Targeted Liposomal Delivery of TLR9 Ligands Activates Spontaneous Antitumor Immunity in an Autochthonous Cancer Model

Abstract

Accessibility of tumors for highly effective local treatment represents a major challenge for anticancer therapy. Immunostimulatory oligodeoxynucleotides (ODN) with CpG motifs are ligands of TLR9, which prime spontaneous antitumor immunity, but are less effective when applied systemically. We therefore developed a liposome-based agent for selective delivery of CpG-ODN into the tumor environment. A peptide that specifically targets angiogenic endothelial cells in a transgenic tumor model for islet cell carcinogenesis was engrafted into CpG-ODN containing liposomes. Intravenous injection of these liposomes resulted in specific accumulation around tumor vessels, increased uptake by tumor-resident macrophages, and retention over time. In contrast, nontargeted liposomes did not localize to the tumor vasculature. Consequently, only vascular targeting of CpG-ODN liposomes provoked a marked inflammatory response at vessel walls with enhanced CD8+ and CD4+ T cell infiltration and, importantly, activation of spontaneous, tumor-specific cytotoxicity. In a therapeutic setting, 40% of tumor-bearing, transgenic mice survived beyond week 45 after systemic administration of vascular-directed CpG-ODN liposomes. In contrast, control mice survived up to 30 wk. Therapeutic efficacy was further improved by increasing the frequency of tumor-specific effector cells through adoptive transfers. NK cells and CD8+ T cells were major effectors which induced tumor cell death and acted in conjunction with antivascular effects. Thus, tumor homing with CpG-ODN-loaded liposomes is as potent as direct injection of free CpG-ODN and has the potential to overcome some major limitations of conventional CpG-ODN monotherapy.

Increased Numbers of Monocyte-Derived Dendritic Cells during Successful Tumor Immunotherapy with Immune-Activating Agents

“…NK cell-derived IFN-g may also contribute to DC and macrophage activation and increase the susceptibility of tumor cells to CTL killing via upregulation of MHC class I expression (50). A similar cooperation between CD8 + T cells and NK cells was reported in other models of local immunotherapy with immune-activating agents (14,18,51).…”

“…In contrast, the TLR4 ligand LPS exhibited low antitumor activity and high toxicity (16). Effective treatments were shown to involve CD8 + T cells, NK cells, or both (15,17,18). Again, the effects of these treatments on DCs have not been examined in detail.…”

Tumor-targeted TNFα stabilizes tumor vessels and enhances active immunotherapy

“…An in vivo CTL assay was used to analyze the capacity of T cells to lyse splenocytes loaded with a tumor-specific peptide (Tag peptide IV; ref. 21). Tumor-bearing controls or TNFα-treated mice do not mount a tumor-specific immune response.…”

Nanoparticle design to induce tumor immunity and challenge the suppressive tumor microenvironment

“…This was shown to result in increased numbers of infiltrating CD4 + and CD8 + T cells into the tumor tissue and the induction of antitumor CTLs [105]. Selectively targeting NPs to the tumor endothelium is possible by targeting tumor endothelial markers such as the vascular endothelial growth factor receptors (VEGF-R) or by using vascular-targeting peptides such as RGR or RGD [106].…”