Skip to main content
SpringerLink
Log in

A Comparison of the Bioconversion Rates and the Caco-2 Cell Permeation Characteristics of Coumarin-Based Cyclic Prodrugs and Methylester-Based Linear Prodrugs of RGD Peptidomimetics

  • Published:
Pharmaceutical Research Aims and scope Submit manuscript

Abstract

Purpose. To compare the bioconversion rates in various biological media and the Caco-2 cell permeation characteristics of coumarin-based cyclic prodrugs (3a, 3b) and methylester-based linear prodrugs (1b, 2b) of two RGD peptidomimetics (la, 2a).

Methods. Bioconversion rates of the prodrugs to the RGD peptidomimetics were determined in Hank balanced salt solution (HBSS), pH 7.4, at 37°C and in various biological media (human blood plasma, rat liver homogenate, Caco-2 cell homogenate) known to have esterase activity. Transport rates of the prodrugs and the RGD peptidomimetics were determined using Caco-2 cell monolayers, an in vitrocell culture model of the intestinal mucosa.

Results. In HBSS, pH 7.4, the coumarin-based cyclic prodrugs 3a and 3b degraded slowly and quantitatively to the RGD peptidomimetics la and 2a, respectively (3a, t1/2= 630 ± 14 min; 3b, t1/2= 301 ± 12 min). The methylester-based linear prodrugs 1b and 2b were more stable to chemical hydrolysis (1b and 2b, t1/2> 2000 min). Both the coumarin-based cyclic prodrugs and the methylester-based linear prodrugs degraded more rapidly in biological media containing esterase activity (e.g., 90% human blood plasma: 1b, t1/2< 5 min; 2b, t1/2< 5 min; 3a, t1/2< 91 ± 1 min; 3b, t1/2< 57 ± 2 min). When the apical (AP)-to-basolateral (BL) permeation characteristics were determined using Caco-2 cell monolayers, it was found that the methylester pro-drugs Ib and 2b underwent esterase bioconversion (>80%) to the RGD peptidomimetics 1a and 2a, respectively. In contrast, the cyclic prodrugs 3a and 3b permeated the cell monolayers intact. Considering the appearance of both the prodrug and the RGD peptidomimetic on the BL side, the methylester prodrugs 1b and 2b were approximately 12-fold more able to permeate than were the RGD peptidomimetics la and 2a. When a similar analysis of the transport data for the coumarin prodrugs 3a and 3b was performed, they were shown to be approximately 6-fold and 5-fold more able to permeate than were the RGD peptidomimetics la and 2a, respectively.

Conclusions. The coumarin-based cyclic prodrugs 3a and 3b were chemically less stable, but metabolically more stable, than the methylester-based linear prodrugs. The esterase stability of the cyclic prodrugs 3a and 3b means that they are transported intact across the Caco-2 cell monolayer in contrast to the methylester prodrugs 1b and 2b, which undergo facile bioconversion during their transport to the RGD peptidomimetics. However, both prodrug systems successfully delivered more (5-12-fold) of the RGD peptidomimetic and/or the precursor (prodrug) than did the RGD peptidomimetics themselves.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

REFERENCES

  1. G. D. Hartman, M. S, Egbertson, W. Halczenko, W. L. Laswell, M. E. Duggan, R. L. Smith, A. M. Naylor, P. D. Manno, R. J. Lynch, G. Zhang, C. T. Chang, and R. J. Gould. Non-peptide fibrinogen receptor antagonists. 1. discovery and design of exosite inhibitors. J. Med. Chem. 35:4640-4642 (1992).

    PubMed  Google Scholar 

  2. D. R. Philips, I. F. Charo, L. V. Parise, and L. A. Fitzgerald. The platelet membrane glycoprotein IIb-IIIa complex. Blood 71:831-843 (1988).

    PubMed  Google Scholar 

  3. J. Zablocki, N. Nicholson, and B. Taite. Selection of an orally active glycoprotein IIb/IIIa receptor antagonist for clinical trials. Thromb. Haemostas. 69:1244 (1993).

