Pharmaceutical Research

, Volume 30, Issue 5, pp 1281–1290 | Cite as

In Vitro and In Vivo Evaluation of Lipofufol, a New Triple Stealth Liposomal Formulation of Modulated 5-Fu: Impact on Efficacy and Toxicity

  • Raphaelle Fanciullino
  • Séverine Mollard
  • Sarah Giacometti
  • Yael Berda-Haddad
  • Mohamed Chefrour
  • Claude Aubert
  • Athanassios Iliadis
  • Joseph CiccoliniEmail author
Research Paper



Drug resistance and severe toxicities are limitations when handling 5-FU. We have developed a triple liposomal formulation of 5-FU combined to 2′-deoxyinosine and folinic acid to improve its efficacy-toxicity balance.


Stealth liposomes were obtained using the thin-film method. Antiproliferative activity was tested on human colorectal and breast cancer models using sensitive (HT29) and resistant (SW620, LS174t, MDA231) cell lines. In vivo, pharmacokinetics, biodistribution and safety studies were performed in rodents. Finally, efficacy was evaluated using two tumor-bearing mice models (LS174 and MDA231) with response and survival as main endpoints.


LipoFufol is a 120-nm pegylated liposome, displaying 20–30% encapsulation rates. In vitro, antiproliferative activities were higher than 5-FU, and matched that of FolFox combination in colorectal models, but not in breast. Drug monitoring showed an optimized pharmacokinetics profile with reduced clearance and prolonged half-life. Liposome accumulation in tumors was shown by fluorescence-based biodistribution studies. Beside, milder neutropenia was observed when giving LipoFufol to animals with transient partial DPD-deficiency, as compared with standard 5-FU. In LS174t-bearing mice, higher response and 55% longer survival were achieved with Lipofufol, as compared with 5-FU.


The issues of drug-resistance and drug-related toxicity can be both addressed using a stealth liposomal formulation of modulated 5-FU.


5-FU colorectal cancer pharmacokinetics stealth liposomes toxicity 



This study was supported by generous grants from the GEFLUC Marseille-Provence and the Association pour la Recherche sur le Cancer (ARC, Grant #1026).


