Virus-like particles show promise as candidates for new vaccine strategies

Abstract

Future VirologyVol. 5, No. 4 EditorialFree AccessVirus-like particles show promise as candidates for new vaccine strategiesMaurizio FedericoMaurizio FedericoNational AIDS Center, Division of Pathogenesis of Retroviruses, Istituto Superiore di Sanità, Viale Regina Elena, 299, 00161 Rome, Italy. Search for more papers by this authorEmail the corresponding author at maurizio.federico@iss.itPublished Online:22 Jul 2010https://doi.org/10.2217/fvl.10.29AboutSectionsPDF/EPUB ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareShare onFacebookTwitterLinkedInRedditEmail Virus-like particles (VLPs) are genomeless virions produced by the assembly of one or more viral structural proteins [1]. They can be generated by engineering the genomes of many virus species, including human papillomavirus (HPV) [2,3], HBV and HCV [4,5], and retro- [6] and lenti-viruses [7]. VLPs can be produced from different mammalian cell lines, various species of yeast (i.e., Saccharomyces cerevisiae and Pichia pastoris) [8,9]Escherichia coli[10] and other bacterial systems, baculovirus/insect cell systems [11,12], and plants [13]. Ease of expression, ability to scale-up and cost of production have made yeast the most popular VLP expression system. However, considerations such as appropriate protein glycosylation, correct folding and assembly favored alternative methods of production. Among these, mammalian cells produce appropriate modifications and authentic assembly, but are a less controllable and more expensive expression system. The ability of VLPs to include heterologous nucleic acids and small molecules has made them novel vessels for gene and drug delivery. For instance, HPV VLPs have been demonstrated to mediate functional delivery of plasmid DNA in vivo[14], MS2 bacteriophage VLPs have been used to deliver antisense oligodeoxynucleotides [15] and VLPs have also been used as scaffolds in nanoparticle biotechnology [16].Virus-like particles mimic the overall structure of virus particles, while having the ability to bind and enter cells using appropriate receptors. They are also potent immunogens owing to their ability to combine key immunogenic properties of viruses into a single entity. In view of their particulate nature and dimensions, VLPs are efficiently taken up by dendritic cells and transported to lymph nodes. In the case of production of VLPs for immunization studies, the choice of expression system might significantly influence direction and outcome of the immune response. In any case, VLPs possess excellent adjuvant properties, supporting both cellular and humoral immune responses [1,17]. Compared with recombinant antigens, which, in many cases, require multiple inoculations in the presence of adjuvants to elicit a valuable immune response, VLPs can induce a strong immunity upon single inoculation [1]. Commercialized VLP-based vaccines have been successful in protecting humans from HBV [18] and HPV infections [19], and their potential to challenge other infectious diseases and cancer are currently being explored. Concerning retro- and lenti-virus-based VLPs, several attempts have been made to induce an effective anti-HIV immune response [7,20]. Retroviral VLPs have also been successfully generated through the inoculation of Gag-Pol-expressing vectors in mice [21].Of note, VLPs can be also engineered to incorporate a variety of peptides and proteins, and, at least in the case of retro- and lenti-viral VLPs, of viral envelope glycoproteins and cell receptors (chimeric VLPs), thus, representing an extraordinarily flexible tool for new vaccine strategies. For example, an antigen delivery system based on recombinant parvovirus-like particles has been developed by exploiting the self-assembly capacity of the VP2 capsid protein of porcine or canine parvoviruses expressed in insect cells through the baculovirus expression system. Of note, porcine parvovirus VLPs containing a CD8 epitope from the lymphocoriomeningitis virus nucleoprotein were found to evoke a potent cytotoxic T lymphocyte (CTL) immune response, which was able to protect mice against the lethal virus infection [22].Unfortunately, in most instances the antigen incorporation into the VLP structural proteins causes major problems in terms of correct assembly of the viral particle. This shortcoming has been bypassed in the case of murine polyoma virus by tagging the heterologous protein to be incorporated in the VLPs with a polyproline motif able to interact with the PPLP-binding domain of the VP1 capsid component [23]. Alternatively, heterologous proteins can be incorporated in polyoma VLPs upon fusion with the minor coat proteins VP2/VP3 by exploiting the hydrophobic interaction between these virion components and the major capsid protein VP1 [24]. Although successful, these strategies led to the incorporation of only a few molecules for each VLP. As another example, HPV-based VLPs have been engineered to carry heterologous peptides as products of fusion with L1 or -2. However, L1 supports a maximum of 60 extra amino acids [25], whereas L2-based fusion proteins incorporate in VLPs at a ratio of approximately 1:30 with respect to the L1 major capsid component [26].Among retro- and lenti-virus-derived VLPs, much insight into immune stimulation and optimum VLP design has been