    Google Scholar 

  4. H. U. Stilz, B. Jabloka, M. Just, J. Knolle, E. F. Paulus, and G. Zoller. Discovery of an orally active non-peptide fibrinogen receptor antagonist. J. Med. Chem. 39:2118-2122 (1996).

    PubMed  Google Scholar 

  5. V. Austel, F. Himmelsbach, and T. Muller. Nonpeptidic fibrinogen receptor antagonists. Drugs of Future 19:757-764 (1984).

    Google Scholar 

  6. J. A. Zablocki, M. Miyano, R. B. Garland, D. Pireh. L. Schretzman, S. N. Rao, R. J. Lindmark, S. G. Panzer-Knodle, N. S. Nicholson, B. B. Taite, A. K. Salyers, L. W. King, J. G. Campion, and L. P. Geigen. Potent in vitro and in vivo inhibitors of platelet aggregation based on the Arg-Gly-Asp sequence of fibrinogen. A proposal on the nature of the binding interaction between the Arg-guanidine of RGDX mimetics and the platelet GP IIb-IIIa receptor. J. Med. Chem. 36:1813-1819 (1993).

    Google Scholar 

  7. J. S. Barrett, G. Murphy, K. Peerlinck, I. DeLepeleire, R. J. Gould, D. Panbianco, E. Hand, H. Deckmyn, J. Vermylen, and J. Arnout. Pharmacokinetics and pharmacodynamics of MK-383, a selective non-peptide platelet glycoprotein-IIb/IIIa receptor antagonist, in healthy men. Clin. Pharmacol. Ther. 56:377-388 (1994).

    PubMed  Google Scholar 

  8. S. Vickers. C. A. Duncan, A. S. Yuan, and K. P. Vyas. Disposition of MK-852, a fibrinogen receptor antagonist, in rats and dogs. Drug Metab. Dispos. 22:631-636 (1994).

    PubMed  Google Scholar 

  9. T. Prueksaritanont. L. M. Gorham. J. D. Ellis, C. Fernandez-Metzler, P. Deluna, J. R. Gehret. K. L. Strong, J. H. Hochman, B. C. Askew, M. E. Duggan, J. D. Gilbert, J. H. Lin, and K. P. Vyas. Species and organ differences in first-pass metabolism of the ester prodrug L-751, 164 in dogs and monkeys. In vivo and in vitro studies. Drug Metab. Dispos. 24:1263-1271 (1996).

    PubMed  Google Scholar 

  10. T. Prueksaritanont, L. M. Gorham, J. A. Naue, T. G. Hamill, B. C. Askew, and K. P. Vyas. Disposition of L-738, 167, a potent and long-acting fibrinogen receptor antagonist, in dogs. Drug Metab. Dispos. 25:355-361 (1997).

    PubMed  Google Scholar 

  11. J. S. Barrett, R. J. Gould, J. D. Ellis, M. M. Holahan, M. T. Stranieri, J. J. Lynch, Jr., G. D. Hartman, N. Ihle, M. Duggan, O. A. Moreno, and A. D. Theoharides. Pharmacokinetics and pharmacodynamics of L-303,014, a potent fibrinogen receptor antagonist, after intravenous and oral administration in the dog. Pharm. Res. 11:426-431 (1994).

    PubMed  Google Scholar 

  12. N. B. Modi, S. A. Baughman, B. D. Paasch, A. Celnitur, and S. Y. Smith. Pharmacokinetics and pharmacodynamics of TP-9201, a GPIIb/IIIa antagonist, in rats and dogs. J. cardiovasc. Pharmacol. 25:888-897 (1995).

    PubMed  Google Scholar 

  13. J. H. Hutchinson, J. J. Cook, K. M. Brashear, M. J. Breslin, J. D. Glass, R. J. Gould, W. Halczenko, M. A. Holahan, R. J. Lynch, G. R. Sitko, M. T. Stranieri, and G. D. Hartman. Non-peptide glycoprotein IIb/IIIa antagonists. 11. Design and in vivo evaluation of 3,4-dihydro-1(1H)-isoquinolinone-based antagonists and ethyl ester prodrugs. J. Med. Chem. 39:4583-4591 (1996).

