Dr. Julia Felipp
Equine herpesvirus 1 (EHV1) is a ubiquitous and highly contagious pathogen that causes a range of disease severity from mild rhinopneumonitis to abortion, neurologic disease, and death. Notable outbreaks in recent years have emphasized the potential virulence of EHV1, the limitations of current vaccines, and ensuing economic impacts. The broad objective of this research is to identify treatments that will 1) significantly decrease EHV1 viral load, and consequent pathology and shedding during lytic infection; 2) complement antiviral drug efficacy during EHV1 infection; and 3) inhibit EHV1 reactivation from latency. The immediate objective of this project is to evaluate the potential of epigenetic drugs to modulate EHV1 infection because EHV1 gene expression is regulated by epigenetic mechanisms, primarily through histone modifications. Sustaining a repressive epigenetic state through inhibition of histone demethylation has been shown to effectively limit human a-herpesviruses in vitro and in vivo1-3. This outcome was observed both during lytic infection and reactivation from latency. Our hypothesis is that maintaining a repressive epigenetic state on the EHV1 genome will decrease viral load during lytic infection, and prevent reactivation from latency. Thus, we propose the following specific aims using in vitro systems:
Aim 1) to determine how histone demethylase inhibition decreases EHV1 viral load and gene expression during lytic infection;
Aim 2) to determine how histone demethylase inhibition decreases EHV1 reactivation from latently infected cells;
Aim 3) to determine the epigenetic state of the EHV1 genome in the presence or absence of histone demethylase inhibition.
In Aim 1, we will infect equine fetal kidney cells with EHV1 in vitro. Cultures will be treated with 1) a histone demethylase inhibitor, 2) an antiviral drug, 3) both a histone demethylase inhibitor and an antiviral drug, or 4) untreated. EHV1 DNA load and gene expression will be measured by quantitative PCR. Statistical analysis will be performed to identify treatments that inhibit EHV1 during lytic infection.
In Aim 2, we will culture peripheral blood mononuclear cells from horses with latent EHV1 infection and stimulate EHV1 reactivation in vitro. Cultures will be treated with 1) a histone demethylase inhibitor, 2) an antiviral drug, 3) both a histone demethylase inhibitor and an antiviral drug, or 4) untreated. EHV1 DNA load and gene expression will be measured by quantitative PCR. Statistical analysis will be performed to identify treatments that inhibit EHV1 reactivation from latency.
In Aim 3, we will assay the viral epigenetic state upon drug treatment of infected cells by assessing the histone methylation present on the EHV1 genome; we will use chromatin immunoprecipitation of histone methylation followed by quantitative PCR targeting the EHV1 genome.From these aims, we expect to determine whether maintaining a repressive epigenetic state on the EHV1 genome alters viral load and gene expression. Success with the proposed in vitro assays would justify future in vivo studies. Ultimately, we anticipate practical application of this treatment in clinical settings.