Effects of the Deletion of Early Region 4 (E4) Open Reading Frame 1 (orf1), orf1-2, orf1-3 and orf1-4 on Virus-Host Cell Interaction, Transgene Expression, and Immunogenicity of Replicating Adenovirus HIV Vaccine Vectors

The global health burden engendered by human immunodeficiency virus (HIV)-induced acquired immunodeficiency syndrome (AIDS) is a sobering reminder of the pressing need for a preventative vaccine. In non-human primate models replicating adenovirus (Ad)-HIV/SIV recombinant vaccine vectors have been shown to stimulate potent immune responses culminating in protection against challenge exposures. Nonetheless, an increase in the transgene carrying capacity of these Ad vectors, currently limited to approximately 3000 base pairs, would greatly enhance their utility. Using a replicating, E3-deleted Ad type 5 host range mutant (Ad5 hr) encoding full-length single-chain HIVBaLgp120 linked to the D1 and D2 domains of rhesus macaque CD4 (rhFLSC) we systematically deleted the genes encoding early region 4 open reading frame 1 (E4orf1) through E4orf4. All the Ad-rhFLSC vectors produced similar levels of viral progeny. Cell cycle analysis of infected human and monkey cells revealed no differences in virus-host interaction. The parental and E4-deleted viruses expressed comparable levels of the transgene with kinetics similar to Ad late proteins. Similar levels of cellular immune responses and transgene-specific antibodies were elicited in vaccinated mice. However, differences in recognition of Ad proteins and induced antibody subtypes were observed, suggesting that the E4 gene products might modulate antibody responses by as yet unknown mechanisms. In short, we have improved the transgene carrying capacity by one thousand base pairs while preserving the replicability, levels of transgene expression, and immunogenicity critical to these vaccine vectors. This additional space allows for flexibility in vaccine design that could not be obtained with the current vector and as such should facilitate the goal of improving vaccine efficacy. To the best of our knowledge, this is the first report describing the effects of these E4 deletions on transgene expression and immunogenicity in a replicating Ad vector.

[1]  R. Webby,et al.  Safety and immunogenicity of an oral, replicating adenovirus serotype 4 vector vaccine for H5N1 influenza: a randomised, double-blind, placebo-controlled, phase 1 study , 2013, The Lancet. Infectious diseases.

[2]  T. Shen,et al.  The role of Cdc25A in the regulation of cell proliferation and apoptosis. , 2012, Anti-cancer agents in medicinal chemistry.

[3]  P. Hearing,et al.  Adenovirus Regulates Sumoylation of Mre11-Rad50-Nbs1 Components through a Paralog-Specific Mechanism , 2012, Journal of Virology.

[4]  P. Bruhns.  Properties of mouse and human IgG receptors and their contribution to disease models. , 2012, Blood.

[5]  J. McCullers.  Influenza , 2012, Textbook of Clinical Pediatrics.

[6]  A. Haase,et al.  Replicating Adenovirus-Simian Immunodeficiency Virus (SIV) Vectors Efficiently Prime SIV-Specific Systemic and Mucosal Immune Responses by Targeting Myeloid Dendritic Cells and Persisting in Rectal Macrophages, Regardless of Immunization Route , 2012, Clinical and Vaccine Immunology.

[7]  A. Haase,et al.  Early events in sexual transmission of HIV and SIV and opportunities for interventions. , 2011, Annual review of medicine.

[8]  D. Venzon,et al.  Construction and immunogenicity of replication-competent adenovirus 5 host range mutant recombinants expressing HIV-1 gp160 of SF162 and TV1 strains. , 2010, Vaccine.

[9]  Utpal Banerjee,et al.  Metabolic control of G1–S transition: cyclin E degradation by p53-induced activation of the ubiquitin–proteasome system , 2010, The Journal of cell biology.

[10]  F. Goodrum,et al.  E4orf1 Limits the Oncolytic Potential of the E1B-55K Deletion Mutant Adenovirus , 2009, Journal of Virology.

[11]  J. Heeney,et al.  Systemic neutralizing antibodies induced by long interval mucosally primed systemically boosted immunization correlate with protection from mucosal SHIV challenge. , 2008, Virology.