  1. 1.
    Longley DB, Harkin DP, Johnston PG. 5-Fluorouracil: mechanisms of action and clinical strategies. Nat Rev Cancer. 2003;3:330–8.PubMedCrossRefGoogle Scholar
  2. 2.
    Zhang N, Yin Y, Xu SJ, Chen WS. 5-Fluorouracil: mechanisms of resistance and reversal strategies. Molecules. 2008;13:1551–69.PubMedCrossRefGoogle Scholar
  3. 3.
    Yang CG, Ciccolini J, Blesius A, Dahan L, Bagarry-Liegey D, Brunet C, Varoquaux A, Frances N, Marouani H, Giovanni A, Ferri-Dessens RM, Chefrour M, Favre R, Duffaud F, Seitz JF, Zanaret M, Lacarelle B, Mercier C. DPD-based adaptive dosing of 5-FU in patients with head and neck cancer: impact on treatment efficacy and toxicity. Cancer Chemother Pharmacol. 2011;67:49–56.PubMedCrossRefGoogle Scholar
  4. 4.
    Zamboni WC. Concept and clinical evaluation of carrier-mediated anticancer agents. Oncologist. 2008;13:248–60.PubMedCrossRefGoogle Scholar
  5. 5.
    Egusquiaguirre SP, Igartua M, Hernández RM, Pedraz JL. Nanoparticle delivery systems for cancer therapy: advances in clinical and preclinical research. Clin Transl Oncol. 2012;14:83–93.PubMedCrossRefGoogle Scholar
  6. 6.
    Paolo D. A, liposomal anticancer therapy: pharmacokinetic and clinical aspects. J Chemother. 2004;16:90–3.PubMedGoogle Scholar
  7. 7.
    Ciccolini J, Peillard L, Evrard A, Cuq P, Aubert C, Pelegrin A, Formento P, Milano G, Catalin J. Enhanced antitumor activity of 5-fluorouracil in combination with 2′-deoxyinosine in human colorectal cell lines and human colon tumor xenografts. Clin Cancer Res. 2000;6:1529–35.PubMedGoogle Scholar
  8. 8.
    Ciccolini J, Cuq P, Evrard A, Giacometti S, Pelegrin A, Aubert C, Cano JP, Iliadis A. Combination of thymidine phosphorylase gene transfer and deoxyinosine treatment greatly enhances 5-fluorouracil antitumor activity in vitro and in vivo. Mol Cancer Ther. 2001;1:133–9.PubMedGoogle Scholar
  9. 9.
    Fanciullino R, Giacometti S, Aubert C, Fina F, Martin PM, Piccerelle P, Ciccolini J. Development of stealth liposome formulation of 2′-deoxyinosine as 5-fluorouracil modulator: in vitro and in vivo study. Pharm Res. 2005;22(12):2051–7.PubMedCrossRefGoogle Scholar
  10. 10.
    Fanciullino R, Giacometti S, Mercier C, Aubert C, Blanquicett C, Piccerelle P, Ciccolini J. In vitro and reversal of resistance to 5-fluorouracil in colorectal cancer cells with a novel stealth double-liposomal formulation. Br J Cancer. 2007;97:919–26.PubMedGoogle Scholar
  11. 11.
    Alley MC, Scudiero DA, Monks A, Hursey ML, Czerwinski MJ, Fine DL, Abbott BJ, Mayo JG, Shoemaker RH, Boyd MR. Feasability of drug screening with panels of human tumor cell lines using a microculture tetrazolium assay. Cancer Res. 1988;48:589–601.PubMedGoogle Scholar
  12. 12.
    Waterhouse DN, Madden TD, Cullis PR, Bally MB, Mayer LD, Webb MS. Preparation, characterization, and biological analysis of liposomal formulations of vincristine. Methods Enzymol. 2005;391:40–57.PubMedCrossRefGoogle Scholar
  13. 13.
    Blanco E, Hsiao A, Ruiz-Esparza GU, Landry MG, Meric-Bernstam F, Ferrari M. Molecular-targeted nanotherapies in cancer: enabling treatment specificity. Mol Oncol. 2011;5:492–503.PubMedCrossRefGoogle Scholar
  14. 14.
    Wang M, Thanou M. Targeting nanoparticles to cancer. Pharmacol Res. 2010;62:90–9.PubMedCrossRefGoogle Scholar
  15. 15.
    Evrard A, Cuq P, Ciccolini J, Vian L, Cano JP. Increased cytotoxicity and bystander effect of 5-fluorouracil and 5-deoxy-5-fluorouridine in human colorectal cancer cells Transfected with thymidine phosphorylase. Br J Cancer. 1999;80:1726–33.PubMedCrossRefGoogle Scholar
  16. 16.
    Petit E, Milano G, Lévi F, Thyss A, Bailleul F, Schneider M. Circadian rhythm-varying plasma concentration of 5-fluorouracil during a 5-day continuous venous infusion at a constant rate in cancer patients. Cancer Res. 1988;48:1676–9.PubMedGoogle Scholar
  17. 17.
    Etienne-Grimaldi MC, Cardot JM, François E, Renée N, Douillard JY, Gamelin E, Milano G. Chronopharmacokinetics of oral tegafur and uracil in colorectal cancer patients. Clin Pharmacol Ther. 2088;83:413–5.CrossRefGoogle Scholar
  18. 18.
    Bocci G, Di Paolo A, Barbara C, Masi G, Fornaro L, Loupakis F, Allegrini G, Falcone A, Del Tacca M, Danesi R. Pharmacokinetics, a main actor in a many-sided approach to severe 5-FU toxicity prediction. Br J Clin Pharmacol. 2009;67:132–4.PubMedCrossRefGoogle Scholar
