Baculovirus as a Tool for Gene Delivery and Gene Therapy

Abstract

Based on its ability to express high levels of protein, baculovirus has been widely used for recombinant protein production in insect cells for more than thirty years with continued technical improvements. In addition, baculovirus has been successfully applied for foreign gene delivery into mammalian cells without any viral replication. However, several CpG motifs are present throughout baculoviral DNA and induce an antiviral response in mammalian cells, resulting in the production of pro-inflammatory cytokines and type I interferon through a Toll-like receptor (TLR)-dependent or -independent signaling pathway, and ultimately limiting the efficiency of transgene expression. On the other hand, by taking advantage of this strong adjuvant activity, recombinant baculoviruses encoding neutralization epitopes can elicit protective immunity in mice. Moreover, immunodeficient cells, such as hepatitis C virus (HCV)- or human immunodeficiency virus (HIV)-infected cells, are more susceptible to baculovirus infection than normal cells and are selectively eliminated by the apoptosis-inducible recombinant baculovirus. Here, we summarize the application of baculovirus as a gene expression vector and the mechanism of the host innate immune response induced by baculovirus in mammalian cells. We also discuss the future prospects of baculovirus vectors.

Keywords: baculovirus expression vectors; gene therapy; insect cells; mammalian cells.

Figure 1

Putative model of internalization of baculovirus into mammalian cells. Baculovirus binds to HSPG or cellular receptor(s) present in the lipid raft. This association induces cellular remodeling through signal transduction. In clathrin-mediated endocytosis, baculovirus is internalized into the clathrin-coated pit. In macropinocytosis [37] or phagocytosis [40], filopodia formed by actin dynamics wrap the baculovirus into a macropinosome or phagosome. The viral genome is released from the endosome, macropinosome or phagosome through membrane fusion induced by low pH (Kataoka et al. [37], with modification).

Figure 2

Induction of innate immune response by viral infection. Induction of IFN and pro-inflammatory cytokines is mainly initiated by the recognition of several viral components in mammalian cells. RIG-like receptors such as RIG-I and MDA5 (cytoplasmic sensors) recognize viral-specific double strand RNA, and then the IPS-1-mediated signaling pathway is activated. On the other hand, Toll-like receptors such as TLR9, TLR7 and TLR3 (endosomal sensors) recognize viral DNA, RNA and replicative intermediate double strand RNA, and then a TRIF- or Myd88-mediated signaling pathway is activated. Finally, these pathways trigger the activation of transcription factors such as NF-κB and IRF3 or IRF7, followed by the production of inflammatory cytokines and Type I IFNs, respectively.

Figure 3

Innate immune response against baculovirus infection in mammalian non-immune cells. Baculovirus infection activates the RIG-I/IPS-1/TBK1/IRF3 or cGAS/STING/TBK1/IRF3 signaling pathway and induces Type I IFN production. Baculovirus DNA is recognized by cytoplasmic sensor RIG-I or cGAS [65], followed by activation of the IPS-1- or STING-mediated signaling pathway. The induction of Type I IFN production by IRF3 suppresses transgene expression by baculovirus [60].

Figure 4

Induction of apoptosis in cells replicating HCV RNA by a recombinant baculovirus expressing a proapoptotic protein. (A) Structure of a recombinant baculovirus, rBV-BIM_S, carrying cDNAs of BIMs 2A peptides, and enhanced green fluorescent protein (EGFP) under the control of the elongation factor 1α promoter. The apoptosis pathway was induced by BIMs. (B) HCV replicon cells (HCV-RNA-replicating Huh7 cells) and cured cells (replicon cells from which HCV-RNA was eliminated) were infected with rBV-BIM_S at an MOI of 500, and cell viability was determined at 24 h post infection. rBV-BIM_S can induce apoptosis more efficiently against HCV replicon cells than cured cells (left). HCV replicon cells and cured cells were infected with rBV-BIM_S at an MOI of 500 in the presence or absence of 20 μM qVD-Oph (caspase inhibitor), and cell viability was determined at 24 h post infection. The results show that infection with rBV-BIM_S selectively induced apoptosis in HCV replicon cells through the activation of Bcl-2 family proteins (right).