title: Unconventional internalization mechanisms underlying functional delivery of antisense oligonucleotides via cationic lipoplexes and polyplexes.
abstract: There is mounting interest in developing antisense and siRNA oligonucleotides into therapeutic entities; however, this potential has been limited by poor access of oligonucleotides to their pharmacological targets within cells. Transfection reagents, such as cationic lipids and polymers, are commonly utilized to improve functional delivery of nucleic acids including oligonucleotides. Cellular entry of large plasmid DNA molecules with the assistance of these polycationic carriers is mediated by some form of endocytosis; however, the mechanism for delivery of small oligonucleotide molecules has not been well established. In this study, splice-shifting oligonucleotides have been formulated into cationic lipoplexes and polyplexes, and their internalization mechanisms have been examined by using pharmacological and genetic inhibitors of endocytosis. The results showed that intercellular distribution of the oligonucleotides to the nucleus governs their pharmacological response. A mechanistic study revealed that oligonucleotides delivered by lipoplexes enter the cells partially by membrane fusion and this mechanism accounts for the functional induction of the target gene. In contrast, polyplexes are internalized by unconventional endocytosis pathways that do not require dynamin or caveolin. These studies may help rationally design novel delivery systems with superior transfection efficiency but lower toxicity.
authors.affiliation: Division of Molecular Pharmaceutics, UNC Eshelman School of Pharmacy, University of North Carolina, Chapel Hill, NC 27599, USA. xming@email.unc.edu
authors.forename: Xin
authors.initials: X
authors.lastname: Ming
authors.forename: Katsuya
authors.initials: K
authors.lastname: Sato
authors.forename: Rudolph L
authors.initials: RL
authors.lastname: Juliano
mesh_terms: D002412:Cations / Q000378:metabolism; D022461:Caveolins / Q000378:metabolism; D002460:Cell Line; D004337:Drug Carriers / Q000378:metabolism*; D004705:Endocytosis / Q000187:drug effects; D006367:HeLa Cells; D006801:Humans; D008055:Lipids / Q000737:chemistry; D016376:Oligonucleotides, Antisense / Q000008:administration & dosage*; D011094:Polyethyleneimine / Q000378:metabolism*; D014162:Transfection
chemical_list: D002412:Cations; D022461:Caveolins; D004337:Drug Carriers; D008055:Lipids; C086724:Lipofectamine; D016376:Oligonucleotides, Antisense; D011094:Polyethyleneimine