  19. 19.
    Kovoor PA, Karim SM, Marshall JL. Is levoleucovorin an alternative to racemic leucovorin? a literature review. Clin Colorectal Cancer. 2009;8:200–6.PubMedCrossRefGoogle Scholar
  20. 20.
    de Gramont A, Louvet C, André T, Tournigand C, Raymond E, Molitor JL, Krulik M, Modulation of 5-fluorouracil with folinic acid in advanced colorectal cancers. Groupe d'étude et de recherche sur les cancers de l'ovaire et digestifs (GERCOD). Rev Med Interne. 4 (1997) 372 s-378s.Google Scholar
  21. 21.
    Zamboni WC. Liposomal, nanoparticle, and conjugated formulations of anticancer agents. Clin Cancer Res. 2005;11:8230–4.PubMedCrossRefGoogle Scholar
  22. 22.
    Thomas AM, Kapanen AI, Hare JI, Ramsay E, Edwards K, Karlsson G, Bally MB. Development of a liposomal nanoparticle formulation of 5-fluorouracil for parenteral administration: formulation design, pharmacokinetics and efficacy. J Control Release. 2011;150:212–9.PubMedCrossRefGoogle Scholar
  23. 23.
    Arias JL, Clares B, Morales ME, Gallardo V, Ruiz MA. Lipid-based drug delivery systems for cancer treatment. Curr Drug Targets. 2011;12:1151–65.PubMedCrossRefGoogle Scholar
  24. 24.
    Takeuchi H, Kojima H, Yamamoto H, Kawashima Y. Evaluation of circulation profiles of liposomes coated with hydrophilic polymers having different molecular weights in rats. J Control Release. 2001;75:83–91.PubMedCrossRefGoogle Scholar
  25. 25.
    Moghimi SM, Szebeni J. Stealth liposomes and long circulating nanoparticles: critical issues in pharmacokinetics, opsonization and protein-binding properties. Prog Lipid Res. 2003;42:463–78.PubMedCrossRefGoogle Scholar
  26. 26.
    Milla P, Dosio F, Cattel L. PEGylation of proteins and liposomes: a powerful and flexible strategy to improve the drug delivery. Curr Drug Metab. 2012;2012(13):105–19.CrossRefGoogle Scholar
  27. 27.
    Diasio RB, Harris BE. Clinical pharmacology of 5-fluorouracil. Clin Pharmacokinet. 1989;16:215–37.PubMedCrossRefGoogle Scholar
  28. 28.
    Ciccolini J, Gross E, Dahan L, Lacarelle B, Mercier C. Routine dihydropyrimidine dehydrogenase testing for anticipating 5-fluorouracil-related severe toxicities: hype or hope? Clin Colorectal Cancer. 2010;9:224–8.PubMedCrossRefGoogle Scholar
  29. 29.
    Schmoll HJ. Dihydropyrimidine dehydrogenase inhibition as a strategy for the oral administration of 5-fluorouracil: utility in the treatment of advanced colorectal cancer. Anticancer Drugs. 2003;14:695–702.PubMedCrossRefGoogle Scholar
  30. 30.
    Fanciullino R, Ciccolini J. Liposome-encapsulated anticancer drugs: still waiting for the magic bullet? Curr Med Chem. 2009;16:4361–71.PubMedCrossRefGoogle Scholar
  31. 31.
    Ciccolini J, Mercier C, Evrard A, Dahan L, Boyer JC, Duffaud F, Richard K, Blanquicett C, Milano G, Blesius A, Durand A, Seitz JF, Favre R, Lacarelle B. A rapid and inexpensive method for anticipating severe toxicity to fluorouracil and fluorouracil-based chemotherapy. Ther Drug Monit. 2006;28:678–85.PubMedCrossRefGoogle Scholar
  32. 32.
    Meta-analysis Group In Cancer. Efficacy of intravenous continuous infusion of fluorouracil compared with bolus administration in advanced colorectal cancer. J Clin Oncol. 1998;16:301–8.Google Scholar
  33. 33.
    Ishikawa T, Sekiguchi F, Fukase Y, Sawada N, Ishitsuka H. Positive correlation between the efficacy of capecitabine and doxifluridine and the ratio of thymidine phosphorylase to dihydropyrimidine dehydrogenase activities in tumors in human cancer xenografts. Cancer Res. 1998;58:685–90.PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  • Raphaelle Fanciullino
    • 1
  • Séverine Mollard
    • 2
  • Sarah Giacometti
    • 1
  • Yael Berda-Haddad
    • 3
  • Mohamed Chefrour
    • 2
  • Claude Aubert
    • 2
  • Athanassios Iliadis
    • 1
  • Joseph Ciccolini
    • 1
    Email author
  1. 1.Pharmacokinetics Unit, School of PharmacyAix Marseille UniversityMarseilleFrance
  2. 2.UMR-MD3, Aix-Marseille UniversityMarseilleFrance
  3. 3.Haematology UnitLa Conception University Hospital of MarseilleMarseilleFrance

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