Elsevier

Vaccine

Volume 18, Issue 15, 14 February 2000, Pages 1506-1514
Vaccine

Immunization with a DNA expression vector encoding the varicella zoster virus glycoprotein E (gE) gene via intramuscular and subcutaneous routes

https://doi.org/10.1016/S0264-410X(99)00428-4Get rights and content

Abstract

In this study we constructed a plasmid containing the gene encoding varicella–zoster virus transmembrane glycoprotein gE (VZV gE) and evaluated its utility for DNA immunization in mice. Our initial work demonstrates that intramuscular and subcutaneous injection of VZV gE DNA, without the use of costimulatory molecules or other adjuvant materials, results in the generation of antigen-specific antibodies of primarily the IgG2a subclass, indicating that this vaccine can stimulate Th1 type immunity. This is the first report of a prototype DNA vaccine for varicella–zoster virus.

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

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    1

    Present address. Charterhouse Therapeutics Ltd, Winchester, UK.

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