Targeting Liposomal Chemotherapy via Both Tumor Cell–Specific and Tumor Vasculature–Specific Ligands Potentiates Therapeutic Efficacy

Abstract

Neuroblastoma, the most common solid tumor of infancy derived from the sympathetic nervous system, continues to present a formidable clinical challenge. Sterically stabilized immunoliposomes (SIL) have been shown to enhance the selective localization of entrapped drugs to solid tumors, with improvements in therapeutic indices. We showed that SIL loaded with doxorubicin (DXR) and targeted to the disialoganglioside receptor GD 2 [aGD 2 -SIL(DXR)] led to a selective inhibition of the metastatic growth of experimental models of human neuroblastoma. By coupling NGR peptides that target the angiogenic endothelial cell marker aminopeptidase N to the surface of DXR-loaded liposomes [NGR-SL(DXR)], we obtained tumor regression, pronounced destruction of the tumor vasculature, and prolonged survival of orthotopic neuroblastoma xenografts. Here, we showed good liposome stability, long circulation times, and enhanced timedependent tumor accumulation of both the carrier and the drug. Antivascular effects against animal models of lung and ovarian cancer were shown for formulations of NGR-SL(DXR). In the chick embryo chorioallantoic assay, NGR-SL(DXR) substantially reduced the angiogenic potential of various neuroblastoma xenografts, with synergistic inhibition observed for the combination of NGR-SL(DXR) with aGD 2 -SIL(DXR). A significant improvement in antitumor effects was seen in neuroblastoma-bearing animal models when treated with the combined formulations compared with control mice or mice treated with either tumor-or vascular-targeted liposomal formulations, administered separately. The combined treatment resulted in a dramatic inhibition of tumor endothelial cell density. Long-term survivors were obtained only in animals treated with the combined tumor-and vascular-targeted formulations, confirming the pivotal role of combination therapies in treating aggressive metastatic neuroblastoma.

The use of the orthotopic model to validate antivascular therapies for cancer

“…It has been shown that 2aG4 treatment not only has an antivascular action when combined with irradiation in this model, but also enhances the immunogenicity of the tumour leading to immunologic control of residual tumour cells. Moreover, antivascular effects and antitumour effects were accompanied by the recruitment of host immune cells in tumour vasculature and the subsequent infiltration into the tumour interstitium, as shown also by other studies (Beck, 2006;Tozer, 2008).To this regard, we evaluated a successful combination strategy in neuroblastoma model using liposomal formulations of DXR targeted against both tumour cells, via anti-GD 2 monoclonal antibodies, and against the tumour vasculature, via the NGR peptide (Pastorino, 2006). The anti-GD 2 -targeted liposomes resulted in direct cell kill, including cytotoxicity against cells that were at the tumour periphery and were independent of the tumour vasculature, whereas NGR peptide-targeted liposomal doxorubicin bind to and killed angiogenic blood vessels and, indirectly, the tumour cells that these vessels supported, mainly in the tumour core.…”

“…The ligand-based therapies include (1) biological response modifiers or cytokines, such as tumour necrosis factor (TNF) (Corti and Ponzoni, 2004), (2) certain chemotherapeutic drugs, such as vinka alkaloids and arsenic trioxide (Griffin, 2005;Pasquier, 2007), and (3) a variety of strategies that use either antibodies, peptides, or growth factors for binding selectively to tumour vessels (Chaplin, 2006;Pastorino, 2006;Pero, 1999;Siemann, 2004a;Siemann, 2004b;Thorpe, 2004). Moreover, different approaches based on linking antibodies or peptides recognizing tumour-associated Orthotopic tumour models and antivascular therapy 549 vasculature, to toxins, pro-coagulant, and pro-apoptotic effector molecules to induce endothelial cell damage have also been explored (Siemann, 2004b;Thorpe, 2004).…”

“…To this regard, we evaluated a successful combination strategy in neuroblastoma model using liposomal formulations of DXR targeted against both tumour cells, via anti-GD 2 monoclonal antibodies, and against the tumour vasculature, via the NGR peptide (Pastorino, 2006). The anti-GD 2 -targeted liposomes resulted in direct cell kill, including cytotoxicity against cells that were at the tumour periphery and were independent of the tumour vasculature, whereas NGR peptide-targeted liposomal doxorubicin bind to and killed angiogenic blood vessels and, indirectly, the tumour cells that these vessels supported, mainly in the tumour core.…”

“…Constitutively, APN is a membrane-bound metallopeptidase that plays multiple functions as a regulator of various hormones and cytokines, protein degradation, antigen presentation, cell proliferation, cell migration, and angiogenesis (Luan and Xu, 2007;Razak and Newland, 1992;Riemann, 1999). More recently, this technique allows the identification of the sequence CPRECES, specific for another aminopeptidase, the aminopeptidase A (APA), expressed on endothelial and perivascular tumour cells (Marchio, 2004).Various compounds, including cytotoxic drugs, cytokines, antiangiogenic agents, viral particles, fluorescent and contrast tracers, DNA complexes, and other biologic response modifiers have been coupled or synthetically added to NGR peptides in order to increase their vasculature-homing effects (Arap, 1998;Buehler, 2006;Corti and Ponzoni, 2004;Curnis, 2005;Curnis, 2000;Grifman, 2001;Pastorino, 2006;Pastorino, 2003a;Zarovni, 2004).Recently, we showed that liposomes are attractive for targeting drugs and other effectors to tumour vasculature because they can carry large payloads. Using nanoparticles of doxorubicin (DOX) targeted to tumour endothelial cells by coupling the liposomes with peptides containing the NGR sequence, we evaluated their efficacy in orthotopic model of human neuroblastoma (Pastorino, 2003a).…”

PLGA-Based Nanoparticles as Cancer Drug Delivery Systems

“…The aptitude of the nanocarrier to be internalized after binding to target cell is so an significant criterion in the selection of proper targeting ligands (Cho et al, 2008;Danhier et al, 2010). In this strategy, ligand targeted nanocarriers will result in direct cell kill, including cytotoxicity against cells that are at the tumor periphery and are independent on the tumor vasculature (Pastorino et al, 2006;Danhier et al, 2010). The more considered internalization-prone receptors are:…”