Targeted delivery system for antisense oligonucleotides: a novel experimental strategy for neuroblastoma treatment

Abstract

Neuroblastoma (NB) is the most common neuroectoderma derived solid tumour of paediatric age. Since conventional treatments are often inefficient, novel therapeutic interventions are required. Among these, the use of antisense oligonucleotides (asODNs) as therapeutic antineoplastic agents has been recently investigated. Oligonucleotide in vivo applicability is impaired from their high sensitivity to cellular nuclease degradation. Encapsulating them within liposomes could nevertheless increase their stability. C-myb gene expression has been reported in several solid tumours of different embryonic origin, including NB, where it is linked to cell proliferation and/or differentiation. We performed a new technique to encapsulate c-myb antisense oligonucleotides within lipid particles. Liposomes resulting from this technique were called coated cationic liposomes (CCLs), since they were made up of a central core of a cationic phospholipid bound to myb-asODNs, and an outer shell of neutral lipids. A monoclonal antibody (mAb) specific for the neuroectoderma antigen disialoganglioside GD 2, has been covalently coupled to their external surface. The resulting anti-GD 2-targeted CCLs showed high loading efficiency for the asODNs, small particle size and good stability. In vitro, they were able to deliver myb-asODNs selectively to GD 2-positive NB cell lines more efficiently than non-targeted liposomes or free asODNs. Consequently, targeted formulations showed greater inhibition of cell proliferation than non-targeted formulations or free asODNs. Furthermore, we demonstrated that the inhibition of cell proliferation was dependent on the down-modulation of c-myb protein expression. Pharmacokinetic studies showed that these targeted liposomal formulations were long circulating in blood. Biodistribution studies presented differences between the free and the encapsulated myb-as ODN profiles, as well. While free myb-as ODNs are widely distributed (mainly liver, kidney and spleen) even after 30 min post-injection, myb-as ODN entrapped into CCL or anti-GD 2-CCL presents only an accumulation in the spleen after 24 h. Future studies will be performed to evaluate the antitumour efficacy of the above formulations in animal models.