gained from HIV-1 Pr55gag-derived VLPs [12,27–29], whose immunogenicity can be broadened by making chimeric Gag molecules [30]. However, VLPs incorporating foreign polypeptides fused in frame with Gag assemble with a strongly diminishing efficiency with both the insertion domains [31] and length of exogenous sequences [32]. In an alternative design, baculovirus-derived HIV-1 VLPs engineered to display clade A HIV-1 Env gp120 molecules on their surface have been proven to efficiently induce anti-Env neutralizing antibodies in mice [33].While in many cases the spread of infecting viruses can be counteracted by virus-specific antibodies through the so-called ‘neutralization’ phenomenon [34,35], in other instances this is not sufficient, and the control and clearance of viral infections would benefit from the induction of alternative immune mechanisms. In this regard, specific CTLs can be crucial in light of their ability to recognize and kill virus-infected cells. While different VLP types have been proven suitable to efficiently accommodate peptides, major obstacles remain for the insertion of large polypeptides and proteins able to specifically elicit unrestricted CTL immune responses. This could also be of major utility for virus-induced and virus-independent cancers, where the induction of CTLs against tumor-associated antigens can be of relevance for the clearance of tumor cells. In fact, an important rationale for the development of cancer vaccines originates from the observation that the priming phase of the antitumor immune response in cancer patients is defective or compromised. The induction of tumor antigen-specific effector T cells elicited by vaccination would, at least in part, fill this gap [36].Within the last few decades, several vaccine strategies have proven suitable for the induction of both unrestricted and strong CTL immune responses. Among these, the most studied immunogens are attenuated viruses [37], recombinant viral vectors [38–41] and DNA vaccines [36,42–44]. However, serious safety concerns, mainly regarding the delivery of exogenous genomic material, limit their usefulness in humans.In an effort to circumvent these limitations, we developed a vaccine platform technology where foreign antigens are efficiently incorporated into lentiviral VLPs. This relies on the ability of an HIV-1 Nef mutant (Nef7) to be incorporated into HIV and retroviral virions approximately 100-fold more than the wild-type protein, while acting as a carrier molecule upon C-terminal fusion [45] with up to 630-amino-acid-long heterologous proteins [Federico M, Unpublished Data].Our approach combines promising VLP nanobiotechnology with the unique advantages of the Nef7 mutant. In addition, pseudotyping the VLPs with envelope glycoproteins supporting pH-dependent cell entry (e.g., from vesicular stomatitis or Rabies viruses) improves the overall efficiency of delivery into antigen-presenting cells, while favoring the CD8+ T-cell immune response through cross-presentation and cross-priming mechanisms. This is expected to be a consequence of the VLP cell entry through endocytosis, fusion in endosomes and delivery of VLP products in the cytoplasm. Here, they can interact with the proteasome, thereby undergoing degradation and association with class I MHC molecules [46]. On this subject, we recently obtained the proof-of-principle that (VSV-G) Nef7-based VLPs allow efficient cross-presentation of the incorporated foreign antigen (i.e., the E7 oncoprotein from HPV), while eliciting a CTL-based immune response strong enough to block the growth of HPV-related tumors in mice [47]. The potential advantages of the Nef7-VLP system over recombinant viral systems or other chimeric VLPs can be summarized as follows: lack of genetic material transfer, high incorporation efficiency of the foreign antigen in the presence of only four additional protein products, flexibility in the envelope protein incorporation and possibility to incorporate the Nef7-based fusion products also in retroviral particles.Lentivirus-based VLPs are typically produced through multiplasmid transient transfection in eukaryotic cells [48]. At present, a major obstacle for the development of vaccine platforms based on lentiviral VLPs is represented by the technical difficulties in their large-scale production. However, significant advances have been made in this field [49], and one would expect that well reproducible and cost-effective methods suitable for fast mass production of lentiviral VLPs will be available in a very short time.The development of new vaccine strategies based on VLPs is certainly a rapidly evolving field. Considering their flexibility, VLPs offer great opportunities of application in vaccinology. Biotechnological industries should pay attention to the development of streamlined processes of production and purification of complex VLPs, which, in the near future, could represent new classes of vaccines possessing unique immunogenic characteristics.AcknowledgementsThe author is indebted to I Ronci (National AIDS Center, ISS, Rome, Italy) for her excellent editorial assistance.Financial & competing interests disclosureThe author has no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.Bibliography1 Grgacic EV, Anderson DA: Virus-like particles: passport to immune recognition. 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