Developing a high throughput screening assay for the discovery of SARS-CoV-2 entry inhibitors

Abstract

Despite the development and global distribution of vaccines, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19, continues to pose a public health challenge. Infections continue to persist today due to rapid mutation in the spike (S) protein, the target for vaccine-induced neutralizing antibodies. These mutations result in reduced vaccine efficacy and subsequent immune evasion by the virus, underscoring the urgent need for antiviral drugs to expand upon disease therapies. Identification of novel antiviral compounds can be achieved through the development of a robust assay suitable for high throughput screening (HTS). This assay must demonstrate high reproducibility and low variability to ensure reliable screening. To evaluate assay robustness, the statistical parameter Z-factor (Z’) was employed. This study aimed to develop a HTS assay to measure SARS-CoV-2 entry, and to validate the assay for HTS applications. To assess SARS-CoV-2 entry, a vesicular stomatitis virus (VSV)-based pseudovirus system was utilized, enabling work under biosafety level 2 (BSL-2) conditions.