title: Development of a miR-92a delivery system for anti-angiogenesis-based cancer therapy.
abstract: BACKGROUND
  RNA interference has received much attention as a novel therapeutic strategy. MicroRNA (miRNA) appears to be promising as a novel nucleic-acid medicine because it is able to suppress a series of protein expression that relates to a specific event such as angiogenesis. In the present study, we used dicetyl phosphate-tetraethylenepentamine-based polycation liposomes (TEPA-PCL) as a delivery system for miR-92a, one of the miRNAs regulating angiogenesis, and attempted to deliver miR-92a to angiogenic endothelial cells for the development of cancer therapy by anti-angiogenesis.
  
  
  METHODS
  Cholesterol-grafted miR-92a (miR-92a-C) was bound to TEPA-PCL, and the ratio of nitrogen of TEPA-PCL to phosphorus of miR-92a-C (N/P ratio) was optimized. This complex was transfected into human umbilical vein endothelial cells (HUVECs), and the intracellular localization of miR-92a-C was observed under a confocal laser-scanning microscope by the use of fluorescein isothiocyanate-labeled miR-92a-C. After transfection of HUVECs with miR-92a-C/TEPA-PCL, the expression of miR-92a-target proteins (e.g. integrin α5, mitogen-activated protein kinase kinase 4, sphingosine-1-phosphate receptor 1) was examined by western blotting, and a tube formation assay was performed.
  
  
  RESULTS
  The complex of miR-92a-C with TEPA-PCL was formed and miR-92a-C remained stable with TEPA-PCL at the N/P ratio of 10. After transfection of HUVECs with miR-92a-C complex, miR-92a-C spread into the whole cytoplasm of the cells without any change of cellular morphology, and the expression of several proteins encoded by miR-92a-target genes was suppressed. Furthermore, the capability of forming capillary tubes was impaired in complex-treated HUVECs.
  
  
  CONCLUSIONS
  We have developed a miR-92a delivery system into angiogenic endothelial cells by the use of TEPA-PCL. These results suggest that miR-92a-C/TEPA-PCL is promising for the treatment of tumors via the suppression of angiogenesis.
authors.affiliation: Department of Medical Biochemistry, University of Shizuoka Graduate School of Pharmaceutical Sciences, Shizuoka, Japan.
authors.forename: Hidenori
authors.initials: H
authors.lastname: Ando
authors.forename: Ayaka
authors.initials: A
authors.lastname: Okamoto
authors.forename: Masafumi
authors.initials: M
authors.lastname: Yokota
authors.forename: Kosuke
authors.initials: K
authors.lastname: Shimizu
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Asai
authors.forename: Takehisa
authors.initials: T
authors.lastname: Dewa
authors.forename: Naoto
authors.initials: N
authors.lastname: Oku
mesh_terms: D020533:Angiogenesis Inhibitors / Q000378:metabolism*; D015153:Blotting, Western; D002478:Cells, Cultured; D005029:Ethylenediamines / Q000494:pharmacology; D005786:Gene Expression Regulation; D055785:Gene Knockdown Techniques; D018014:Gene Transfer Techniques; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D039482:Integrin alpha5 / Q000235:genetics / Q000378:metabolism; D008081:Liposomes; D048670:MAP Kinase Kinase 4 / Q000235:genetics / Q000378:metabolism; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D018613:Microscopy, Confocal; D009369:Neoplasms / Q000628:therapy; D009389:Neovascularization, Pathologic / Q000628:therapy; D034622:RNA Interference; D049349:Receptors, Lysosphingolipid / Q000235:genetics / Q000378:metabolism; D000081025:Sphingosine-1-Phosphate Receptors
chemical_list: D020533:Angiogenesis Inhibitors; D005029:Ethylenediamines; D039482:Integrin alpha5; D008081:Liposomes; C541874:MIRN92 microRNA, human; D035683:MicroRNAs; D049349:Receptors, Lysosphingolipid; C477362:S1PR1 protein, human; D000081025:Sphingosine-1-Phosphate Receptors; D048670:MAP Kinase Kinase 4; C034269:tetraethylenepentamine