Non-canonical cell death by reassortant reovirus

Triple-negative breast cancer (TNBC) constitutes 12% of all breast cancer and is associated with worse prognosis compared to other subtypes of breast cancer. Current therapies are limited to cytotoxic chemotherapy, radiation, and surgery, leaving a need for targeted therapeutics to improve outcomes for TNBC patients. Mammalian orthoreovirus (reovirus) is a nonenveloped, segmented, dsRNA virus in the Reoviridae family. Reovirus preferentially kills transformed cells and is in clinical trials to assess its efficacy against several types of cancer. We previously engineered a reassortant reovirus, r2Reovirus, that infects TNBC cells more efficiently and induces cell death with faster kinetics than parental reoviruses. In this study, we sought to understand the mechanisms by which r2Reovirus induces cell death in TNBC cells. We show that r2Reovirus infection of TNBC cells of a mesenchymal-stem like (MSL) lineage downregulates the MAPK/ERK pathway and induces non-conventional cell death that is caspase dependent, but caspase 3-independent. Infection of different MSL lineage TNBC cells with r2Reovirus results in caspase 3-dependent cell death. We map the enhanced oncolytic properties of r2Reovirus in TNBC to epistatic interactions between the Type 3 Dearing M2 gene segment and Type 1 Lang genes. These findings suggest that the genetic composition of the host cell impacts the mechanism of reovirus-induced cell death in TNBC. Together, our data show that understanding host and virus determinants of cell death can identify novel properties and interactions between host and viral gene products that can be exploited for the development of improved viral oncolytics. Importance Triple negative breast cancer (TNBC) is unresponsive to hormone therapies, leaving patients afflicted with this disease with limited treatment options. We previously engineered an oncolytic reovirus (r2Reovirus) with enhanced infective and cytotoxic properties in TNBC cells. However, how r2Reovirus promotes TNBC cell death is not known. In this study, we show that reassortant r2Reovirus can promote non-conventional caspase-dependent but caspase 3-independent cell death and that the mechanism of cell death depends on the genetic composition of the host cell. We also map the enhanced oncolytic properties of r2Reovirus in TNBC to interactions between a Type 3 M2 gene segment and Type 1 genes. Our data show that understanding the interplay between the host cell environment and the genetic composition of oncolytic viruses is crucial for the development of efficacious viral oncolytics.