Delivery of foreign antigens by engineered outer membrane vesicle vaccines

Abstract

As new disease threats arise and existing pathogens grow resistant to conventional interventions, attention increasingly focuses on the development of vaccines to induce protective immune responses. Given their admirable safety records, protein subunit vaccines are attractive for widespread immunization, but their disadvantages include poor immunogenicity and expensive manufacture. We show here that engineered Escherichia coli outer membrane vesicles (OMVs) are an easily purified vaccine-delivery system capable of greatly enhancing the immunogenicity of a low-immunogenicity protein antigen without added adjuvants. Using green-fluorescent protein (GFP) as the model subunit antigen, genetic fusion of GFP with the bacterial hemolysin ClyA resulted in a chimeric protein that elicited strong anti-GFP antibody titers in immunized mice, whereas immunization with GFP alone did not elicit such titers. Harnessing the specific secretion of ClyA to OMVs, the ClyA-GFP fusion was found localized in OMVs, resulting in engineered recombinant OMVs. The anti-GFP humoral response in mice immunized with the engineered OMV formulations was indistinguishable from the response to the purified ClyA-GFP fusion protein alone and equal to purified proteins absorbed to aluminum hydroxide, a standard adjuvant. In a major improvement over current practice, engineered OMVs containing ClyA-GFP were easily isolated by ultracentrifugation, effectively eliminating the need for laborious antigen purification from cell-culture expression systems. With the diverse collection of heterologous proteins that can be functionally localized with OMVs when fused with ClyA, this work signals the possibility of OMVs as a robust and tunable technology platform for a new generation of prophylactic and therapeutic vaccines.protein subunit | adjuvant

A Non-classical Assembly Pathway of Escherichia coli Pore-forming Toxin Cytolysin A

“…ClyA toxin is also believed to follow a similar strategy when attacking host cells (1). However, unlike the well-studied ␤-toxins ␣-hemolysin and protective antigen from anthrax toxin, which are secreted by Gram-positive bacteria into the extracellular environment as a soluble monomer (19,20), ClyA is secreted from E. coli via a vesicle-mediated pathway (21)(22)(23). Similar to the budding of yeast cells, the outer membrane of E. coli bubbles out and pinches off to form the outer membrane vesicles (OMVs) (24).…”

Mucosal Immunization with Vibrio cholerae Outer Membrane Vesicles Provides Maternal Protection Mediated by Antilipopolysaccharide Antibodies That Inhibit Bacterial Motility

“…route elicits an antibody response that significantly reduces small-intestinal colonization of suckling neonates challenged orally with V. cholerae (71). In addition, by using OMVs as a delivery vehicle, responses to heterologous antigens have been observed with mice without the need for additional adjuvants (22,70). Therefore, OMVs may represent a versatile vaccine delivery system with natural mucosal adjuvant properties.…”

E. coli Alpha Hemolysin and Properties

“…The anti-GFP humoral response in mice immunized with the engineered OMV formulations was indistinguishable from the response to the purified ClyA-GFP fusion protein alone and was equal to the response to purified proteins adsorbed to aluminum hydroxide, a standard adjuvant. Engineered OMVs containing ClyA-GFP were easily isolated by ultracentrifugation, thus effectively eliminating the need for a laborious antigen purification from cell-culture expression systems (Chena et al, 2010). The retention of hemolytic-protein activity indicated that ClyA-antigen fusions maintained their conformations.…”