Neurons are protected from excitotoxic death by p53 antisense oligonucleotides delivered in anionic liposomes.
The potential of anionic liposomes for oligonucleotide delivery was explored because the requirement for a net-positive charge on transfection-competent cationic liposome-DNA complexes is ambiguous. Liposomes composed of phosphatidylglycerol and phosphatidylcholine were monodisperse and encapsulated oligonucleotides with 40-60% efficiency. Ionic strength, bilayer charge density, and oligonucleotide chemistry influenced encapsulation. To demonstrate the biological efficacy of this vector, antisense oligonucleotides to p53 delivered in anionic liposomes were tested in an in vitro model of excitotoxicity. Exposure of hippocampal neurons to glutamate increased p53 protein expression 4-fold and decreased neuronal survival to approximately 35%. Treatment with 1 microm p53 antisense oligonucleotides in anionic liposomes prevented glutamate-induced up-regulation of p53 and increased neuronal survival to approximately 75%. Encapsulated phosphorothioate p53 antisense oligonucleotides were neuroprotective at 5-10-fold lower concentrations than when unencapsulated. Replacing the anionic lipid with phosphatidylserine significantly decreased neuroprotection. p53 antisense oligonucleotides complexed with cationic liposomes were ineffective. Neuroprotection by p53 antisense oligonucleotides in anionic liposomes was comparable with that by glutamate receptor antagonists and a chemical inhibitor of p53. Anionic liposomes were also capable of delivering plasmids and inducing transgene expression in neurons. Anionic liposome-mediated internalization of Cy3-labeled oligonucleotides by neurons and several other cell lines demonstrated the universal applicability of this vector.
Lakkaraju
A
A
Department of Pharmaceutics, University of Minnesota, Minneapolis, Minnesota 55455, USA.
Dubinsky
J M
JM
Low
W C
WC
Rahman
Y E
YE
D000818:Animals; D000838:Anions; D001483:Base Sequence; D052160:Benzothiazoles; D016923:Cell Death / Q000187:drug effects* / Q000502:physiology; D002460:Cell Line; D006224:Cricetinae; D017931:DNA Primers; D004337:Drug Carriers; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D018698:Glutamic Acid / Q000502:physiology*; D006801:Humans; D008081:Liposomes; D009474:Neurons / Q000166:cytology / Q000187:drug effects*; D018696:Neuroprotective Agents / Q000737:chemistry / Q000494:pharmacology*; D016376:Oligonucleotides, Antisense / Q000737:chemistry / Q000494:pharmacology*; D009994:Osmolar Concentration; D051381:Rats; D013844:Thiazoles / Q000494:pharmacology; D014050:Toluene / Q000031:analogs & derivatives* / Q000494:pharmacology; D016159:Tumor Suppressor Protein p53 / Q000235:genetics*
D000838:Anions; D052160:Benzothiazoles; D017931:DNA Primers; D004337:Drug Carriers; D018691:Excitatory Amino Acid Antagonists; D008081:Liposomes; D018696:Neuroprotective Agents; D016376:Oligonucleotides, Antisense; D013844:Thiazoles; D016159:Tumor Suppressor Protein p53; D014050:Toluene; D018698:Glutamic Acid; C121565:pifithrin