Epstein-Barr virus vectors for gene delivery to B lymphocytes.

Abstract

Basic research in Epstein-Barr virus (EBV) molecular genetics has provided means to maintain episomes in human cells, to efficiently deliver episomes with up to 150 kbp of heterologous DNA to human B lymphocytes, and to immortalize human B lymphocytes with EBV recombinants that can maintain up to 120 kbp of heterologous DNA. Episome maintenance requires an EBV nuclear protein, EBNA1, whereas immortalization of cells with EBV recombinants requires EBNA1, EBNA2, EBNA3A, EBNA3C, EB-NALP, and LMP1. EBV-derived vectors are useful for experimental genetic reconstitution in B lymphocytes, a cell type frequently used in establishing repositories of human genetic deficiencies. The ability of EBV-infected cells to establish a balanced state of persistence in normal humans raises the possibility that cells infected with EBV recombinants could be useful for genetic reconstitution, in vivo.The Epstein-Barr virus (EBV) genome has yielded reagents that have been useful in vector design. Genetic engineering of specifically mutated EBV recombinants has also had a major impact on basic EBV research and is useful for experimental genetic reconstitution in human cells. The purpose of this article is to review previous work, indicate potential utilities and liabilities, and describe recent experiments that should enable intensive investigation of an under explored and important part of the EBV genome.EBV is similar in its replication to other herpes viruses and many of the genetic approaches have substantial precedence in alpha herpes virus research. However, EBV also differs from alpha herpes viruses in that it establishes latency in and alters the growth of human B lymphocytes (for reviews, see refs. 1 and 2). These properties derive in large measure from a unique set of genes that encode nuclear and integral membrane proteins that have been given the acronyms, EBNAs (EBV nuclear antigens) and LMPs (latent membrane proteins). In initiation of latent infection in B lymphocytes, EBV first expresses EBNALP and EBNA2. EBNA2 upregulates the EBNA promoter leading to a longer primary transcript from which EBNALP, EBNA2, EBNA3A, EBNA3B, EBNA3C, and EBNA1 transcripts are derived (Fig. 1). EBNA2 also activates the LMP1 and LMP2 promoters. These proteins act in concert to alter B-lymphocyte growth and enable the maintenance of the EBV genome as a multicopy episome in a state of latent infection. Each of these proteins expressed in latently infected cells, with the exception of EBNA1, has epitopes that are presented on the B-cell surface in the context of common class I major histocompatibility complex (MHC) molecules and are recognized by immune human cytotoxic T lymphocytes. This high level of cytotoxic T-cell recognition and the ability of latently infected cells to shift between full latent gene expression with cell proliferation and an EBNA1 only type of latent infection that is immunologically privileged enables latently infected cells to achieve a balanced state of long-term persistence in humans. This state of long-term persis...

Sustained transgene expression in vitro and in vivo using an Epstein–Barr virus replicon vector system combined with HVJ liposomes

“…[5][6][7] From the viewpoint of sustained gene expression, a gene delivery vector based on EBV has been constructed. 8 However, this EBV vector can transfer genes to B lymphocytes but not to other somatic cells. Another approach is the co-introduction of the EBNA-1 and oriP sequences, which are the minimum EBV components for the maintenance of a transgene.…”

Latest Developments in Gene Transfer Technology: Achievements, Perspectives, and Controversies over Therapeutic Applications

“…Preclinical tests have been carried out to characterize the gene delivery properties of vectors based on foami virus [95][96][97], lentiviruses (such as HIV-1 [98][99][100][101][102][103][104] and feline immunodeficiency virus (FIV) [105][106][107]), human cytomegalovirus (CMV) [108], Epstein-Barr virus [109], negative-strand RNA viruses (influenza virus) [110], alphaviruses [111], herpesvirus saimiri [112], hybrid adenoviral/retroviral vector systems [113,114], and hybrid alphavirus/retroviral vector [115]. Other preclinical studies are also assessing the level of vector design improvement that has been reported for a variety of gene transfer models.…”

Viral vectors for gene delivery and gene therapy within the endocrine system

“…Vaccinia viral vectors have also been suggested for use in cancer gene therapy, as they are able to infect a large variety of tumours and have a large cloning capacity (for review, see Peplinski et al 1998). Other vector systems have been based on the Sindbis and Semliki Forest alphaviruses (Polo et al 1999), Sendai virus (Nakanishi et al 1999 and EBV (Robertson et al 1996).…”