Multicomponent Vaccine Delivery Systems for Subcellular Targeting of Antigen and Molecular Adjuvant.

Abstract

Vaccines are a clinically proven, cost-effective means of improving public health by preventing infectious disease; however, diseases remain that lack efficacious vaccines. The development of a safe and effective multicomponent carrier is a considerable challenge in the field of vaccine delivery systems. Stimulation of cellular immune responses may be particularly desirable in vaccination against cancers or intracellular infections such as viruses. To elicit robust cellular immune responses, protein antigens must be delivered to antigen-presenting cells (APCs) in the context of proper co-stimulation. One approach is to design vaccine delivery systems such that antigens and adjuvant are co-delivered to the same APCs; however, to most effectively stimulate the desired response, the antigen and adjuvant are both delivered to a targeted subcellular location. Our lab has previously demonstrated that co-encapsulating antigen and endosomolytic listeriolysin O (LLO) into pH-sensitive liposomes results in the efficient delivery of antigen to the cytosol of APCs where the antigen can follow the cytosolic pathway of antigen presentation to stimulate cytotoxic T lymphocytes (CTLs). T helper 1 (Th1)-type immune responses may further aid in the removal of cancerous or virally infected cells; therefore, to enhance the CTL response and skew towards the Th1-type, we have incorporated CpG-oligodeoxyribonucleotides (ODNs), which are recognized by Toll-like receptor (TLR9), an endosomal receptor of the innate immune pathway. First, we demonstrate the potential of CpG-ODNs, co-encapsulated in LLO-liposomes, to enhance the CTL response and to skew a Th1-type response utilizing the model antigen ovalbumin. Next, we show that the cellular immune response can be further enhanced by conjugating CpG-ODNs to the liposome surface as a means of promoting interaction between the CpG-ODNs and TLR9, before LLO ruptures the endosomal membrane to release the antigen into the cytosol. We demonstrate that CpG-ODNs in the lipid conjugate form retain immunostimulatory capabilities after incorporation onto liposome bilayers. Lastly we evaluate both methods (co-encapsulation and lipid-conjugation) of incorporating CpG-ODN in LLO-liposomes with the clinically relevant antigen influenza A nucleoprotein. Taken together, this work indicates the utility of LLO-liposomes with CpG-ODNs, either encapsulated or conjugated, as a vaccine delivery system to stimulate a robust cellular immune response.