In Vivo Delivery of Antisense Olignucleotides in pH-Sensitive Liposomes Inhibits Lipopolysaccharide-Induced Production of Tumor Necrosis Factor-#alpha# in Rats

摘要

Kupffer cells play an important role in the pathogenesis of liver diseases. During endotoxemia and alcohol-induced liver dis- ease, tissue injury is preceded by an excessive release of cytokines by these macrophages. Tumor necrosis factor-a (TNF-a) is one of the key cytokines associated with liver injury. Pre-exposure of animals to TNF-a antibodies has been shown to prevent macrophage-mediated liver injury in experimental animals. In this article, we describe a method to encapsulate in pH-sensitive liposomes and to deliver an antisense phospho- rothioate oligonucleotide {T JU-2755) against TNF-a. We de- scribe the efficacy of this formulation in inhibiting endotoxin- mediated production of TNF-a. The liposomes prepared were stable for over 4 weeks at pH 7.4, but readily released their contents when exposed to an acidic environment below pH 6, similar to the pH that exists in early endosomes. Male Sprague- Dawlev rats were administered li.v. lioosome-encaosLllated TJU-2755 (1-2 mg/kg body wt.). Empty liposomes served as controls. Forty-eight hours postinjection, the animals were ad- ministered a single dose of lipopolysaccharide (50 JLg/kg body wt.) and were sacrificed 90 min later. The TNF-a produced by excised liver incubated ex vivo and the levels of plasma TNF-a were determined. After a single administration of liposome- encapsulated antisense T JU-2755, a 30% reduction in TNF-a produced by liver slices was observed. Two daily doses of the antisense oligonucleotide inhibited TNF-a production by 50%. This was associated with a 65 to 70% reduction in plasma levels of TNF-a, compared with controls. These results indicate that oligonucleotide T JU-2755 encapsulated in pH-sensitive liposomes can be used to effectively reduce endotoxin-medi- ated production of TNF-a in macro phages in vivo and thus may be of value in attenuating or preventing macrophage-mediated liver lnjury.