In the mid 1960s, pathologists first observed the formation of nanoscale particles from virus-infected human and animal tissue samples that resembled the virus in morphology but were non-infectious. These viral-like particles (VLPs) are supramolecular assemblies with the same or similar structure as native virions of about 10–200 nm in diameter. They are made up of copies of one or more viral proteins which self-assemble into nanoparticles in an icosahedral or rod-shaped structure. In addition, VLPs are replication deficient because they contain no viral genetic material (DNA or RNA) and thus are not infectious for the vaccinated individuals. VLPs can be experimentally generated in the laboratory using recombinant viral proteins that are expressed in a range of expression systems including prokaryotic cells, yeast, insect cell lines, plants and mammalian cell lines. The use of recombinant methods to produce VLP marked the second generation in VLP production, leading to higher production yields and the ability to produce VLPs derived from viruses of diverse genetic backgrounds.
Fig. 1 An overview of the evolutionary stages in the development of VLPs (Brett D. Hill, et al. 2017)
Different viruses present different structures for generation of VLPs. Based on the presence or absence of lipid envelopes, VLPs are classified into two main types: enveloped and non-enveloped VLPs and the presence of proteins organized into single-layered, two-layered or multi-layered.
Non-enveloped VLPs are typically composed of one or more viral structural proteins, whereas enveloped VLPs (eVLPs) consist of the host cell membrane with viral proteins displayed on the outer surface. Non-enveloped VLPs are easier to produce and purify, but eVLPs are more flexible, as antigens from different pathogens can be integrated. However, they may also contain host’s proteins, which may affect downstream applications.
The simplest available non-enveloped model of VLP consists of single capsid VLP structure like human papillomavirus (HPV) VLP vaccines. These simple VLPs are composed of a single capsid protein that can be expressed in both eukaryotic and prokaryotic systems. In contrast, production of the multi-capsid non-enveloped VLPs are more complex and challenging. These complex VLPs are usually made in eukaryotic expression systems such as yeast, insect cells and plants. Examples of such multi-capsid proteins VLPs that have been successfully assembled into multiple layers and produced in a heterogeneous hosts include VLPs of bluetongue virus, enterovirus 71, infectious bursal disease virus, poliovirus and rotavirus. Self-assembly of enveloped VLPs involves two steps including the formation of an internal protein (nucleocapsid, and/or matrix) and then acquisition of the membrane. The specific constraints to the large-scale manufacture and purification of eVLPs, which are complex assemblies of membranes and viral glycoproteins. Insect cells are usually ideal substrates for correct glycoprotein folding and posttranslational modification. Examples of eVLP production include VLPs of dengue virus, Japanese encephalitis virus, chikungunya virus.
Fig. 2 A schematic diagram of the classification of virus-like particles (D Yan et al. 2015)
The expression system chosen for VLP production must take into consideration the requirements for protein folding and post-translational modifications. The main production platform includes E. coli, yeast, insect cells, mammalian cells and plants. The E. coli and yeast represent easily scaled up and cost-effective production systems. The baculovirus/insect cell (B/IC) expression system process is divided into two phases: an infection phase and a production phase. Baculovirus design is a fast and easy procedure, which makes it suitable for the production of vaccines for viruses whose surface protein can vary between each outbreak. Mammalian cells are typically used to produce complex enveloped VLPs composed of multiple structural proteins. The Chinese Hamster Ovary (CHO) and HEK293 cell lines are most extensively used cell lines for recombinant protein production. Transgenic plants have also been used for VLP production. Agrobacterium tumefaciens is commonly used for infection and transformation of the cells. The most commonly used plants for recombinant protein production are Nicotiana tabacum and Arabidopsis thaliana.
VLPs are capable of inducing strong cellular and humoral responses as direct immunogens. VLP size appears to be favourable for uptake by dendritic cells (DC) via macropinocytosis and endocytosis that play a central role in activating innate and adaptive immune responses. A growing body of data indicates that Gag VLPs that per se contain many immunogenic epitopes are capable of stimulating cellular immune responses via both, the MHC class-I and MHC class-II pathway. Owing to the repetitive particle structure, uptake of a single VLP feeds thousands of contained epitopes into the processing and presentation machinery of APCs, a process thought to be supported by the fusogenic activity and the lipid nature of VLPs.
VLPs show an expanding spectrum of applications such as gene therapy, nanotechnology, vaccination, and diagnostics. In addition, it can also be used for development for various purposes including imaging reagents, template synthesis, and catalysts. Some applications of VLPs against viral diseases as following:
VLPs have the ability to present viral epitopes via authentic, repetitive, highly organized structures similar to those in a native virion but without the risk of infection. These features have attracted significant interest in developing VLPs as novel immunogens in vaccines. VLP can also be applied to chronic disease treatment. VLP display of self-antigens has been used successfully to target molecules that are involved in the pathogenesis of a variety of chronic disease, including Alzheimer’s disease (AD), rheumatoid arthritis and certain cancers.
Fig. 3 VLP-based strategies for vaccine design (Christine L et al. 2007)
VLPs also can be exploited as carriers for the delivery of bio- and nanomaterials, such as drugs, vaccines, quantum dots and imaging substances by virtue of the cavity within their structure. As virus are potent immune activators as well as natural delivery vehicles of genetic materials to their host cells, VLPs are especially well suited for antigen and drug delivery applications.
Lipoparticles are virus-like particles (VLPs) that incorporate high concentrations of target membrane proteins in their native conformation. Lipoparticles are produced in mammalian cells by co-expressing a retroviral structural core polyprotein, Gag, along with a user-specified membrane protein which is correctly translated, folded, and post-translationally modified. Lipoparticles can be used for phage/yeast display, ELISA-based antibody screening and radioligand and fluorescent binding assays, etc.
VLP has biocompatibility and low toxicity. It can be conjugated to fluorescent dyes, probes or contrast agents to obtain better contrast with low toxicity in magnetic resonance imaging and positron emission tomography. This makes them an excellent tool for in vitro and in vivo bioimaging and live cell imaging. In addition, the potential of VLPs to target specific cell types increases the signal-to-noise ratio in diagnostic imaging, thereby providing better resolution. VLPs that display specific antigen proteins can also be used as diagnostic agents.
Cell type-specific ligands, such as proteins, aptamers, and small molecules, can be chemically or genetically coupled to the outer surface of VLPs to achieve cell-specific targeting, which can be further used in medicine and research. The multivalent display of this targeting moiety also contributes to the specific localization of VLPs.
On the basis of their flexibility and stability, simple production and distinctive immunogenic properties, VLPs offer vast opportunities of application in the fields of vaccine development, gene therapy as well as nanobiotechnology. Creative Diagnostics offers a range of high-quality VLP products for use in a wide range of research applications. If you are interested in finding out more about our VLP technology, please feel free to contact us.