(705c) Influenza Virus-Like Particles as a Platform for Novel Bionanomaterials

Virus-like particles (VLPs) are virus bodies without the genome. VLPs mimic viruses in structure and morphology but are safe because they lack natural viral infectivity. The first commercial use of VLPs was as a recombinant hepatitis B vaccine in 1986. Recently, much attention has been focused on VLPs as a source of novel bionanomaterials. VLPs can be fabricated utilizing their natural ability to self-assemble into nanometer-sized biocomplexes, can be genetically tailored to incorporate new traits and can be targeted to specific cells using the natural infective mechanism of viruses, without the genome to actually cause infection. Many VLPs are constructed based on non-enveloped viruses (i.e., viruses that naturally do not possess an outer lipid bilayer) by self-assembling protein subunits comprising the viral protective protein cage. In this study we have fabricated influenza VLPs, which in contrast contain a lipid envelope and offer the advantage of anchoring membrane bound proteins with different functions for enhanced versatility.

Using recombinant molecular biology, baculovirus expression vectors were first synthesized to express hemagglutinin (HA type H1) and matrix protein (type M1) of the influenza virus A PR/8. Insect cell cultures (SF9 cells) were then used as an expression system for baculovirus vectors to produce VLPs. VLPs were separated from cell culture, concentrated and purified over a discontinuous sucrose gradient to yield 8 mg VLPs from 1L cell culture stock. VLPs were characterized through transmission electron microscopy and were found to be morphologically similar to influenza virus. Purity of VLPs was assessed through SDS-page electrophoresis and directly compared to influenza A PR/8 virus. The cleavage pattern of HA protein incorporated into VLPs suggests that protein translation and post-translational modification in insect cells yielded HA molecule similar to the native HA protein. As an in vivo test, VLPs were injected into mice to study the immune response and subsequent protection from exposure to influenza virus. VLP immunized mice were protected from lethal challenge while naïve non-immunized mice succumbed to infection. This shows that functionally HA protein of VLPs is similar to native HA protein and VLPs can induce antibodies against influenza virus.

In conclusion, we fabricated influenza VLPs that mimic influenza A PR/8 virus, using an insect cell culture system. Influenza VLPs have the potential to encapsulate DNA for combined DNA/protein vaccination and for gene therapy. As in the native virus, hemagglutinin molecule should also enable influenza VLPs to fuse with host cell membrane to deliver encapsulated DNA. Furthermore, influenza VLPs can also be used for imaging and targeted delivery of therapeutics, for example in cancer medications.