title: Recombinant vaccinia virus-induced T-cell immunity: quantitation of the response to the virus vector and the foreign epitope.
abstract: Recombinant vaccinia viruses (rVV) have been extensively used as vaccines, but there is little information about the total magnitude of the VV-specific T-cell response and how this compares to the immune response to the foreign gene(s) expressed by the rVV. To address this issue, we quantitated the T-cell responses to both the viral vector and the insert following the infection of mice with VV expressing a cytotoxic T lymphocyte (CTL) epitope (NP118-126) from lymphocytic choriomeningitis virus (LCMV). The LCMV epitope-specific response was quantitated by intracellular cytokine staining after stimulation with the specific peptide. To analyze the total VV-specific response, we developed a simple intracellular cytokine staining assay using VV-infected major histocompatibility complex class I and II matched cells as stimulators. Using this approach, we made the following determinations. (i) VV-NP118 induced potent and long-lasting CD8 and CD4 T-cell responses to the vector; at the peak of the response (approximately 1 week), there were approximately 10(7) VV-specific CD8 T cells (25% of the CD8 T cells) and approximately 10(6) VV-specific CD4 T cells (approximately 5% of the CD4 T cells) in the spleen. These numbers decreased to approximately 5 x 10(5) CD8 T cells (approximately 5% frequency) and approximately 10(5) CD4 T cells (approximately 0.5% frequency), respectively, by day 30 and were then stably maintained at these levels for >300 days. The size of this VV-specific T-cell response was comparable to that of the T-cell response induced following an acute LCMV infection. (ii) VV-specific CD8 and CD4 T cells were capable of producing gamma interferon (IFN-gamma), tumor necrosis factor alpha (TNF-alpha), and interleukin-2; all cells were able to make IFN-gamma, a subset produced both IFN-gamma and TNF-alpha, and another subset produced all three cytokines. (iii) The CD8 T-cell response to the foreign gene (LCMV NP118-126 epitope) was coordinately regulated with the response to the vector during all three phases (expansion, contraction, and memory) of the T-cell response. The total number of CD8 T cells responding to NP118-126 were approximately 20- to 30-fold lower than the number responding to the VV vector (approximately 1% at the peak and 0.2% in memory). This study provides a better understanding of T-cell immunity induced by VV-based vaccines, and in addition, the technique described in the study can be readily extended to other viral vectors to determine the ratio of the T-cell response to the insert versus the vector. This information will be useful in optimizing prime-boost regimens for vaccination.
authors.affiliation: Emory Vaccine Center and Department of Microbiology and Immunology, Emory University School of Medicine, Atlanta, Georgia 30322, USA.
authors.forename: Laurie E
authors.initials: LE
authors.lastname: Harrington
authors.forename: Robbert van der
authors.initials: Rv
authors.lastname: Most Rv
authors.forename: J Lindsay
authors.initials: JL
authors.lastname: Whitton
authors.forename: Rafi
authors.initials: R
authors.lastname: Ahmed
mesh_terms: D000818:Animals; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology; D002452:Cell Count; D016207:Cytokines / Q000032:analysis; D000939:Epitopes / Q000276:immunology; D005822:Genetic Vectors / Q000276:immunology*; D007156:Immunologic Memory; D016169:Lymphocyte Function-Associated Antigen-1 / Q000032:analysis; D008217:Lymphocytic choriomeningitis virus / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D009698:Nucleoproteins / Q000276:immunology; D010446:Peptide Fragments / Q000276:immunology; D011995:Recombination, Genetic; D013154:Spleen / Q000276:immunology; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*; D013997:Time Factors; D014616:Vaccinia virus / Q000235:genetics / Q000276:immunology*; D014764:Viral Proteins / Q000235:genetics / Q000276:immunology
chemical_list: D016207:Cytokines; D000939:Epitopes; D016169:Lymphocyte Function-Associated Antigen-1; D009698:Nucleoproteins; D010446:Peptide Fragments; D014764:Viral Proteins; C094246:nucleoprotein peptide 118-126, lymphocytic choriomeningitis virus