drug transport in human intestinal epithelial Caco-2 cell layers. Pharm. Res. 10:743–749 (1993).Google Scholar
- 8.J. M. Zamora, H. L. Pearce, and W. T. Beck. Physical-chemical properties shared by compounds that modulate multidrug resistance in human leukemic cells. Mol. Pharmacol. 33:454–462 (1988).Google Scholar
- 9.J. M. Ford and W. N. Hait. Pharmacology of drugs that alter multidrug resistance in cancer. Pharmacol. Rev. 42:155–198 (1990).Google Scholar
- 10.D. M. Woodcock, S. Jefferson, M. E. Linsenmeyer, P. J. Crowther, G. M. Chojnwski, B. Williams, and I. Bertoncello. Reversal of the multidrug resistance phenotype with Cremophor EL, a common vehicle for water insoluble vitamins and drugs. Cancer Res. 50:4199–4203 (1990).Google Scholar
- 11.R. A. Conradi, A. R. Hilgers, N. F. H. Ho, and P. S. Burton. The influence of peptide structure on transport across Caco-2 cells. Pharm. Res. 8:1453–1460 (1991).Google Scholar
- 12.R. A. Conradi, A. R. Hilgers, N. F. H. Ho, and P. S. Burton. The influence of peptide structure on transport across Caco-2 cells. II. Peptide bond modification which results in improved permeability. Pharm. Res. 9:435–439 (1992).Google Scholar
- 13.J. Fogh, J. M. Fogh, and T. Orfeo. One hundred and twenty seven cultured human tumor cell lines producing tumors in nude mice. J. Nat. Cancer. Inst. 59:221–226 (1977).Google Scholar
- 15.P. Artursson. Epithelial transport of drugs in cell culture. I. A model for studying the passive diffusion of drugs over intestinal absorptive (Caco-2) cells. J. Pharm. Sci. 79:476–482 (1990).Google Scholar
- 16.A. R. Hilgers, R. A. Conradi, and P. S. Burton. Caco-2 cell monolayers as a model for drug transport across the intestinal mucosa. Pharm. Res. 7:902–910 (1990).Google Scholar
- 17.D. M. Woodcock, M. E. Linsenmeyer, G. M. Chojnowski, A. B. Kriegler, V. Nink, L. K. Webster, and W. H. Sawyer. Reversal of multidrug resistance by surfactants. Br. J. Cancer. 66:62–68 (1992).Google Scholar
- 18.E. Friche, P. B. Jensen, M. Sehested, E. J. F. Demant, and N. N. Nissen. The solvents Cremophor EL and Polysorbate 80 modulate daunorubin resistance in the multidrug resistant Ehrlich Ascites Tumor. Cancer Commun. 2:297–303 (1990).Google Scholar
- 19.A. Helenius and K. Simons. Solubilization of membranes by detergents. Biochim. Biophys. Acta. 415:29–79 (1975).Google Scholar
- 20.N. F. H. Ho, P. S. Burton, R. A. Conradi, and C. L. Barsuhn. A biophysical model of passive and polarized active transport processes in Caco-2 cells: approaches to uncoupling apical and basolateral membrane events in the intact cell. J. Pharm. Sci. 84:21–27 (1995).Google Scholar
- 21.T. Zordan-Nudo, V. Ling, Z. Liu, and E. Georges. Effects of nonionic detergents on P-glycoprotein drug binding and reversal of mutidrug resistance. Cancer Res. 53:5994–6000 (1993).Google Scholar
- 22.M. M. Gottesman and I. Pastan. Biochemistry of multidrug resistance mediated by multidrug transporter. Annu. Rev. Biochem. 62:385–427 (1993).Google Scholar