Neurons are protected from excitotoxic death by p53 antisense oligonucleotides delivered in anionic liposomes

The potential of anionic liposomes for oligonucleotide delivery was explore d because the requirement for a net-positive charge on transfection-compete nt cationic liposome-DNA complexes is ambiguous. Liposomes composed of phos phatidylglycerol and phosphatidylcholine were monodisperse and encapsulated oligonucleotides with 40-60% efficiency. Ionic strength, bilayer charge de nsity, and oligonucleotide chemistry influenced encapsulation. To demonstra te the biological efficacy of this vector, antisense oligonucleotides to p5 3 delivered in anionic liposomes were tested in an in vitro model of excito toxicity. Exposure of hippocampal neurons to glutamate increased p53 protei n expression 4-fold and decreased neuronal survival to similar to 35%. Trea tment with 1 muM p53 antisense oligonucleotides in anionic liposomes preven ted glutamate-induced up-regulation of p53 and increased neuronal survival to similar to 35%. Encapsulated phosphorothioate p53 antisense oligonucleot ides were neuroprotective at 5-10-fold lower concentrations than when unenc apsulated. Replacing the anionic lipid with phosphatidylserine significantl y decreased neuroprotection. p53 antisense oligonucleotides complexed with cationic liposomes were ineffective. Neuroprotection by p53 antisense oligo nucleotides in anionic liposomes was comparable with that by glutamate rece ptor antagonists and a chemical inhibitor of p53. Anionic liposomes were al so capable of delivering plasmids and inducing transgene expression in neur ons. Anionic liposome-mediated internalization of Cy3-labeled oligonucleoti des by neurons and several other cell lines demonstrated the universal appl icability of this vector.