Engineered Nanoparticle Applications for Recombinant Influenza Vaccines

Abstract

Influenza viruses cause seasonal epidemics and represent a pandemic risk. With current vaccine methods struggling to protect populations against emerging strains, there is a demand for a next-generation flu vaccine capable of providing broad protection. Recombinant biotechnology, combined with nanomedicine techniques, could address this demand by increasing immunogenicity and directing immune responses toward conserved antigenic targets on the virus. Various nanoparticle candidates have been tested for use in vaccines, including virus-like particles, protein and carbohydrate nanoconstructs, antigen-carrying lipid particles, and synthetic and inorganic particles modified for antigen presentation. These methods have yielded some promising results, including protection in animal models against antigenically distinct influenza strains, production of antibodies with broad reactivity, and activation of potent T cell responses. Based on the evidence of current research, it is feasible that the next generation of influenza vaccines will combine recombinant antigens with nanoparticle carriers.

Keywords: antigens; influenza; nanoparticles; particles; vaccine.

Fig 1.. Various methods for producing nanoparticles with recombinant antigens.

In solution, recombinant full-sequence hemagglutinin can form radial oligomeric particles termed “rosettes”. Cellular hosts, including plant and insect cells, can be induced to produce virus-like particles (VLPs) through transfection with viral surface proteins. Antigen sequences can be formed into protein subunits which naturally assemble into structures including protein cages, as protein is a type of biopolymer nanoparticle. Synthetic lipids can be used to form liposomes which encapsulate antigens and adjuvants on their surface or interior. Inorganic particles can serve as a core scaffold for the attachment of antigens; in this case, gold can be functionalized with surface antigens using either sulfur association or charge layering methods.

Fig. 2 –

A comparison of soluble antigen binding against nanoparticle- bound antigen binding. Presentation of antigens on a particle results in a high concentration of localized receptor binding on the surface of B cells. Furthermore, while reversible binding limits the exposure time of receptors to soluble antigens, the multivalent binding of particles prolongs the duration of receptor activation to promote a stronger response. This response elicits both intracellular and intercellular signaling, increasing T_FH cell engagement, cytokine production, and production of antibodies, resulting in a superior long-term immunity. Reproduced with permission from.

Fig. 3 –

Ribbon diagram representing the three-dimensional structure of a hemagglutinin monomer and trimer. HA1 indicates the head domain, bearing the sialic acid receptor-binding site. HA2 indicates the stalk domain, possessing the fusion peptide which facilitates cellular entry. Figure modified with permission from.

Fig. 4 –

A structural comparison of influenza virus (left) and VLPs (right). (a) Native virus presents both HA (green) and NA (orange) surface proteins; VLPs may be produced with both proteins or may be produced with HA presented only, as shown. (b) Unlike the virus, VLPs contain no internal proteins or nucleic acids. (c) When viewed with electron microscopy, VLPs bear physical resemblance to the native virus. Reproduced with permission from.

Fig. 5 –

The design of ferritin nanoparticles for influenza antigen presentation. Subunits of ferritin form three-fold axial symmetry, which allows HA trimers to form at the axis. Assembled ferritin particles are octahedral and have an HA trimer valence of 6, which is valuable for the characterization and quantification of HA for influenza vaccines. Reproduced with permission from.

Fig. 6 –

Induction of cross-reactive immunity with multivalent nanoparticles. Utilizing the principles discussed in Fig. 3, it is predicted that multivalent ‘mosaic’ particles will preferentially activate B cells with cross-reactive receptors, thus initiating an adaptive immune response favoring cross-reactive B cell proliferation and antibody production. Reproduced with permission from.

Fig. 7 –

Design schematic for liposomal vaccine nanoparticles. Both the lipid membrane and interior lumen can act as vehicles for antigen and adjuvant transport. This allows B cell activation with membrane-associated surface antigens such as HA, while also facilitating cellular response to conserved internal antigens such as nucleoprotein. Lipid adjuvants, such as monophosphoryl lipid A, can be presented in the liposomal membrane, while cytokines can be delivered to the intracellular compartment. Reproduced with permission from.

Fig. 8 –

Routes of immune stimulation with virosomes. Virosomes can activated B cells directly through the binding of HA to receptors. Virosome HA can also facilitate entry of the particle into antigen presenting cells. Surface antigens are degraded in lysosomes and enter the MHC II pathway for presentation to T helper cells. Internal antigens enter the cytosol through virosomal fusion escape, where they undergo proteasomal degradation and are presented on MHC I to cytotoxic T cells. Reproduced with permission from.

Fig. 9 –

Methods for associating biomolecules to gold-core nanoparticles. (A) Thiol groups associate with gold and can be used to bind molecules, such as the oligonucleotide adjuvant CpG (shown) to the surface of the nanoparticle. (B) Layers of adjuvant and antigen can also be deposited to the surface of the particle through electrostatic interactions. Schematic (A) reproduced with permission from. Schematic (B) reproduced with permission from.