Viral Sensitizers Potentiate Infection of Cancer Cells via NFκB
Michael Phan1,2, Ramya Krishnan1,2, Nader El Sayes1,2, Jean-Simon Diallo1,2 1Ottawa Hospital Research Institute; 2University of Ottawa
Genetically engineered oncolytic viruses (OVs) can be effective anti-cancer agents. However, the heterogeneity of tumours and attenuation of OVs can limit their efficacy. Our group has previously shown that small molecules called Viral Sensitizers (VSes) used in combination with OVs can potentiate the infection of cancer cells over 1000-fold. Of particular interest is a novel class of small molecules, including VSe1 and its analogs, which have been previously shown to down-regulate antiviral genes including IFNβ and downstream targets through an unknown mechanism. As a way to identify their molecular target(s), affinity capture experiments revealed that these small molecules interact with glutathione-s-transferase- π (GSTP1-1), a detoxification enzyme induced by oxidative stress. Here, we investigated the effects of VSe1 and analogs on innate cellular antiviral signalling to further refine their mechanism of action and relationship to oxidative stress. The expression of various antiviral cytokines, activation and nuclear translocation of transcription factors involved in the antiviral response were analyzed using immunoblotting, ELISA, and real-time PCR. GST inhibition assays and knockdown experiments were used to assess VSes effects on GST. Our results show that VSe1 analogs inhibit OV-induced nuclear translocation of NFκB and expression of antiviral cytokines such as IFNβ, TNFα, and IL-6. While VSe1 and analogs induce oxidative stress and deplete cellular glutathione, they do not ubiquitously inhibit GSTP1-1 enzymatic activity, and siRNA knockdown of GSTP1-1 does not alter viral sensitivity of cancer cells. However, we demonstrate that inducing oxidative stress with H2O2 enhances OV activity and inhibits NFκB nuclear translocation and transcriptional activity in a similar manner to our VSes. Our results so far demonstrate that VSe1 and analogs inhibit nuclear translocation and transcriptional activity of NFκB in response to OV infection and dampens the expression of antiviral and pro-inflammatory cytokines. Our data suggests that inhibition of NFκB may be a result of redox-sensitive post-translational modifications. Future work will aim to determine whether VSe1 and analogs act through direct binding or modification of redox-sensitive residues of NFκB.
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