Advertisement

Pharmaceutical Research

, Volume 22, Issue 12, pp 2051–2057 | Cite as

Development of Stealth Liposome Formulation of 2′-Deoxyinosine as 5-Fluorouracil Modulator: In Vitro and In Vivo Study

  • Raphaelle Fanciullino
  • Sarah Giacometti
  • Claude Aubert
  • Frederic Fina
  • Pierre-Marie Martin
  • Philippe Piccerelle
  • Joseph CiccoliniEmail author
Research Paper

Purpose

The aims of this study were to develop a stealth, pegylated liposomal formulation of 2′-deoxyinosine (d-Ino), a 5-fluorouracil (5-FU) modulator, to evaluate its efficacy in vitro and in tumor-bearing mice, and to study its pharmacokinetics in rats.

Method

After designing a pegylated liposome encapsulating d-Ino (L-d-Ino), we evaluated its efficacy as 5-FU modulator in vitro. Antiproliferative assays, thymidylate synthase (TS) inhibition, and apoptosis studies were carried out to check whether an optimization of 5-FU action was achieved on the 5-FU-resistant SW620 cell line. Animal pharmacokinetic and ex vivo studies were next performed to confirm that L-d-Ino displayed a slower plasma elimination pattern than free d-Ino. Finally, effects on tumor growth of L-d-Ino + 5-FU combination was evaluated in xenografted mice.

Results

We developed a stable, sterile, and homogenous 100-nm population of pegylated liposomes encapsulating 30% of d-Ino. Liposomal d-Ino exhibited a strong potential as 5-FU modulator in vitro by enhancing TS inhibition and subsequent apoptosis induction, while displaying a better pharmacokinetic profile in animals, with a near seven times clearance reduction as compared with the free form. When used in tumor-bearing mice in combination with 5-FU, our results showed next that the association led to 70% of tumor reduction with a doubling median survival time as compared with untreated animals, whereas 5-FU alone was ineffective.

Conclusion

Our data show that liposomal d-Ino, through an optimized pharmacokinetic profile, displays apotenteffect as fluoropyrimidines modulator, both in vitro and in xenografted mice. Besides, we showed here that itispossible to reverse a resistant phenotype to 5-FU, a major drug extensively described in clinical oncology.

