Inflammasome-activating nanoparticles as modular systems for optimizing vaccine efficacy,”

a b s t r a c t Innate immune system activation is a critical step in the initiation of an effective adaptive immune response; therefore, activation of a class of innate pathogen receptors called pattern recognition receptors (PRR) is a central feature of many adjuvant systems. It has recently been shown that one member of an intracellular PRR, the NLRP3 inflammasome, is activated by a number of classical adjuvants including alu- Inflammasome activation in vitro requires signaling of both the Toll-like receptor (TLR) and NLRP3 in antigen-presenting cells. Here we present a class of nanomaterials endowed with these two signals for rapid optimization of vaccine design. We constructed this system using a simple approach that incorporates lipopolysaccharides (LPS) onto the surface of nanoparticles constructed from a biocompatible polyester, poly(lactic-co-glycolic acid) (PLGA), loaded with antigen. We demonstrate that LPS-modified particles are preferentially internalized by dendritic cells compared to uncoated nanoparticles and the system, when administered to mice, elicits potent humoral and cellular immunity against a model antigen, ovalbumin. Wild-type macrophages pulsed with LPS-modified nanoparticles resulted in production of the proinflammatory cytokine IL-1␤ consistent with inflammasome activation. In comparison, NLRP3-deficient and caspase-1-deficient macrophages showed negligible production of IL-1␤. Furthermore, when endocytosis and lysosomal destabilization were inhibited, inflammasome activity was diminished, supporting the notion that nanoparticles rupture lysosomal compartments and behave as 'danger signals' [Hornung V, Bauernfeind F, Halle A, Samstad EO, Kono H, Rock KL, et al. Silica crystals and aluminum salts activate the NALP3 inflammasome through phagosomal destabilization. Nat Immunol 2008;9(August (8)):847-56]. The generality of this vaccination approach is tested by encapsulation of a recombinant West Nile envelope protein and demonstrated by protection against a murine model of West Nile encephalitis. The design of such an antigen delivery mechanism with the ability to stimulate two potent innate immune pathways represents a potent new approach to simultaneous antigen and adjuvant delivery

A West Nile virus recombinant protein vaccine that coactivates innate and adaptive immunity.

A chimeric protein West Nile virus (WNV) vaccine capable of delivering both innate and adaptive immune signals was designed by fusing a modified version of bacterial flagellin (STF2 Delta ) to the EIII domain of the WNV envelope protein. This fusion protein stimulated interleukin-8 production in a Toll-like receptor (TLR)-5-dependent fashion, confirming appropriate in vitro TLR5 bioactivity, and also retained critical WNV-E-specific conformation-dependent neutralizing epitopes as measured by enzyme-linked immunosorbent assay. When administered without adjuvant to C3H/HeN mice, the fusion protein elicited a strong WNV-E-specific immunoglobulin G antibody response that neutralized viral infectivity and conferred protection against a lethal WNV challenge. This potent EIII-specific immune response requires a direct linkage of EIII to STF2 Delta , given that a simple mixture of the 2 components failed to induce an antibody response or to provide protection against virus challenge. The presence of a functional TLR5 gene in vivo is also required--TLR5-deficient mice elicited only a minimal antigen-specific response. These results confirm that vaccines designed to coordinately regulate the innate and adaptive immune responses can induce protective immune responses without the need for potentially toxic adjuvants. They also support the further development of an effective WNV vaccine and novel monovalent and multivalent vaccines for related flaviviruses

Abrogation of macrophage migration inhibitory factor decreases West Nile virus lethality by limiting viral neuroinvasion

The flavivirus West Nile virus (WNV) is an emerging pathogen that causes life-threatening encephalitis in susceptible individuals. We investigated the role of the proinflammatory cytokine macrophage migration inhibitory factor (MIF), which is an upstream mediator of innate immunity, in WNV immunopathogenesis. We found that patients suffering from acute WNV infection presented with increased MIF levels in plasma and in cerebrospinal fluid. MIF expression also was induced in WNV-infected mice. Remarkably, abrogation of MIF action by 3 distinct approaches (antibody blockade, small molecule pharmacologic inhibition, and genetic deletion) rendered mice more resistant to WNV lethality. Mif–/– mice showed a reduced viral load and inflammatory response in the brain when compared with wild-type mice. Our results also indicate that MIF favors viral neuroinvasion by compromising the integrity of the blood-brain barrier. In conclusion, the data obtained from this study provide direct evidence for the involvement of MIF in viral pathogenesis and suggest that pharmacotherapeutic approaches targeting MIF may hold promise for the treatment of WNV encephalitis