CN54gp140(Imperial College London)

Liposomes have shown promise as constituents of adjuvant formulations in vaccines to parasitic and viral diseases. A particular type of liposomal construct, referred to as Army Liposome Formulation (ALF), containing neutral and anionic saturated phospholipids, cholesterol, and monophosphoryl lipid A (MPLA), has been used as an adjuvant for many years. Here we investigated the effects of physical and chemical changes of ALF liposomes on adjuvanted immune responses to CN54 gp140, a recombinant HIV-1 envelope protein. While holding the total amounts of liposomal MPLA and the gp140 antigen constant, different liposome sizes and liposomal MPLA:phospholipid molar ratios, and the effect of adding QS21 to the liposomes were compared for inducing immune responses to the gp140. For liposomes lacking QS21, higher titers of IgG binding antibodies to gp140 were induced by small unilamellar vesicle (SUV) rather than by large multilamellar vesicle (MLV) liposomes, and the highest titers were obtained with SUV having the MPLA:phospholipid ratio of 1:5.6. ALF plus QS21 (ALFQ) liposomes induced the same maximal binding antibody titers regardless of the MPLA:phospholipid ratio. ALF MLV liposomes induced mainly IgG1 and very low IgG2a antibodies, while ALF SUV liposomes induced IgG1≥IgG2a>IgG2b antibodies. Liposomes containing QS21 induced IgG1>IgG2a>IgG2b>IgG3 antibodies. ELISPOT analysis of splenocytes from immunized mice revealed that ALF liposomes induced low levels of IFN-γ, but ALFQ induced high levels. ALF and ALFQ liposomes each induced approximately equivalent high levels of IL-4. Based on antibody subtypes and cytokine secretion, we conclude that ALF liposomes predominantly stimulate Th2, while ALFQ strongly induces both Th1 and Th2 immunity. When CN54 gp140 was adjuvanted with either ALF or ALFQ liposomes, antibodies were induced that neutralized two HIV-1 tier 1 clade C strain pseudoviruses.

It is accepted that an effective prophylactic HIV-1 vaccine is likely to have the greatest impact on viral transmission rates. As previous reports have implicated DNA-priming, protein boost regimens to be efficient activators of humoral responses, we sought to optimize this regimen to further augment vaccine immunogenicity. Here we evaluated single versus concurrent intradermal (i.d.) and intramuscular (i.m.) vaccinations as a DNA-priming strategy for their abilities to elicit humoral and cellular responses against a model HIV-1 vaccine antigen, CN54-gp140. To further augment vaccine-elicited T and B cell responses, we enhanced cellular transfection with electroporation and then boosted the DNA-primed responses with homologous protein delivered subcutaneously (s.c.), intranasally (i.n.), i.m., or transcutaneously (t.c.). In mice, the concurrent priming regimen resulted in significantly elevated gamma interferon T cell responses and high-avidity antigen-specific IgG B cell responses, a hallmark of B cell maturation. Protein boosting of the concurrent DNA strategy further enhanced IgG concentrations but had little impact on T cell reactivity. Interestingly protein boosting by the subcutaneous route increased antibody avidity to a greater extent than protein boosting by either the i.m., i.n., or t.c. route, suggesting that this route may be preferential for driving B cell maturation. Using an alternative and larger animal model, the rabbit, we found the concurrent DNA-priming strategy followed by s.c. protein boosting to again be capable of eliciting high-avidity humoral responses and to also be able to neutralize HIV-1 pseudoviruses from diverse clades (clades A, B, and C). Taken together, we show that concurrent multiple-route DNA vaccinations induce strong cellular immunity, in addition to potent and high-avidity humoral immune responses.IMPORTANCE The route of vaccination has profound effects on prevailing immune responses. Due to the insufficient immunogenicity and protection of current DNA delivery strategies, we evaluated concurrent DNA delivery via simultaneous administration of plasmid DNA by the i.m. and i.d. routes. The rationale behind this study was to provide clear evidence of the utility of concurrent vaccinations for an upcoming human clinical trial. Furthermore, this work will guide future preclinical studies by evaluating the use of model antigens and plasmids for prime-boost strategies. This paper will be of interest not only to virologists and vaccinologists working in the HIV field but also to researchers working in other viral vaccine settings and, critically, to the wider field of vaccine delivery.