The genetic approach to the Epstein-Barr virus: from basic virology to gene therapy

Abstract

The Epstein-Barr virus (EBV) infects humans and the genome of this infectious agent has been detected in several tumour types, ranging from lymphomas to carcinomas. The analysis of the functions of the numerous viral proteins encoded by EBV has been impeded by the large size of the viral genome, which renders the construction of viral mutants diYcult. To overcome these limitations, several genetic systems have been developed that allow the modification of the viral genome. Two diVerent approaches, depending on the host cell type in which the viral mutants are generated, have been used in the past. Traditionally, mutants were constructed in EBV infected eukaryotic cells, but more recently, approaches that make use of a recombinant EBV cloned in Escherichia coli have been proposed. The phenotype associated with the inactivation or modification of nearly 20 of the 100 EBV viral genes has been reported in the literature. In most of the reported cases, the EBV latent genes that mediate the ability of EBV to immortalise infected cells were the targets of the genetic analysis, but some virus mutants in which genes involved in DNA lytic replication or infection were disrupted have also been reported. The ability to modify the viral genome also opens the way to the construction of viral strains with medical relevance. A cell line infected by a virus that lacks the EBV packaging sequences can be used as a helper cell line for the encapsidation of EBV based viral vectors. This cell line will allow the evaluation of EBV as a gene transfer system with applications in gene therapy. Finally, genetically modified non-pathogenic strains will provide a basis for the design of an attenuated EBV live vaccine.

Epstein-Barr Virus Downregulates MicroRNA 203 through the Oncoprotein Latent Membrane Protein 1: a Contribution to Increased Tumor Incidence in Epithelial Cells

“…The EBV-infected cell line 293-EBV was previously established by creating a stably transfected cell line (32) that contained the EBV genome, Maxi-EBV (15,16). The EBV-negative cell line, Lm, was derived from 293-EBV cells from a mixture of 20 clones that had lost the EBV genome during subculture (32).…”

Epstein–Barr virus vector-mediated gene transfer into human B cells: potential for antitumor vaccination

“…10 This plasmid termed maxi-EBV is approximately 180 kbp in size and accessible to genetic manipulation in Escherichia coli. 20,21 The GM-CSF-EBV vectors used in this study are based on the maxi-EBV technology. 21 Three major EBV genes (LMP1, EBNA2, and BZLF1) were deleted by homologous recombination to meet the safety aspects for a prospective application in gene therapy (Figure 1a).…”

“…20,21 The GM-CSF-EBV vectors used in this study are based on the maxi-EBV technology. 21 Three major EBV genes (LMP1, EBNA2, and BZLF1) were deleted by homologous recombination to meet the safety aspects for a prospective application in gene therapy (Figure 1a). The latent EBV gene LMP1 is considered to be essential for in vitro growth transformation of B lymphocytes by EBV, 22,23 and is critically involved in the efficiency of this process.…”

EBV LMP2A-specific T Cell Immune Responses Elicited by Dendritic Cells Loaded with LMP2A Protein

“…EBV antigens expressing in all the lymphomas restricted to EBNA1, LMP1, and LMP2 (16)(17)(18)(19)(20). It has been shown LMP2 is the ideal tumor antigen to be chosen as target for EBV associated malignancies in immunotherapy (16,(21)(22)(23)(24)(25). Our previous study has shown that by using purified LMP2A protein loaded Daces, strong EBV-LMP2A specific catatonic T lymphocytes (CTL) responses can be elicited (26).…”