A heterologous DNA prime/protein boost immunization strategy for rhesus cytomegalovirus

Abstract

A previous study in nonhuman primates demonstrated that genetic immunization against the rhesus cytomegalovirus phosphoprotein 65-2 (pp65-2) and glycoprotein B (gB) antigens both stimulated antigen-specific antibodies and CD8 T cell responses, and significantly reduced plasma viral loads following intravenous challenge with RhCMV. It was also noted in this study that weak CD4 T cell and neutralizing antibody responses were generated by DNA alone. To broaden the type of immune responses, a DNA prime/protein boost strategy was used in seronegative macaques, consisting of four DNA immunizations against pp65-2, gB, and immediate-early 1 (IE1), followed by two boosts with formalin-inactivated RhCMV virions. This heterologous prime/boost strategy elicited robust antigen-specific CD4 and CD8 T cell responses in addition to biologically relevant neutralizing antibody titers. Animals were challenged with RhCMV delivered into four sites via a subcutaneous route. Skin biopsies of one of the inoculation sites 7 days post challenge revealed marked differences in the level of RhCMV replication between the vaccinated and control monkeys. Whereas the inoculation site in the controls was noted for a prominent inflammatory response and numerous cytomegalic, antigen-positive (IE1) cells, the inoculation site in the vaccinees was characterized by an absence of inflammation and antigen-positive cells. All five vaccinees developed robust recall responses to viral antigens, and four of them exhibited long-term viral immune responses consistent with effective control of viral expression and replication. These results demonstrate that a heterologous DNA prime/protein boost strategy greatly expands the breadth of antiviral immune responses and greatly reduces the level of viral replication at the primary site of challenge infection.