Abstract
Rhinoviruses (RV) comprise three species, RV-A, RV-B, and RV-C, with approximately 170 types. RV-C is associated with severe respiratory illness, particularly in children and individuals with asthma or chronic obstructive pulmonary disease, underscoring the need for effective antiviral strategies. Progress in RV-C research and drug discovery has been limited by the lack of robust, scalable cell-based infection models that recapitulate the complete RV-C replication cycle. Here, we describe a high-content imaging (HCI)-based high-throughput infection system for RV-C. Rather than relying solely on receptor overexpression, we used a genetically stable fluorescent reporter virus (RV-C15a-mGL) to screen ~300 monoclonal cell lines expressing the RV-C receptor variant CDHR3-Tyr529. This approach identified a clone that efficiently supports RV-C replication and revealed that productive infection depends on determinants beyond receptor abundance alone. Using this clone, we established and validated a robust, scalable screening platform with Z′ > 0.75 in both 96- and 384-well formats. The system was readily adapted to additional RV-C types (C11 and C41), as well as RV-A and RV-B. A pilot screen of approximately 10,000 small molecules identified both known and novel RV-C inhibitors, supporting the utility of this platform for antiviral discovery and for advancing the study of RV-C biology.
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Acknowledgements
HeLa Rh was kindly provided by Dr. K. Andries (Janssen Pharmaceutica, Belgium). We thank Jasper Rymenants for technical assistance with the experiments involving human epithelial cell cultures, and Dirk Jochmans for supervision of these experiments. We also thank Nelleke Cloet for preparing and providing the pre-spotted assay plates, and Hugo Klaassen for supervising this work. The HTS was supported by the National Research Foundation of Korea (NRF) grant funded by the Korean government Ministry of Science and ICT (No. RS-2024-00432287). The has been posted to bioRxiv50.
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Lyoo, H., Alpizar, Y.A., Sablon, C. et al. A robust cell-based infection model for Rhinovirus C research and antiviral drug discovery. npj Viruses (2026). https://doi.org/10.1038/s44298-026-00194-5
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DOI: https://doi.org/10.1038/s44298-026-00194-5