    PubMed  Google Scholar 

  14. T. Weller, L. Alig, M. Beresini, B. Blackburn, S. Bunting, P. Hadvary, M. Hürzeler Müller, D. Knopp, B. Levet-Trafit, M. Terry Lipari. N. B. Modi, M. Müller, C. J. Refino, M. Schmitt, P. Schönholzer, S. Weiss, and B. Steiner. Orally active fibrinogen receptor antagonists. 2. Amidoximes as prodrugs of amidines. J. Med. Chem. 39:3139-3147 (1996).

    PubMed  Google Scholar 

  15. S. Gangwar, G. M. Pauletti, B. Wang, T. J. Siahaan, V. S. Stella, and R. T. Borchardt. Prodrug strategies to enhance the intestinal absorption of peptides. Drug Disc. Today 2:22-29 (1997).

    Google Scholar 

  16. B. Wang, S. Gangwar, G. M. Pauletti, T. J. Siahaan, and R. T. Borchardt. Synthesis of a novel esterase-sensitive cyclic prodrug system for peptides that utilizes a trimethyl lock-facilitated lactonization reaction. J. Org. Chem. 62:1363-1367 (1997).

    Google Scholar 

  17. G. M. Pauletti, S. Gangwar, F. W. Okumu, T. J. Siahaan, V. J. Stella, and R. T. Borchardt. Esterase-sensitive cyclic prodrugs of peptides: Evaluation of an acyloxyalkoxy promoiety in a model hexapeptide. Pharm. Res. 13:1615-1623 (1996).

    PubMed  Google Scholar 

  18. G. M. Pauletti, S. Gangwar, B. Wang, and R. T. Borchardt. Esterase-sensitive cyclic prodrugs of peptides: Evaluation of a phenyl-propionic acid promoiety in a model hexapeptide. Pharm. Res. 14:11-17 (1997).

    PubMed  Google Scholar 

  19. B. Wang, W. Wang, and H. Zhang. Chemical feasibility studies of a potential coumarin-based prodrug system. Bioorg. Med. Chem. Lett. 6:945-950 (1996).

    Google Scholar 

  20. B. Wang, W. Wang, H. Zhang, D. Shan, and T. D. Smith. Coumarin-based prodrugs. 2. Synthesis and bioreversibility studies of an esterase-sensitive cyclic prodrug of DADLE, an opioid peptide. Bioorg. Med. Chem. Lett. 6:2823-2826 (1996).

    Google Scholar 

  21. B. Wang, W. Wang, G. Camenisch, J. Elmo, H. Zhang, and R. T. Borchardt. Coumarin-based prodrugs 4. Synthesis and evaluation of esterase-sensitive cyclic prodrugs of antithrombotic RGD analogs. Bioorg. Med. Chem.: submitted (1998).

  22. I. J. Hidalgo, T. J. Raub, and R. T. Borchardt. Biochemical, histological and physicochemical characterization of human adenocarcinoma cells (Caco-2) as a model system for studying mucosal transport and metabolism of drugs. Gastroenterology 96:736-749 (1989).

    PubMed  Google Scholar 

  23. R. Oliyai. Prodrugs of peptides and peptidomimeties for improved formulation and delivery. Adv. Drug Delivery Rev. 19:275-286 (1996).

    Google Scholar 

  24. W. N. Aldrige. Serum esterases. 1. Two types of esterase (A and B) hydrolysing p-nitrophenyl acetate, proprionate and butyrate, and a method for their determination. Biochem. J. 53:110-117 (1953).

    PubMed  Google Scholar 

  25. D. s. Auld and B. Holmquist. Carboxypeptidase A. Differences in the mechanism of ester and peptide hydrolysis. Biochemistry 13:4355-4361 (1974).

    PubMed  Google Scholar 

  26. G. M. Pauletti, S. Gangwar, G. T. Knipp, M. M. Nerukar. F. W. Okumu. K. Tamura, T. J. Siahaan, and R. T. Borchardt. Structural requirements for intestinal absorption of peptide drugs. J. Controlled Rel. 41:3-17 (1996).

