Immunization with a DNA expression vector encoding the varicella zoster virus glycoprotein E (gE) gene via intramuscular and subcutaneous routes
Introduction
Varicella–zoster virus (human herpesvirus 3) manifests as chickenpox (varicella) and shingles (zoster), caused by primary and reactivated infection, respectively. Infection with varicella is of particular concern in neonates, adults and immunocompromised patients, while zoster is frequently associated with postherpetic neuralgia. Currently, the only vaccine available is a live attenuated form (Oka strain) which can become latent and cause recurrent infection. As a result, efforts to develop alternative vaccines are underway.
DNA based immunization has provided protection against viral, bacterial and parasitic diseases in experimental systems and offers numerous advantages over conventional protein based preparations (for review see Hasan et al [1]). In this study, we have produced and evaluated a gene encoding VZV glycoprotein E (VZV gE, gpI nucleotides 115789–117694) inserted into a pRc/CMV DNA plasmid vector for immunization. Since the host range of VZV is highly restricted, alternatives for animals models are limited. The lack of a good animal model of human VZV infections has hampered the understanding of the pathogenesis and therapeutic strategies for the disease. However, mice are widely used as model systems to evaluate immediate immune responses in genetic immunization experiments and were used for the purpose of this study. VZV is difficult to grow and historically its proteins and genome organisation were compared to that of herpes simplex virus (HSV). However, with the advancement in molecular biological tools, the complete VZV sequence was published in 1986 by Davison and Scott [2]. The genome is a linear double stranded molecule and approximately 125 kb pairs in size [3]. Approximately half of the genes of VZV have been identified, mostly by analogy with HSV. Sixty-nine unique open reading frames (ORFs) have been identified within the VZV genome [4]. VZV encodes at least six groups of glycoproteins with molecular weights ranging from 20 to 118 kDa, named (for their HSV equivalent) VZV gE, gB, gH, gI, gC and gL [5]. gE is the product of VZV ORF 68 and the primary translation product of 73 kDa is processed to the mature glycosylated form of 88–98 kDa. gE is the most abundant VZV glycoprotein on the surface of virus infected cells. VZV gE induces both neutralizing antibodies and T helper cell responses and contains epitopes recognised by CD4+ and CD8+ cells [6]. Our work has focused on the construction and characterization of a plasmid containing the VZV gE gene and evaluating its antigenicity using nucleic acid immunization technology.
Section snippets
Preparation of DNA
The vector used in this study was the pRc/CMV plasmid vector (Invitrogen, San Diego, CA, USA), in which the BamH1 fragment containing the neomycin cassette was removed (2044 bp deleted, leaving 3482 bp). This vector was also used as the negative control (pRc/CMV). The gE gene was amplified from viral DNA using PCR with BamHI restriction sites encoded on either end. The forward primer (5′-CCTAGGTACCCCTGTCAATT-3′) was designed to produce a BamH1 before the gE start codon. The backward primer
Plasmid characterization
All DNA preparations were checked for purity and molecular weight conformation by agar gel electrophoresis using Bam H1 restriction endonuclease enzyme (data not shown). pRc/CMV and gEpRc/CMV plasmid constructs were transfected into a COS-7 cell line to verify that only the latter expressed authentic gE protein. Pooled zoster serum (PZ7) was used to detect the 93-kDa protein species. The protein was only detected in transfected cells with the gEpRc/CMV DNA plasmid as would be expected and was
Discussion
Immunization with naked DNA has proved to be a valuable means of inducing immunity to a variety of HSV antigens [10], [11]. HSV and VZV share genomic and functional homology, and it is likely that a similar approach will be effective against the latter virus. VZV DNA encodes seven glycoproteins designated gE, gB, gH, gI, gC gL and gK [12], [13], the first six of which are highly immunogenic. However gE is the most abundant of these antigens and appears to be the most immunogenic following
Acknowledgements
This work was funded by the Joint Research Board of St. Bartholomew's Hospital. The author would like to thank Dr. K.J. Williams, Dr. N. Dorrell, Richard Stabler, Lynne Batty and Dr. T. Ng (Protein Phosphorylation Unit, Imperial Cancer Research Fund) for assistance with the molecular biology work; Arif Mustafa and other members of the Biological Service Unit to help with the animal studies, as well as Dr. S. Foynes, Umaima Al-Alem, Brendan Murphy, Dr. R. Zheng, Dr. K. Nye, Chris White, Victoria
References (23)
- et al.
