Human Recombinant Apolipoprotein E-Enriched Liposomes Can Mimic Low-Density Lipoproteins as Carriers for the Site-Specific Delivery of Antitumor Agents

  1. Patrick C. N. Rensen1,
  2. Raymond M. Schiffelers1,
  3. A. Jenny Versluis1,
  4. Martin K. Bijsterbosch1,
  5. Marly E. M. J. Van Kuijk-Meuwissen2 and
  6. Theo J. C. Van Berkel1
  1. 1Division of Biopharmaceutics (P.C.N., R.M.S., A.J.V., M.K.B., T.J.C.V.B.) and 2Pharmaceutical Technology (M.E.M.J.M.), Leiden-Amsterdam Center for Drug Research, University of Leiden, Leiden, The Netherlands

    Abstract

    Progressive hypocholesterolemia is a feature associated with a number of cancers of different origin, and it is caused by the high expression of low-density lipoprotein (LDL) receptors (LDLrs) on many tumor cell types. Selective delivery of chemotherapeutics using LDL as a carrier has therefore been proposed, but the endogenous nature of LDL hampers its pharmaceutical application. In the current study, we explored the possibility of synthesizing liposomes that mimic LDL from commercially available lipids and proteins. Small unilamellar liposomes were created (28.9 ± 0.9 nm) and complexed with 5.8 ± 0.7 molecules of human recombinant apolipoprotein E (apoE). On intravenous injection into rats, the liposomes retained their aqueous core, structural integrity, and the majority of the preassociated apoE. [3H]Cholesteryl oleate-labeled apoE-enriched liposomes showed a relatively long serum half-life (>5 hr), and a low uptake by cells of the reticuloendothelial system was observed (<0.8% of the injected dose at 30 min after injection). Pretreatment of rats with 17α-ethinyl estradiol, which induces the expression of the LDLr on the liver and adrenals, led to a 2.5-fold accelerated serum clearance (t½ = 123 ± 10 min) and a selectively increased uptake of liposomes by the liver (2.0-fold) and adrenals (3.8-fold). The liver association of the liposomes was coupled to the lysosomal uptake route, similarly as for LDL. In vitro studies using B16 melanoma cells showed that the liposomes bound exclusively to the LDLr via their apoE moiety (90,000 liposomes/cell), with a 14-fold higher affinity (Kd = 0.77 ± 0.09 nm) than LDL itself. Because of their favorable properties, we anticipate that these apoE-enriched liposomes are advantageous compared with native LDL in the development of a selective LDLr-targeted antitumor therapy.

    Footnotes

    • Send reprint requests to: Dr. Patrick C. N. Rensen, Division of Biopharmaceutics, Leiden-Amsterdam Center for Drug Research, University of Leiden, Sylvius Laboratories, P.O. Box 9503, 2300 RA Leiden, The Netherlands. E-mail: p.rensen{at}lacdr.leidenuniv.nl

    • ↵1 P. C. N. Rensen and T. J. C. van Berkel, unpublished observations.

    • ↵2 Versluis, A.J., M.K. Bysterbosch, and T.J.C. van Berkel, unpublished observations.

    • Abbreviations:
      LDL
      low-density lipoprotein
      apo
      apolipoprotein
      BSA
      bovine serum albumin
      CO
      cholesteryl oleate
      DOPC
      dioleoyl phosphatidylcholine
      EE
      ethinyl estradiol
      EYPC
      egg yolk phosphatidylcholine
      HDL
      high-density lipoprotein
      LDLr
      low-density lipoprotein receptor
      HEPES
      4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid
      PBS
      phosphate-buffered saline
      RES
      reticuloendothelial system
      VLDL
      very low-density lipoprotein
      • Received December 2, 1996.
      • Accepted May 27, 1997.
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    1. Molecular Pharmacology September 1, 1997 vol. 52 no. 3 445-455
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