Oncolytic Activity of Wild-type Newcastle Disease Virus HK84 Against Hepatocellular Carcinoma Associated with Activation of Type I Interferon Signaling

Abstract

<p><strong>Background and aims: </strong>Hepatocellular carcinoma (HCC) is listed as one of the most common causes of cancer-related death. Oncolytic therapy has become a promising treatment because of novel immunotherapies and gene editing technology, but biosafety concerns remain the biggest limitation for clinical application. We studied the the antitumor activity and biosafety of the wild-type Newcastle disease virus HK84 strain (NDV/HK84) and 10 other NDV strains.</p><p><strong>Methods: </strong>Cell proliferation and apoptosis were determined by cell counting Kit-8 and fluorescein isothiocyanate Annexin V apoptosis assays. Colony formation, wound healing, and a xenograft mouse model were used to evaluate <em>in vivo</em> and <em>in vitro</em> oncolytic effectiveness. The safety of NDV/HK84 was tested in nude mice by an <em>in vivo</em> luciferase imaging system. The replication kinetics of NDV/HK84 in normal tissues and tumors were evaluated by infectious-dose assays in eggs. RNA sequencing analysis was performed to explore NDV/HK84 activity and was validated by quantitative real-time PCR.</p><p><strong>Results: </strong>The cell counting Kit-8 assays of viability found that the oncolytic activity of the NDV strains differed with the multiplicity of infection (MOI). At an MOI of 20, the oncolytic activity of all NDV strains except the DK/JX/21358/08 strain was >80%. The oncolytic activities of the NDV/HK84 and DK/JX/8224/04 strains were >80% at both MOI=20 and MOI=2. Only NDV/HK84 had >80% oncolytic activities at both MOI=20 and MOI=2. We chose NDV/HK84 as the candidate virus to test the oncolytic effect of NDV in HCC in the <em>in vitro</em> and <em>in vivo</em> experiments. NDV/HK84 killed human SK-HEP-1 HCC cells without affecting healthy cells.</p><p><strong>Conclusions: </strong>Intratumor infection with NDV/HK84 strains compared with vehicle controls or positive controls indicated that NDV/HK84 strain specifically inhibited HCC without affecting healthy mice. High-throughput RNA sequencing showed that the oncolytic activity of NDV/HK84 was dependent on the activation of type I interferon signaling.</p>