Abstract LB-242: Mammalian orthoreovirus downregulates HIF-1α in hypoxic prostate tumor cells via RACK1-mediated proteasomal degradation and translational inhibition

Abstract

Abstract The purpose of our study was to investigate the oncolytic properties of mammalian orthoreovirus (MRV) on hypoxic prostate tumor cells. Outgrowth from the blood supply and abnormal vascularization leads to poor oxygenation, or hypoxia, within tumors. Hypoxia results in modification of many genes, largely through the activation of the hypoxia inducible factor alpha (HIF-1α) transcription factor. Hypoxia-induced changes lead to tumor propagation through the selection of mutant cells that are capable of clonal expansion and metastases. For these reasons, the extent of hypoxia in tumors correlates with cancer progression, therapy resistance and mortality. MRV is a clinically benign member of the family Reoviridae and is a potent oncolytic agent that replicates in cell lines derived from common tumors. MRV therapy has been tested in numerous clinical trials, and is a safe and effective treatment for many tumor types. We investigated MRV replication in hypoxic prostate tumor cells and found that the virus exhibited robust replication in cells grown in hypoxic conditions. We measured cell cytotoxicity and apoptosis and found that the virus induces cell death by apoptosis in these cells. We additionally examined HIF-1α levels in MRV infected hypoxic prostate tumor cells and found that there are substantially diminished levels of HIF-1α protein in these cells. MRV-induced loss of HIF-1α was reversed when proteasome inhibitors were added at early times to infected hypoxic cells, suggesting that infection results in increased degradation of the HIF-1α protein. However, HIF-1α protein did not accumulate when proteasome inhibitors were added at later times in infection. There were no significant changes in HIF-1α mRNA levels; suggesting MRV also induces downregulation of HIF-1α protein through a second pathway involving virus-induced translational inhibition. siRNA knockdown of the receptor for activated kinase C (RACK1) protein rescued HIF-1α protein expression in MRV infected cells, suggesting that the proteasome-dependent degradation of HIF-1α in MRV infected cells occurs through a RACK1-mediated pathway. Release of RACK1 from cellular stress granule sequestration in infected hypoxic prostate tumor cells was also demonstrated providing a link between virus-induced cellular dysregulation and HIF-1α degradation. Based on these data, we conclude that MRV induces apoptotic cell death and downregulation of the master regulator of hypoxia, HIF-1α, via RACK1-dependent, proteasome-mediated degradation and virus induced translational shutoff in hypoxic prostate tumor cells. This study demonstrates the potential value of inclusion of MRV in clinical treatment modalities to target hypoxic prostate tumor cells at all stages of prostate cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-242. doi:1538-7445.AM2012-LB-242