Key Words

2′-deoxyinosine 5-FU liposome pharmacokinetics thymidylate synthase xenografts 

References

  1. 1.
    Cohen, A. M., Minsky, B. D., Schilsky, R. L. 1993Colon cancer. InDevita,  V.T.Hellman, S.Rosenberg, S. A. eds. Cancer, Principles and Practises of Oncology4th edn.JB LippincottPhiladelphia929977Google Scholar
  2. 2.
    Gotto, A. M., Belkhode, M. L., Touster, O. 1969Stimulatory effects of inosine and deoxyinosine on the incorporation of uracil-2-14-C, and 5-bromouracil-2-14-C into nucleic acids by Ehrlich ascites tumor cells in vitro Cancer Res.29807811PubMedGoogle Scholar
  3. 3.
    Evans, R. M., Laskin, J. D., Hakala, M. T. 1981Effect of excess folates and deoxyinosine on the activity and site of action of 5-fluorouracilCancer Res.4132883295PubMedGoogle Scholar
  4. 4.
    Inaba, M., Mistsuhashi, J., Sawada, H., Miike, N.,  et al. 1996Reduced activity of anabolizing enzymes in 5-fluorouracil-resistant human stomach cancer cellsJpn. Cancer Res.87212220Google Scholar
  5. 5.
    Ciccolini, J., Peillard, L., Aubert, C., Formento, P.,  et al. 2000Monitoring of the intracellular activation of 5-fluorouracil to deoxyribonucleotides in HT29 human colon cell line: application to modulation of metabolism and cytotoxicity studyFundam. Clin. Pharmacol.14147154PubMedGoogle Scholar
  6. 6.
    Perignon, J. L., Bories, D. M., Houllier, A. M., Thuillier, L.,  et al. 1987Metabolism of pyrimidine bases and nucleotides by pyrimidine-nucleoside phosphorylases in cultured human lymphoid cellsBiochim. Biophys. Acta928130136PubMedCrossRefGoogle Scholar
  7. 7.
    Singhal, D., Ho, N. F., Anderson, B. D. 1998Absorption and intestinal metabolism of purine dideoxynucleosides and an adenosine deaminase-activated prodrug of 2′,3′-dideoxyinosine in the mesenteric vein cannulated rat ileumJ. Pharm. Sci.87569577PubMedCrossRefGoogle Scholar
  8. 8.
    Kewn, S., Hoggard, P. G., Henry-Mowatt, J. S., Veal, G. J.,  et al. 1999Intracellular activation of 2′,3′-dideoxyinosine and drug interactions in vitro AIDS Res. Hum. Retrovir.15793802PubMedCrossRefGoogle Scholar
  9. 9.
    Lionetti, F. J., Fortier, N. L. 1966Metabolism of deoxyinosine by human erythrocyte ghostsBiochim. Biophys. Acta119462469PubMedGoogle Scholar
  10. 10.
    Ciccolini, J., Peillard, L., Evrard, A., Cuq, P., Aubert, C.,  et al. 2000Enhanced antitumor activity of 5-fluorouracil in combination with 2′-deoxyinosine in human colorectal cell lines and human colon tumor xenograftsClin. Cancer Res.615291535PubMedGoogle Scholar
  11. 11.
    Ciccolini, J., Cuq, P., Evrad, A., Giacometti, S., Pelegrin, A., Aubert, C., Cano, J. P., Iliadis, A. 2001Combination of thymidine phosphorylase gene transfer and deoxyinosine treatment greatly enhances 5-fluorouracil antitumor activity in vitro and in vivo Mol. Cancer Ther.1133139PubMedGoogle Scholar
  12. 12.
    Lian, T., Ho, R. 2000Trends and developments in liposome drug delivery systemsJ. Pharm. Sci.90667680CrossRefGoogle Scholar
  13. 13.
    Bangham, A. D., Standish, M. M., Watkins, J. C. 1965Diffusion of univalent ions across the lamellae of swollen phospholipidsJ. Mol. Biol.13238252PubMedGoogle Scholar
  14. 14.
    Roberts, D. 1966An isotopic assay for dihydrofolate reductaseBiochemistry535493551PubMedCrossRefGoogle Scholar
  15. 15.
    Workman, P., Balmain, A., Hickman, J. A.,  et al. 1988UKCCR guidelines for welfare of animals in experimental neoplasiaLab Anim.22195201PubMedCrossRefGoogle Scholar
  16. 16.
    Iliadis, A., Brown, A. C., Huggins, M. L. 1992APIS: a software for model identification, simulation and dosage regimen calculations in clinical and experimental pharmacokineticsComput. Methods Programs Biomed.38227239PubMedCrossRefGoogle Scholar
  17. 17.
    Myers, C. E., Young, R. C., Chabner, B. A. 1975Biochemical determinants of 5-fluorouracil response in vivo. The role of deoxyuridylate pool expansionJ. Clin. Invest.5612311238PubMedCrossRefGoogle Scholar
  18. 18.
    Myers, C. E., Diasio, R., Eliot, H. M., Chabner, B. A. 1976Pharmacokinetics of the fluoropyrimidines: implications for their clinical useCancer Treat. Rev.3175183PubMedCrossRefGoogle Scholar
  19. 19.
    Rustum, Y. M. 1999Clinical implications of 5-FU modulationOncology (Huntingt.)132225Google Scholar
  20. 20.
    Peters, G. J., Laurensse, E., Lankelma, J., Leyva, A.,  et al. 1984Separation of several 5-fluorouracil metabolites in various melanoma cell lines. Evidence for the synthesis of 5-fluorouracil-nucleotide sugarsEur. J. Cancer Clin. Oncol.2014251431PubMedCrossRefGoogle Scholar
  21. 21.
