Reactivation of latent cytomegalovirus (CMV) is frequently observed in recipients of solid organs and bone marrow transplants and is also a significant cause of morbidity and mortality in newborns and in immunocompromised hosts. Currently available anti-viral therapies which target viral replication are associated with narrow therapeutic margins resulting in significant toxicities, including bone marrow suppression, especially in patients with renal dysfunction. Also, the increasing rates of resistance to these agents necessitates the use of efficacious but more toxic agents. It is generally accepted that immediate early (IE) genes are transcriptionally silent in latent infection, and that reactivation of IE gene expression is a critical first step in reactivation of the virus. Treatment with agents which prevent reactivation of IE gene expression would be a novel approach which would avoid problems associated with attempting to combat viral replication. The mechanisms by which reactivation of IE gene expression occurs are not well understood. Because of the lack of animal models to study HCMV infection in vivo, we and others have used MCMV as a model to study CMV latency and reactivation. We have previously utilized a mouse kidney transplant model to induce transcriptional reactivation of MCMV in vivo. Our preliminary studies indicate that genes induced by Toll like receptors are up-regulated in allogeneic kidney transplants. We hypothesize that transcriptional reactivation of MCMV immediate early (IE) gene expression in response to transplantation is initiated by an innate immune response through interaction of Toll-like receptors with cellular ligands damaged by ischemic injury, and this is further amplified by recognition of MHC-incompatible cells. We hypothesize that this leads to increased expression of inflammatory cytokines such as TNF which can themselves activate the enhancer, and that signaling pathways activated by these factors leads to chromatin remodeling and reactivation of IE gene expression. Our preliminary data suggests a major role for TLR2 in activation of the cellular immune response to allogeneic transplants, and thus, for reactivation of MCMV. Through the use of TLR2 deficient mice and the use of siRNA technology to block expression in vivo, we propose to test the requirement for TLR2 as well as signaling adapters MyD88 and Trif in reactivation of MCMV IE gene expression in this model. Treatment of the donor organ prior to transplantation with agents such as siRNA that target steps required for initiation of reactivation would be a novel approach which would significantly reduce the problem of toxicity to the recipient. These studies may therefore lead to the development of new strategies to prevent reactivation of CMV and its sequelae.
Reactivation of latent cytomegalovirus can cause significant morbidity or mortality in immunocompromised transplant patients. Currently available anti-viral therapies, which target viral replication, are associated with severe toxicities and increasing rates of resistance. The goal of this proposal is to understand the molecular pathways that trigger reactivation of latent virus and to develop new strategies to prevent reactivation of latent virus.
