title: Efficient delivery of an antisense oligodeoxyribonucleotide formulated in folate receptor-targeted liposomes.
abstract: BACKGROUND
  Folate receptors (FRs) are cellular surface markers for numerous solid tumors and myeloid leukemias. The aim of this study was to develop an antisense oligodeoxyribonucleotide (ODN) carrier targeting FR-overexpressing cancer cells using folate (FA) as the targeting moiety. G3139, a phosphorothioate antisense ODN against human bcl2 mRNA, was evaluated in this study.
  
  
  METHODS
  G3139-containing liposomes were prepared using an ethanol dilution method. For the targeted formulation, 0.5 mol% of folate-PEG-DSPE was incorporated as a targeting ligand into cationic liposomes composed of DC-Chol/egg PC/PEG-DSPE at 25:65:10 mol/mol. Particle size and surface charge were measured and cellular uptake was assessed by fluorescence microscopy and flow cytometry. The ODN-containing formulations were evaluated in FR+ KB cells for Bcl2 down-regulation measured by Western blot. The cytotoxicity of the formulations was determined by MTT assay.
  
  
  RESULTS
  The G3139-containing liposomes had an average diameter of 80-90 nm with high ODN entrapment efficiency (70-80%). Incorporation of the folate ligand did not significantly alter the particle size and entrapment efficiency. The formulation exhibited colloidal stability in a serum-containing environment. In uptake studies, the folate-targeted formulation showed ligand concentration-dependent uptake that was up to 6-fold more efficient than that of the non-targeted formulation (p < 0.05). The uptake could be blocked by an excess amount of free folate, thus indicating an FR-dependent mechanism.
  
  
  CONCLUSIONS
  FR-targeted G3139-containing liposomes showed promising transfection activity in KB cells. FR-targeted formulations were capable of specific targeting to FR-overexpressing cell lines and optimizing the amount of folate ligand in the liposomal formulation can result in more efficient antisense delivery.
authors.affiliation: Division of Pharmaceutics, College of Pharmacy, The Ohio State University, Columbus, OH 43210, USA.
authors.forename: Shih-Jiuan
authors.initials: SJ
authors.lastname: Chiu
authors.forename: Guido
authors.initials: G
authors.lastname: Marcucci
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Lee
mesh_terms: D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism*; D003630:Daunorubicin / Q000494:pharmacology; D015536:Down-Regulation; D058973:Folate Receptors, GPI-Anchored; D005492:Folic Acid / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D007624:KB Cells; D008081:Liposomes / Q000008:administration & dosage / Q000138:chemical synthesis / Q000737:chemistry; D008856:Microscopy, Fluorescence; D010316:Particle Size; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D011956:Receptors, Cell Surface / Q000378:metabolism*; D013873:Thionucleotides / Q000008:administration & dosage* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D000903:Antibiotics, Antineoplastic; D002352:Carrier Proteins; D058973:Folate Receptors, GPI-Anchored; D008081:Liposomes; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D013873:Thionucleotides; C408162:oblimersen; D005492:Folic Acid; D003630:Daunorubicin