Peptide-engrafted liposomes : effective delivery vehicles for antigen and DNA to modulate immunity

Abstract

Since dendritic cells (DC) possess the unique ability to prime na{u00EF}ve T cells and orchestrate subsequent antigen (Ag)-specific effector functions, a new generation of cancer therapies is emerging, which seek to restore host defence by triggering DCs to generate anti-tumour immunity. Currently, a wide range of nanoparticle-based vaccines are being developed to harness the immune system to recognize and eradicate cancer cells. However, the search for an effective, versatile and 'easy to scale-up' delivery system to deliver antigenic payloads to target cells still remains elusive. Liposomes have long been used as carriers for delivering Ag and nucleic acids to DC and to other antigen presenting cells (APC) for induction of both humoral and cellular immunity. Unfortunately, the lack of suitable targeting strategy to selectively direct the liposomal payload to APC has limited their extensive use in clinical settings. Interestingly, incorporation of the chelator lipid 3(nitrilotriacetic acid)-ditetradecylamine (NTA{u2083}-DTDA) into liposomes and membrane vesicles enables engraftment of histidine-tagged forms of targeting proteins/peptides by metal chelating linkage. This thesis aims to explore the use of short synthetic peptides bearing polyhistidine tags as targeting molecules on Ag/DNA-bearing liposomes to facilitate delivery of the payload to APCs in vivo, in an attempt to induce Ag-specific immunity, and to address the drawbacks associated with current liposomal delivery systems. To take advantage of the receptor targeting for selective delivery of Ag to cells expressing CD11c/CD18 (i.e., DC), we initially explored the potential of CD11c/CD18-interacting peptides (pCD11c) to target NTA{u2083}-DTDA-containing liposomes to DCs. Similarly, peptides containing sequences related to regions of High-mobility group box-1 protein (pHMGB1) and of flagellin (pFlg) were also assessed for their ability to promote liposomal interaction with APCs. Two novel peptides that interact with TLR5 (Toll-like receptor-5) have been identified. The results indicate that the engraftment of pCD11c, pHMGB1 and pFlg peptides onto liposomes promotes binding of the liposomes to DCs and facilitates delivery of liposomal Ag to DCs; thereby eliciting Ag-specific and anti-tumour immunity. To examine the versatility of the approach, the delivery of plasmid DNA to APC through pFlg-engrafted liposome also was explored. The results demonstrate that DODAP-containing lipoplexes engrafted with pFlg peptide bind to DCs and can mediate transgene expression in vivo. Moreover, the vaccination of mice with such lipoplexes also was shown to induce potent Ag-specific and anti-tumour immunity in the B16-OVA tumour model. To summarize, the research performed for this thesis has identified a number of peptides that when produced with a His-tag and engrafted onto NTA{u2083}-DTDA containing liposomes and membrane vesicles have potential for eliciting Ag-specific responses and hence for vaccine development. Because the peptides also exert adjuvant effects, the liposomal delivery system developed enables convenient delivery of Ag/DNA and DC maturation stimuli in a single delivery unit; thereby, simplifying the vaccination approach. More importantly, the demonstration that tumour-derived membrane vesicles engrafted with these His-tagged targeting peptides can elicit potent Ag-specific and anti-tumour immunity in murine tumour models reflects the potential of the targeting strategies developed to be useful for clinical applications.