    Google Scholar 

  27. P. S. Burton, R. A. Conradi, A. R. Hilgers, 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. 190:760-765 (1993).

    PubMed  Google Scholar 

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

    PubMed  Google Scholar 

  29. H. Van de Waterbeemd. G. Camenisch. G. Folkers, and O. A. Raevsky. Estimation of Caco-2 cell permeability using calculated molecular descriptors. Quant. Struct. Act. Relat. 15:480-490 (1996).

    Google Scholar 

  30. G. Camenisch, H. Van de Waterbeemd. and G. Folkers. Review on theoretical passive drug absorption models. Pharm. Acta Helv. 71:309-327 (1996).

    PubMed  Google Scholar 

  31. P. Artursson. Cell cultures as models for drug absorption across the intestinal mucosa. Crit. Rev. Ther. Drug Carrier Syst. 8:305-330 (1991).

    PubMed  Google Scholar 

  32. S. Gan, C. Eads, T. Niederer, A. Bridgers, S. Yanni, P. H. Hsyu, F. J. Pritchard, and D. Thakker. Use of Caco-2 as an in-vitro intestinal absorption and metabolism model. Drug Develop. Ind. Pharm. 20:615-631 (1994).

    Google Scholar 

  33. K. M. Hillgren, A. Kato, and R. T. Borchardt. In vitro systems for studying intestinal drug absorption. Med. Res. Rev. 15:83-109 (1995).

    PubMed  Google Scholar 

  34. S. Ong, H. Liu, X. Qiu, G. Bhat, and Ch. Pidgeon. Membrane partition coefficients chromatographically measured using immobilized artificial surfaces. Anal. Chem. 67:755-762 (1995).

    PubMed  Google Scholar 

  35. F. Barbato. M. La Rotonda, and F. Quaglia. Chromatographic indices determined on an immobilized artificial membrane (IAM) column as descriptors of lipophilic and polar interactions of 4-phenyldihydropyridine calcium-channel blockers with biomembranes. Eur. J. Med. Chem. 31:311-318 (1996).

    Google Scholar 

  36. N. El Tayar, H. Van de Waterbeemd, and B. Testa. Lipophilicity measurements of protonated basic compounds by reversed-phased high-performance liquid chromatography. I. Relationship between capacity factors and the methanol concentration in methanolwater eluents. J. Chromatogr. 320:293-304 (1985).

    Google Scholar 

  37. N. EI Tayar, H. Van de Waterbeemd, and B. Testa. Lipophilicity measurements of protonated basic compounds by reversed-phased high-performance liquid chromatography. II. Procedure for the determination of a lipophilic index measured by reversed-phased high-performance liquid chromatography. J. Chromatogr. 320:305-312 (1985).

    Google Scholar 

  38. 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. 7:902-909 (1990).

    PubMed  Google Scholar 

  39. P. Annaert, R. Kinget, L. Naesens, E. de Clerq, and P. Augustijns, Transport, uptake, and metabolism of the bis(pivaloxymethyl)-ester prodrug of 9-(2-phosphonyl-methoxyethyl)adenine in an in vitro cell culture system of the intestinal mucosa (Caco-2). Pharm. Res. 14:492-496 (1995).

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Pharmaceutical Chemistry, The University of Kansas, Lawrence, Kansas, 66047

    Gian P. Camenisch & Ronald T. Borchardt

  2. Department of Chemistry, North Carolina State University, Raleigh, North Carolina, 27695-8204

    Wei Wang & Binghe Wang

Authors
  1. Gian P. Camenisch
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Wei Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Binghe Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Ronald T. Borchardt
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Camenisch, G.P., Wang, W., Wang, B. et al. A Comparison of the Bioconversion Rates and the Caco-2 Cell Permeation Characteristics of Coumarin-Based Cyclic Prodrugs and Methylester-Based Linear Prodrugs of RGD Peptidomimetics. Pharm Res 15, 1174–1181 (1998). https://doi.org/10.1023/A:1011975404789

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1023/A:1011975404789

Search

Navigation