Anticancer efficiency of reovirus in normoxia and hypoxia.

Oncolytic viruses infect, replicate in, and lyse tumour cells but spare the normal ones. One of oncolytic viruses is a naturally occurring replication-competent reovirus (RV), which preferentially kills tumour cells with activated Ras signaling pathways. The aim of this study was to survey effects of RV on brain tumour-derived cells in vitro under hypoxic conditions since hypoxia causes resistance to radio- and chemotherapy. This study demonstrates that RV replicates preferentially in tumour cells and that the virus is able to overcome cellular adaptation to hypoxia and infect and kill hypoxic tumour cells. RV can both replicate in hypoxic tumour microenvironment and cause the cytopathic effect, subsequently inducing cell death. We found that a large proportion of cells are killed in hypoxia (1% O2) by caspase-independent mechanisms. Furthermore, we learned that the cell death induced by RV in hypoxic conditions is not caused by autophagy.

In vitro and in vivo effects of reovirus on HPV16-transformed mice cells.

UNLABELLED: Oncolytic viruses are examined to serve as anticancer therapeutics. It is expected that in addition to direct oncolytic effect their action will also help eliciting a solid antitumor immunity. In presented series of experiments we have employed two HPV16-transformed mouse (strain C57/B6) cell lines, TC-1 and MK16/III/ABC (MK16), and reovirus type 3, strain Dearing (RV). Both cell lines are highly susceptible to RV and produce large amounts of infectious virus in vitro while normal human are not susceptible to RV. Still, some differences were encountered. TC-1 cells produced moderately lesser amounts of infectious virus, but, paradoxically, were more efficient producers of delta1 antigen of RV and as a consequence of virus infection died more rapidly than simultaneously infected MK16 cells. Minor differences between the cell lines were observed in the percentage of cells arrested in theG2/M phase of the cell cycle and in some markers of apoptosis. When inoculating high doses (5x106) of infected cells (MOI 10 PFU/cell) into syngeneic animals their oncogenic activity was strongly suppressed, nearly completely in the case of MK16 cells and somewhat less efficiently in the case of more oncogenic TC-1 cells. Immunizing experiments in which non-oncogenic doses (106) of RV infected TC-1 cells were tested in parallel with the same doses of irradiated cells brought surprising results. When immunized animals were challenged with TC-1 cells, the irradiated cells proved to be a much better immunogen that the infected cells. However, when challenged with MK16 cells the opposite was true. It is believed that this difference was associated with the different biological properties of the cell lines tested. KEYWORDS: reovirus type 3, HPV16-transformed mouse cell lines, apoptosis, cell cycle, immunization/challenge experiments.

Reovirus - possible therapy of cancer.

Oncolytic viruses infect, replicate in, and eventually lyse tumor cells but spare normal ones. In addition to direct lysis, a result of viral replicative cycle, viruses also mediate tumor cell destruction by inducing nonspecific and specific antitumor immunity. Some viruses express proteins that are cytotoxic to tumor cells. Viruses recognized as oncolytic agents can therefore be divided into three categories: 1/ naturally occurring viruses (e.g. Newcastle disease virus, vesicular stomatitis virus, autonomous parvoviruses, some measles virus strains, reovirus) that selectively replicate in tumor cells, in some instances owing to their relative resistance to interferon action; 2/ virus mutants in which some genes essential for replication in normal cells but evitable in cancer cells have been deleted (e.g.adenovirus ONYX 015 that replicates only in cells with defected p53 or herpes virus G207 which exacts the presence of ribonucleotide reductase); 3/ virus mutants modified by the introduction of tissue-specific transcriptional elements that drive viral genes (e.g.adenovirus CV706 that has PSA restricted expression of E1A and E1B and adenovirus adMycTK that binds selectively on myc protein). Reovirus is prevalent in the human population but not associated with any known human disease. Studies have shown that reovirus multiplicate preferentially in tumor cells with activated gene of ras family or ras-signaling pathway while sparing normal cells. Activated ras or its pathway could be found in as many as 60-80% of human malignancies. In our studies we used cell lines that demonstrably express activated ras. We showed the cytopathic effect of reovirus (serotype 3 strain Dearing) on medulloblastoma cell lines and compared it with its acting on normal human fibroblasts. Oncolytics Biotech Inc. is currently guiding three Phase I or Phase I/II Reolysin studies, and has completed two clinical studies and concluded enrolment in a third one.