[12]  M. Robert-Guroff,et al.  Replicating adenovirus vector prime/protein boost strategies for HIV vaccine development , 2008, Expert opinion on biological therapy.

[13]  K. McMasters,et al.  Adenovirus E1B55K Region Is Required To Enhance Cyclin E Expression for Efficient Viral DNA Replication , 2008, Journal of Virology.

[14]  N. Reich,et al.  Adenovirus E4 ORF3 Protein Inhibits the Interferon-Mediated Antiviral Response , 2007, Journal of Virology.

[15]  D. Montefiori,et al.  A Replication-Competent Adenovirus-Human Immunodeficiency Virus (Ad-HIV) tat and Ad-HIV env Priming/Tat and Envelope Protein Boosting Regimen Elicits Enhanced Protective Efficacy against Simian/Human Immunodeficiency Virus SHIV89.6P Challenge in Rhesus Macaques , 2007, Journal of Virology.

[16]  D. Venzon,et al.  Durable protection of rhesus macaques immunized with a replicating adenovirus-SIV multigene prime/protein boost vaccine regimen against a second SIVmac251 rectal challenge: role of SIV-specific CD8+ T cell responses. , 2006, Virology.

[17]  E. Cundari,et al.  E1B55K-Deleted Adenovirus (ONYX-015) Overrides G1/S and G2/M Checkpoints and Causes Mitotic Catastrophe and Endoreduplication in p53-Proficient Normal Cells , 2006, Cell cycle.

[18]  D. Montefiori,et al.  Replicating Rather than Nonreplicating Adenovirus-Human Immunodeficiency Virus Recombinant Vaccines Are Better at Eliciting Potent Cellular Immunity and Priming High-Titer Antibodies , 2005, Journal of Virology.

[19]  B. Clurman,et al.  Cyclin E in normal and neoplastic cell cycles , 2005, Oncogene.

[20]  F. McCormick,et al.  Adenoviral proteins mimic nutrient/growth signals to activate the mTOR pathway for viral replication , 2005, The EMBO journal.

[21]  Sheng Wei,et al.  Small Rho GTPases Regulate Antigen Presentation in Dendritic Cells1 , 2005, The Journal of Immunology.

[22]  R. Shepard,et al.  Diverse Roles for E4orf3 at Late Times of Infection Revealed in an E1B 55-Kilodalton Protein Mutant Background , 2004, Journal of Virology.

[23]  D. Venzon,et al.  Protection against Mucosal Simian Immunodeficiency Virus SIVmac251 Challenge by Using Replicating Adenovirus-SIV Multigene Vaccine Priming and Subunit Boosting , 2004, Journal of Virology.

[24]  Zbigniew Darzynkiewicz,et al.  Analysis of cell cycle by flow cytometry. , 2004, Methods in molecular biology.

[25]  G. Nebl,et al.  Interaction of cofilin with the serine phosphatases PP1 and PP2A in normal and neoplastic human T lymphocytes. , 2003, Advances in enzyme regulation.

[26]  Jin-Hyun Ahn,et al.  Proto-oncogene PML enhances antigen presentation by MHC class I molecules in human lung cancer cells. , 2002, Molecules and cells.

[27]  W. Alvord,et al.  Insertion of HIV-1 genes into Ad4DeltaE3 vector abrogates increased pathogenesis in cotton rats due to E3 deletion. , 2002, Virology.

[28]  Z. Ruzsics,et al.  Subversion of host defense mechanisms by adenoviruses. , 2002, Current topics in microbiology and immunology.

[29]  X. Graña,et al.  E1A modulates phosphorylation of p130 and p107 by differentially regulating the activity of G1/S cyclin/CDK complexes , 2001, Oncogene.

[30]  P. Branton,et al.  Identification of Three Functions of the Adenovirus E4orf6 Protein That Mediate p53 Degradation by the E4orf6-E1B55K Complex , 2001, Journal of Virology.

