Measles remains one of the most important causes of child morbidity and mortality worldwide. Studies of measles in children and in a well-characterized rhesus macaque model have shown that clearance of wild type (WT) measles virus (MeV) RNA is ongoing for many months after clearance of infectious virus with persistence in peripheral blood mononuclear cells and lymphoid tissues. RNA persistence is accompanied by ongoing immune stimulation with continued production of MeV-specific plasmablasts, antibody (Ab) maturation and multiple waves of functionally distinct T cells. These immune responses result in lifelong immunity to measles, but comparable data are not available for the live attenuated MeV vaccine. The MeV vaccine was developed empirically by attenuation of a WT MeV isolate by passage in chicken cells. The resultant live attenuated vaccine was licensed in 1963 and has been remarkably safe and successful although delivery by injection creates hurdles to sustained high coverage that might be alleviated with respiratory delivery. Its safety record, combined with advances in reverse genetics for negative strand viruses, have led to development of recombinant versions as vectors for immunization against other infections and as oncolytic agents for a variety of tumors. However, limited understanding of fundamental aspects of MeV vaccine virus in vivo biology hinders development. For instance, there is little knowledge of where the vaccine virus replicates, the mechanism(s) of attenuation of virulence or how the immune responses induced differ from those induced by WT infection except to note that antibody titers are lower and protection is less durable. We hypothesize that a central difference between infection with vaccine and WT strains is the ability to replicate and persist in lymphoid tissues. This proposal will address this knowledge gap by: 1) Identifying the target cell(s) in which vaccine virus replicates less well than WT virus. We hypothesize that attenuated replication is cell type-specific and that vaccine strains of MeV replicate well in myeloid, endothelial and epithelial cells, but poorly in lymphoid cells. 2) Identifying the viral determinants of inefficient MeV vaccine virus replication. We hypothesize that the hemagglutinin (H) and matrix (M) proteins are the primary determinants of inefficient replication in lymphocytes through effects on TLR2 signaling, virus assembly and release and will test the hypothesis by constructing recombinant strains of EZ vaccine that will tested for replication in lymphocytes. 3) Determining the in vivo sites of vaccine virus replication and dynamics of viral RNA clearance in rhesus macaques. We hypothesize that vaccine strains of MeV do not spread efficiently from lymphoid sites of infection and that both infectious virus and viral RNA are cleared quickly. 4) Identifying differences in the CD4+ T cell and Ab responses to infection with vaccine and WT MeV. We hypothesize that vaccine strains induce fewer MeV-specific Ab-secreting cells for a shorter period of time associated with a less robust and less polyfunctional CD4+ T cell response than WT strains.
Because of inadequate population coverage with the live virus vaccine, measles remains one of the 10 most important causes of death due to infectious diseases. The vaccine is safe, effective and now being modified to produce new vaccines and cancer treatments, but the biological differences from virulent wild type strains of measles virus have never been defined. This project will advance the use of measles vaccine by identifying how the attenuated virus differs from virulent wild type measles virus with respect to infection of myeloid and lymphoid cells and ability to stimulate a protective immune response.
