Targeted liposomal c-myc antisense oligodeoxynucleotides induce apoptosis and inhibit tumor growth and metastases in human melanoma models.
OBJECTIVE
Melanoma is a highly malignant and increasingly common tumor. Because the cure rate of metastatic melanoma by conventional treatment is very low, new therapeutic approaches are needed. We previously reported that coated cationic liposomes (CCL) targeted with a monoclonal antibody against the disialoganglioside (GD(2)) and containing c-myb antisense oligodeoxynucleotides (asODNs) resulted in a selective inhibition of the proliferation of GD(2)-positive neuroblastoma cells in vitro.


METHODS
Here, we tested the in vivo antitumor effects of this novel antisense liposomal formulation by targeting the c-myc oncogene on melanoma, a neuroectodermal tumor sharing with neuroblastoma the expression of GD(2).


RESULTS
Our methods produced GD(2)-targeted liposomes that stably entrapped 90% of added c-myc asODNs. These liposomes showed a selective binding for GD(2)-positive melanoma cells in vitro. Melanoma cell proliferation was inhibited to a greater extent by GD(2)-targeted liposomes containing c-myc asODNs (aGD(2)-CCL-myc-as) than by nontargeted liposomes or free asODNs. The pharmacokinetic results obtained after i.v. injection of [(3)H]-myc-asODNs, free or encapsulated in nontargeted CCLs or GD(2)-targeted CCLs, showed that free c-myc-asODNs were rapidly cleared, with less than 10% of the injected dose remaining in blood at 30 min after injection. c-myc-asODNs encapsulated within either CCL or aGD(2)-CCL demonstrated a more favorable profile in blood, with about 20% of the injected dose of each preparation remaining in vivo at 24 h after injection. In an in vivo melanoma experimental metastatic model, aGD(2)-CCL-myc-as, at a total dose of only 10 mg of asODN per kilogram, significantly inhibited the development of microscopic metastases in the lung compared with animals treated with myc-asODNs, free or entrapped in nontargeted liposomes, or aGD(2)-CCL encapsulating scrambled asODNs (P < 0.01). Moreover, mice bearing established s.c. human melanoma xenografts treated with aGD(2)-CCL-myc-as exhibited significantly reduced tumor growth and increased survival (P < 0.01 versus control mice). The mechanism for the antitumor effects appears to be down-regulation of the expression of the c-myc protein and interruption of c-myc-mediated signaling: induction of p53 and inhibition of Bcl-2 proteins, leading to extensive tumor cell apoptosis.


CONCLUSIONS
These results suggest that inhibition of c-myc proto-oncogene by GD(2)-targeted antisense therapy could provide an effective approach for the treatment of melanoma in an adjuvant setting.
Pastorino
Fabio
F
Differentiation Therapy Unit, Laboratory of Oncology, G. Gaslini Children's Hospital, 16148 Genoa, Italy.
Brignole
Chiara
C
Marimpietri
Danilo
D
Pagnan
Gabriella
G
Morando
Adriana
A
Ribatti
Domenico
D
Semple
Sean C
SC
Gambini
Claudio
C
Allen
Theresa M
TM
Ponzoni
Mirco
M
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D017209:Apoptosis / Q000187:drug effects*; D004195:Disease Models, Animal; D016503:Drug Delivery Systems; D005260:Female; D005732:Gangliosides / Q000276:immunology; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D008081:Liposomes; D008546:Melanoma, Experimental / Q000235:genetics / Q000473:pathology* / Q000556:secondary*; D051379:Mice; D008807:Mice, Inbred BALB C; D020319:Oligodeoxyribonucleotides, Antisense / Q000008:administration & dosage* / Q000493:pharmacokinetics; D000090063:Proto-Oncogene Mas; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics*; D012878:Skin Neoplasms / Q000235:genetics / Q000473:pathology*; D015996:Survival Rate; D014183:Transplantation, Heterologous; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism
D000911:Antibodies, Monoclonal; D005732:Gangliosides; D008081:Liposomes; C000715470:MAS1 protein, human; D020319:Oligodeoxyribonucleotides, Antisense; D000090063:Proto-Oncogene Mas; D019253:Proto-Oncogene Proteins c-bcl-2; D016271:Proto-Oncogene Proteins c-myc; D016159:Tumor Suppressor Protein p53; C019403:ganglioside, GD2