Donor-derived airway air-liquid interface model for high-throughput screening of antiviral combinations with concurrent analysis of antiviral efficacy and epithelial toxicity using ciliR.

BACKGROUND: Respiratory RNA viral infections significantly impact quality of life and productivity, in terms of both pandemics (e.g. severe acute respiratory syndrome coronavirus 2, influenza A viruses) and seasonal infections, such as respiratory syncytial virus (RSV). Globally, there are an estimated 33.8 million cases of RSV annually in children aged <5 years, leading to 2.8-4.3 million hospital admissions and up to 199 000 deaths. Human-relevant drug screening models are key to addressing the urgent need for effective antiviral treatments for RSV, particularly in vulnerable patient groups.

METHODS: Here, we present a donor-derived differentiated primary human nasal epithelial cell, high-throughput screening (HTS) Transwell-96 air-liquid interface (ALI) model designed to investigate the effects of combination antiviral therapies on RSV infection in primary human ciliated airway epithelium. In addition, we describe novel analytical tools using R (ciliR) to screen drug combinations by concurrently measuring efficacy and ciliary beat frequency, which we used as a sensitive marker of cell toxicity.

RESULTS: Our results demonstrate that the higher-throughput HTS Transwell-96 ALI cultures retain epithelial composition and ciliary function consistent within donor replicates. These cultures are permissible to infection with an RSV-green fluorescent protein (GFP) reporter virus, enabling quantitative comparison and combination treatment across multiple epithelial cultures from the same donor.

CONCLUSION: We anticipate that our disease-relevant system will serve as a foundation for larger-scale experiments aimed at optimising combination therapy for RSV and other respiratory viruses.