Development of a Novel in-Vitro Hcov-Oc43 Screening Method

ABSTRACT

Aim: To develop a simple, inexpensive antiviral screening assay, applicable to SARS-CoV-2, using a plate-based bioassay approach to assess the in-vitro activity of compounds against HCoV-OC43. Background(s) Despite the successful deployment of vaccines against SARS-CoV-2 there remains a need for effective antivirals for acute infection treatment. A distinct problem facing the search for new anti-coronavirus compounds is the cost of antiviral screening, compounded by the biosecurity concerns of live SARSCoV- 2 culture. In concert with low pathogenic surrogate virus use, the resazurin reduction assay, which is often employed for compound cytotoxicity assessments can be employed for safe, rapid and inexpensive antiviral screening. Method(s) In-vitro cell based resazurin reduction assays were optimised using remdesivir as a control compound for the assessment of anti-HCoV-OC43 activity. Following optimisation, 246 purified natural compounds from the University of Western Australia's compound collection,were screened using the resazurin bioassay as a primary screen, under pre-treatment and cotreatment conditions. Five compounds, which demonstrated anti- HCoV-OC43 activity, were chosen for secondary screening with dose responses determined using qRT-PCR. Result(s) Primary screens of the 246 compounds using the resazurin bioassay identified five compounds with a relative viral inhibition >60% and a relative cell viability >70% (Table 1). The Z factor of the pre-treatment and co-treatment assays was >0.5 (average +/- SD;0.85 +/- 0.07, 0.91 +/- 0.03 respectively). Further dose response analysis of the top five compounds identified one compound with an IC50 value <10 muM. Conclusion(s) The method developed is an appropriate primary screening tool for the identification of novel compounds with anti-HCoV-OC43 activity.Copyright © 2023 Southern Society for Clinical Investigation.