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Immunotherapeutic herpes simplex virus 2 (HSV-2) vaccine efficacy depends upon the promotion of antigen-specific immune responses that inhibit reactivation or reactivated virus, thus controlling both recurrent lesions and viral shedding. In the present study, a candidate subunit vaccine, GEN-003/MM-2, was evaluated for its ability to induce a broad-spectrum immune response in mice and therapeutic efficacy in HSV-2-infected guinea pigs. GEN-003 is comprised of HSV-2 glycoprotein D2 (gD2ΔTMR340-363) and a truncated form of infected cell polypeptide 4 (ICP4383-766), formulated with Matrix M-2 (MM-2) adjuvant (GEN-003/MM-2). In addition to eliciting humoral immune responses, CD4(+) and CD8(+) T cells characterized by the secretion of multiple cytokines and cytolytic antigen-specific T cell responses that were able to be recalled at least 44 days after the last immunization were induced in immunized mice. Furthermore, vaccination with either GEN-003 or GEN-003/MM-2 led to significant reductions in both the prevalence and severity of lesions in HSV-2-infected guinea pigs compared to those of phosphate-buffered saline (PBS) control-vaccinated animals. While vaccination with MM-2 adjuvant alone decreased recurrent disease symptoms compared to the PBS control group, the difference was not statistically significant. Importantly, the frequency of recurrent viral shedding was considerably reduced in GEN-003/MM-2-vaccinated animals but not in GEN-003- or MM-2-vaccinated animals. These findings suggest a possible role for immunotherapeutic GEN-003/MM-2 vaccination as a viable alternative to chronic antiviral drugs in the treatment and control of genital herpes disease.

A model system to study the pathogenesis of gammaherpesvirus infections is the infection of mice with murine gammaherpesvirus 68 (MHV-68). To define the kinetics of infection, we developed an RNase protection assay to quantitate gene expression from lytic (K3, Rta, M8, DNA polymerase [DNA pol], and gB) and candidate latency (M2, M3, M9, M11, ORF73, and ORF74) genes. All candidate latency genes were expressed during lytic infection of 3T3 cells. Four kinetic classes of transcripts were observed following infection of 3T3 cells: immediate-early (K3, Rta, M8, and ORF73), early (DNA pol), early-late (M3, M11, and ORF74), and late (M2, M9, and gB). To assess the kinetics of viral gene expression in vivo, lungs, spleens, and mediastinal lymph nodes (MLN) were harvested from MHV-68-infected mice. All transcripts were expressed between 3 and 6 days postinfection (dpi) in the lungs. In the spleen, K3, M3, M8, and M9 transcripts were expressed between 10 and 16 dpi when latency is established. The K3, M3, M8, M9, and M11 transcripts were detected in the MLN from 2 through 16 dpi. This is the first demonstration of MHV-68 gene expression in the MLN. Importantly, our data showed that MHV-68 has different kinetics of gene expression at different sites of infection. Furthermore, we demonstrated that K3, a gene recently shown to encode a protein that downregulates major histocompatibility complex class I on the surface of cells, is expressed during latency, which argues for a role of K3 in immune evasion during latent infection.

Respiratory challenge with the murine gammaherpesvirus 68 (gammaHV-68) results in productive infection of the lung, the establishment of latency in B lymphocytes and other cell types, transient splenomegaly, and prolonged clonal expansion of activated CD8(+) CD62L(lo) T cells, particularly a Vbeta4(+) CD8(+) population that is found in mice with different major histocompatibility complex (MHC) haplotypes. Aspects of the CD8(+)-T-cell response are substantially modified in mice that lack B cells, CD4(+) T cells, or the CD40 ligand (CD40L). The B-cell-deficient mice show no increase in Vbeta4(+) CD8(+) T cells. Similar abrogation of the Vbeta4(+) CD8(+) response is seen following antibody-mediated depletion of the CD4(+) subset, through the numbers of CD8(+) CD62L(lo) cells are still significantly elevated. Virus-specific CD4(+)-T-cell frequencies are minimal in the CD40L(-/-) mice, and the Vbeta4(+) CD8(+) population remains unexpanded. Apparently B-cell-CD4(+)-T-cell interactions play a part in the gammaHV-68 induction of both splenomegaly and non-MHC-restricted Vbeta4(+) CD8(+)-T-cell expansion.