    Peters, G. J., Laurensse, E., Leyva, A., Lankelma, J.,  et al. 1986Sensitivity of human, murine, and rat cells to 5-fluorouracil and 5′-deoxy-5-fluorouridine in relation to drug-metabolizing enzymesCancer Res.462028PubMedGoogle Scholar
  22. 22.
    Schwartz, E. L., Baptiste, N., O'Connor, C. J., Wadler, S.,  et al. 1994Potentiation of the antitumor activity of 5-fluorouracil in colon carcinoma cells by the combination of interferon and deoxyribonucleosides results from complementary effects on thymidine phosphorylaseCancer Res.5414721478PubMedGoogle Scholar
  23. 23.
    Schwartz, E. L., Baptiste, N., Wadler, S., Makower, D. 1995Thymidine phosphorylase mediates the sensitivity of human colon carcinoma cells to 5-fluorouracilJ. Biol. Chem.2701907319077PubMedCrossRefGoogle Scholar
  24. 24.
    Damen, J., Regts, J., Scherphof, G. 1981Transfer and exchange of phospholipid between small unilamellar liposomes and rat plasma high density lipoproteins. Dependence on cholesterol content and phospholipid compositionBiochim. Biophys. Acta665538545PubMedGoogle Scholar
  25. 25.
    Ishida, O., Maruyama, K., Sasaki, K., Iwatsuru, M. 1999Size-dependent extravasation and interstitial localization of polyethyleneglycol liposomes in solid tumor-bearing miceInt. J. Pharm.1904956PubMedCrossRefGoogle Scholar
  26. 26.
    Harrington, K. J., Syrigos, K. N., Vile, R. G. 2002Liposomally targeted cytotoxic drugs for the treatment of cancerJ. Pharm. Pharmacol.5415731600PubMedCrossRefGoogle Scholar
  27. 27.
    Allen, T. M., Cleland, L. G. 1980Serum-induced leakage of liposome contentsBiochim. Biophys. Acta597418426PubMedCrossRefGoogle Scholar
  28. 28.
    Maruyama, K., Yuda, T., Okamoto, A., Kojima, S., Suginaka, A., Iwatsuru, M. 1992Prolonged circulation time in vivo of large unilamellar liposomes composed of distearoyl phosphatidylcholine and cholesterol containing amphipathic poly(ethylene glycol)Biochim. Biophys. Acta11284449PubMedGoogle Scholar
  29. 29.
    Allen, T. M., Hansen, C., Martin, F., Redemann, C., Yau-Young, A. 1991Liposomes containing synthetic lipid derivatives of poly(ethylene glycol) show prolonged circulation half-lives in vivo Biochim. Biophys. Acta10662936PubMedCrossRefGoogle Scholar
  30. 30.
    Hong, R. L., Huang, C. J., Tseng, Y. L., Pang, V. F., Chen, S. T., Liu, J. J., Chang, F. H. 1999Direct comparison of liposomal doxorubicin with or without polyethylene glycol coating in C-26 tumor-bearing mice: is surface coating with polyethylene glycol beneficial?Clin. Cancer Res.536453652PubMedGoogle Scholar
  31. 31.
    Gabizon, A., Shmeeda, H., Horowitz, A. T., Zalipsky, S. 2004Tumor cell targeting of liposome-entrapped drugs with phospholipid-anchored folic acid-PEG conjugatesAdv. Drug Deliv. Rev.5611771192PubMedCrossRefGoogle Scholar
  32. 32.
    Ciccolini, J., Peillard, L., Aubert, C., Formento, P.,  et al. 2000Monitoring of the intracellular activation of 5-fluorouracil to deoxyribonucleotides in HT29 human colon cell line: application to modulation of metabolism and cytotoxicity studyFundam. Clin. Pharmacol.14147154PubMedGoogle Scholar
  33. 33.
    Canman, C. E., Tang, H. Y., Normolle, D. P., Lawrence, T. S.,  et al. 1992Variations in patterns of DNA damage induced in human colorectal tumor cells by 5-fluorodeoxyuridine: implications for mechanisms of resistance and cytotoxicityProc. Natl. Acad. Sci. USA.891047410478PubMedCrossRefGoogle Scholar
  34. 34.
    Sadzuka, Y., Hirota, S., Sonobe, T. 2000Intraperitoneal administration of doxorubicin encapsulating liposomes against peritoneal disseminationToxicol. Lett.1165159PubMedCrossRefGoogle Scholar
  35. 35.
    Antimisiaris, S. G., Klepetsanis, P., Zachariou, V., Giannopoulou, E., Ioannou, P. V. 2004 In vivo distribution of arsenic after i.p. injection of arsonoliposomes in balb-c miceInt. J. Pharm.289151158PubMedCrossRefGoogle Scholar
  36. 36.
    Sadzuka, Y., Nakai, S., Miyagishima, A., Nozawa, Y., Hirota, S. 1997Effects of administered route on tissue distribution and antitumor activity of polyethyleneglycol-coated liposomes containing adriamycinCancer Lett.1117786PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science + Business Media, Inc. 2005

Authors and Affiliations

  • Raphaelle Fanciullino
    • 1
  • Sarah Giacometti
    • 1
  • Claude Aubert
    • 1
  • Frederic Fina
    • 2
  • Pierre-Marie Martin
    • 2
  • Philippe Piccerelle
    • 3
  • Joseph Ciccolini
    • 1
    Email author
  1. 1.Laboratoire de Pharmacocinétique, UFR PharmacieMarseille cedex 05France
  2. 2.EMI359UFR Médecine NordMarseilleFrance
  3. 3.Laboratoire de Pharmacie Galénique et IndustrielleUFR PharmacieMarseilleFrance

Personalised recommendations