Formulation, high throughput in vitro screening and in vivo functional characterization of nanoemulsion-based intranasal vaccine adjuvants.

Vaccine adjuvants have been reported to induce both mucosal and systemic immunity when applied to mucosal surfaces and this dual response appears important for protection against certain pathogens. Despite the potential advantages, however, no mucosal adjuvants are currently approved for human use. Evaluating compounds as mucosal adjuvants is a slow and costly process due to the need for lengthy animal immunogenicity studies. We have constructed a library of 112 intranasal adjuvant candidate formulations consisting of oil-in-water nanoemulsions that contain various cationic and nonionic surfactants. To facilitate adjuvant development we first evaluated this library in a series of high-throughput, in vitro assays for activities associated with innate and adaptive immune activation in vivo. These in vitro assays screened for the ability of the adjuvant to bind to mucin, induce cytotoxicity, facilitate antigen uptake in epithelial and dendritic cells, and activate cellular pathways. We then sought to determine how these parameters related to adjuvant activity in vivo. While the in vitro assays alone were not enough to predict the in vivo adjuvant activity completely, several interesting relationships were found with immune responses in mice. Furthermore, by varying the physicochemical properties of the surfactant components (charge, surfactant polar head size and hydrophobicity) and the surfactant blend ratio of the formulations, the strength and type of the immune response generated (TH1, TH2, TH17) could be modulated. These findings suggest the possibility of using high-throughput screens to aid in the design of custom adjuvants with unique immunological profiles to match specific mucosal vaccine applications.

Intranasal immunization with W 80 5EC adjuvanted recombinant RSV rF-ptn enhances clearance of respiratory syncytial virus in a mouse model.

Respiratory Syncytial Virus (RSV) is a ubiquitous virus that infects almost all people by age two and is a major source of respiratory illness in infants, the elderly and others with compromised immune systems. Currently there is no available vaccine. Prior efforts using formalin-inactivated RSV (FI-RSV) were associated with enhanced respiratory disease upon viral exposure following clinical vaccine trials. Several researchers and pharmaceutical companies have utilized vector-associated live attenuated RSV vaccines in pre-clinical and clinical studies. Another attractive approach, however, is a subunit vaccine which would be easier to produce and quality control. Our group has previously demonstrated in a murine model of infection that intranasal immunization with nanoemulsion-inactivated and adjuvanted RSV induces humoral and cellular immune responses, resulting in protection against RSV infection. The present studies characterize the immune responses elicited by intranasal RSV F protein adjuvanted with nanoemulsion. Intranasal application of nanoemulsion adjuvanted F protein induced a rapid and robust systemic and mucosal antibody response, as well as protection against subsequent RSV challenge. Importantly, RSV challenge in immunized animals did not elicit airway hyper-reactivity, a Th2-skewed immune response or immunopathology associated with hypersensitivity reactions with formalin-inactivated vaccine. These results suggest that RSV F protein adjuvanted with nanoemulsion may be a good mucosal vaccine candidate. Formulating RSV F protein in nanoemulsion creates a well-defined and well-controlled vaccine that can be delivered intranasally to induce T cell mediated immunity without inducing enhanced disease associated with the mouse model of FI-RSV vaccination and infection.

Nanoemulsion nasal adjuvant W805EC induces dendritic cell engulfment of antigen-primed epithelial cells.

Nanoemulsions are adjuvants that enhance antigen penetration in the nasal mucosa, increase cellular uptake of antigens by both epithelial dendritic cells, and promote migration of antigen-loaded dendritic cells to regional lymph nodes within a day of vaccine administration. The objective of this study was to determine whether the W(80)5EC nanoemulsion adjuvant enhances immune response not only by direct uptake of antigen by dendritic cells, but also indirectly, by phagocytosis of antigen-primed, apoptotic, epithelial cells. Consistent with this, we show that exposure of both epithelial cells (TC-1s) and dendritic cells (JAWS II or bone marrow derived dendritic cells (BMDCs)) to nanoemulsion exhibited augmented antigen uptake in cell culture. TC-1 cells subsequently underwent G(2)/M cell cycle arrest and apoptosis, and when co-cultured with JAWS II or BMDCs were rapidly engulfed by the dendritic cells, which responded by up-regulating dendritic cell maturation marker CD86. Altogether these results suggest that the effectiveness of nanoemulsions as adjuvants stems, at least in part, from the engulfment of antigen-loaded epithelial cells, leading to enhanced antigen processing and a strong and balanced mucosal and systemic immune response.