A pseudovirus system for SARS-CoV-2 for screening antiviral agents and phytochemical extracts

The development of BSL-2-safe antiviral screening platforms is essential for addressing emerging viral outbreaks such as SARS-CoV-2. This study aimed to construct SARS-CoV-2 spike pseudoviruses (PSV) and assess their infectivity and applicability for antiviral screening across multiple human cell lines. PSVs were generated using a human immunodeficiency virus (HIV)-based vector system. PSV production was confirmed through viral gene expression, green fluorescent protein expression, and transmission electron microscopy. Cell susceptibility against PSV infection was assessed via fluorescent readouts and cell viability assays. Antiviral screening was performed using remdesivir, chloroquine, and previously characterized plant extracts. Viral entry gene expression was evaluated using reverse transcription-polymerase chain reaction (RT-PCR). The developed PSV system produced infectious titers comparable to those reported in previously established systems. Cellular susceptibility to PSV infection was likely associated with variations in the expression levels of viral entry receptor genes. These findings also demonstrate that antiviral response was cell-type dependent and suggest <i>Clinacanthus nutans</i> stem ethyl acetate extract (CNSEA) as a promising candidate for therapeutic investigation. This PSV system is safe, practical, and well-suited for antiviral screening. The findings contribute to emerging evidence that cancer cell susceptibility to SARS-CoV-2 is influenced by distinct viral entry receptor profiles.