Induction of neutralising antibody and T cell responses to varicella–zoster virus (VZV) using Ty-virus-like particles carrying fragments of glycoprotein E (gE)
Vaccine
(1997) - et al.
Protection against tuberculosis by a plasmid DNA vaccine
Vaccine
(1997) - et al.
Protection against murine cysticercosis using cDNA expression library immunization
Immunol. Lett.
(1998) - et al.
DNA-based vaccination using Schistosoma japonicum (Asian blood-fluke) genes
Vaccine
(1997) - Hasan UA, Abai A, Harper DR, Wren BW, Morrow WJW. Nucleic acid immunization: concepts and techniques associated with...
- et al.
The complete DNA sequence of varicella–zoster virus
J. Gen. Virol.
(1986) Varicella–zoster virus
J. Gen. Virol.
(1991)- et al.
Varicella–zoster virus and its replication
- et al.
New common nomenclature for glycoproteins genes of varicella–zoster virus and there glycosylated products
J. Virol.
(1986) - et al.
Transfection
Antibody responses in recipients of varicella vaccine assayed by immunoblotting
J. Med. Virol.
Cited by (17)
A highly conserved epitope-vaccine candidate against varicella-zoster virus induces neutralizing antibodies in mice
2016, VaccineCitation Excerpt :With the development of biochemistry, molecular vaccinology and genetic engineering technology and increased demand for vaccine safety, the need for more safe and stable vaccines in various forms continues to grow. Plasmids containing the gene encoding the full-length or truncated form of gE (prototype DNA vaccines), which could stimulate immunity in mice, were constructed by Hasan et al. [6,7]. Mullane et al. [8] reported that the heat-treated zoster vaccine was generally safe and immunogenic in immunocompromised adults.
Immune responses of mice against recombinant bovine herpesvirus 5 glycoprotein D
2014, VaccineCitation Excerpt :We speculate that this might be due to the route of vaccination used. Intramuscular inoculations are performed in poorly vascularized tissues [37], and might induce a local response as a result of antigen presentation and accumulation at the draining lymph nodes [38]; however, the cytokine assay was performed in cells obtained from spleens, and we did not examined the cytokine transcripts in draining lymph nodes at this time. The higher levels of IFN-γ in spleen cells stimulated in vitro with whole live BoHV-5 after vaccination with rgD5 suggests that it was able to prime those cells against the virus, which was also observed in animals vaccinated with rgD5 + Em-D, who had higher transcript levels of GM-CSF and IL-17 when spleen cells were stimulated with BoHV-5.
Cell-mediated immune responses to a varicella-zoster virus glycoprotein E vaccine using both a TLR agonist and QS21 in mice
2012, VaccineCitation Excerpt :An MPL-adjuvanted subunit vaccine based on truncated gE has been clinically shown to boost pre-existing anti-gE humoral responses [37]. In preclinical models, recombinant protein and DNA vaccines including gE as an antigen have demonstrated high immunogenicity and/or were able to induce partial efficacy against infection [38–44]. Since VZV does not efficiently replicate in animal models [45], it is not possible to assess the efficacy of a gE candidate vaccine against VZV pathogenesis, latency and reactivation in the preclinical setting.
DNA vaccine strategies: Candidates for immune modulation and immunization regimens
2003, MethodsCitation Excerpt :A study of various forms of hepatitis B virus surface antigens showed that, while all forms elicited comparable levels of CTL, the secreted forms elicited higher antibody titers and a higher IgG1/IgG2a ratio [28]. In addition, a study by Hasan et al. [29] showed that immunization of mice with a plasmid expressing a truncated form of the Varicella zoster virus gE antigen tended to promote a Th2-type response, whereas a construct expressing a non-truncated form induced a Th1 response. A naturally secreted protein can be modified to enhance the rate of secretion by replacement of its signal sequence with that of a highly expressed protein, such as tissue-type plasminogen activator (t-PA) [30–33].
Intradermal immunization with novel plasmid DNA-coated nanoparticles via a needle-free injection device
2003, Journal of Biotechnology
- 1
Present address. Charterhouse Therapeutics Ltd, Winchester, UK.