[31]  M. Reitz,et al.  Expression and Characterization of a Single-Chain Polypeptide Analogue of the Human Immunodeficiency Virus Type 1 gp120-CD4 Receptor Complex , 2000, Journal of Virology.

[32]  M. Lusky,et al.  Regulation of Adenovirus-Mediated Transgene Expression by the Viral E4 Gene Products: Requirement for E4 ORF3 , 1999, Journal of Virology.

[33]  J. Yewdell,et al.  Cutting edge: adenovirus E19 has two mechanisms for affecting class I MHC expression. , 1999, Journal of immunology.

[34]  J. Kaplan,et al.  Novel Role for E4 Region Genes in Protection of Adenovirus Vectors from Lysis by Cytotoxic T Lymphocytes , 1999, Journal of Virology.

[35]  S. Zolla-Pazner,et al.  Vaccine Protection against a Heterologous, Non-Syncytium-Inducing, Primary Human Immunodeficiency Virus , 1998, Journal of Virology.

[36]  D. Levy,et al.  Proto-oncogene PML controls genes devoted to MHC class I antigen presentation , 1998, Nature.

[37]  O. Mühlemann,et al.  Regulation of adenovirus alternative RNA splicing by dephosphorylation of SR proteins , 1998, Nature.

[38]  C. Whiteford,et al.  Wortmannin, a phosphatidylinositol 3-kinase inhibitor, blocks the assembly of peptide-MHC class II complexes. , 1997, International immunology.

[39]  R. Padmanabhan,et al.  Early region 4 modulates adenovirus DNA replication by two genetically separable mechanisms. , 1997, Virology.

[40]  S. Zolla-Pazner,et al.  Long-term protection of chimpanzees against high-dose HIV-1 challenge induced by immunization , 1997, Nature Medicine.

[41]  A. Pavirani,et al.  Humoral immune response to the capsid components of recombinant adenoviruses: Routes of immunization modulate virus‐induced Ig subclass shifts , 1997, European journal of immunology.

[42]  E. Karsenti,et al.  S-phase induction by adenovirus E1A requires activation of cdc25a tyrosine phosphatase. , 1996, Oncogene.

[43]  J. Howe,et al.  Induction of the cell cycle in baby rat kidney cells by adenovirus type 5 E1A in the absence of E1B and a possible influence of p53 , 1993, Journal of virology.

[44]  R. Chanock,et al.  Coexpression of the simian immunodeficiency virus Env and Rev proteins by a recombinant human adenovirus host range mutant , 1992, Journal of virology.

[45]  P. Chanda,et al.  High level expression of the envelope glycoproteins of the human immunodeficiency virus type I in presence of rev gene using helper-independent adenovirus type 7 recombinants. , 1990, Virology.

[46]  P. Hearing,et al.  Adenovirus early region 4 encodes two gene products with redundant effects in lytic infection , 1989, Journal of virology.

[47]  E. Bridge,et al.  Redundant control of adenovirus late gene expression by early region 4 , 1989, Journal of virology.

[48]  Stephen H. Friend,et al.  Association between an oncogene and an anti-oncogene: the adenovirus E1A proteins bind to the retinoblastoma gene product , 1988, Nature.

[49]  J. Snick,et al.  IgG2a restriction of murine antibodies elicited by viral infections , 1987, The Journal of experimental medicine.

[50]  P. Lemay,et al.  Flow cytophotometric and time-lapse cinematographic study of human cells infected by adenovirus type 2 wild-type and two DNA-negative temperature-sensitive mutants. , 1984, The Journal of general virology.

[51]  I. Taylor,et al.  Flow cytometric analysis of DNA content of mouse liver cells following in vivo infection by human adenovirus type 5. , 1981, The Journal of general virology.

[52]  A. Braithwaite,et al.  Alterations to controls of cellular DNA synthesis by adenovirus infection , 1981, Journal of virology.

[53]  N. Jones,et al.  Isolation of adenovirus type 5 host range deletion mutants defective for transformation of rat embryo cells , 1979, Cell.

[54]  J. Lindsay,et al.  Hydroxyurea reversal of inhibition and use as a cell-synchronizing agent. , 1967, The Journal of biological chemistry.