Stably-Inverted Apical-Out Human Upper Airway Organoids for SARS-CoV-2 Infection and Therapeutic Testing

Lee, J.-H.; LeCher, J. C.; Parigoris, E.; Shinagawa, N.; Sentosa, J.; Manfredi, C.; Goh, S. L.; De, R.; Tao, S.; Zandi, K.; Amblard, F.; Sorscher, E. J.; Spence, J. R.; Tirouvanziam, R.; Schinazi, R. F.; Takayama, S.

doi:10.1101/2024.01.02.573939

bioRxiv · 10.1101/2024.01.02.573939

Stably-Inverted Apical-Out Human Upper Airway Organoids for SARS-CoV-2 Infection and Therapeutic Testing

Lee, J.-H.·LeCher, J. C.·Parigoris, E.·Shinagawa, N.·Sentosa, J.·Manfredi, C.·Goh, S. L.·De, R.·Tao, S.·Zandi, K.·Amblard, F.·Sorscher, E. J.·Spence, J. R.·Tirouvanziam, R.·Schinazi, R. F.·Takayama, S.

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Abstract

While breakthroughs with organoids have emerged as next-generation in vitro tools, standardization for drug discovery remains a challenge. This work introduces human airway organoids with reversed biopolarity (AORBs), cultured and analyzed in a high-throughput, single-organoid-per-well format, enabling milestones towards standardization. AORBs exhibit a spatio-temporally stable apical-out morphology, facilitating high-yield direct intact-organoid virus infection. Single-cell RNA sequencing and immunohistochemistry confirm the physiologically relevant recapitulation of differentiated human airway epithelia. The cellular tropism of five severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) strains along with host response differences between Delta, Washington, and Omicron variants, as observed in transcriptomic profiles, also suggest clinical relevance. Dose-response analysis of three well-studied SARS-CoV-2 antiviral compounds (remdesivir, bemnifosbuvir, and nirmatrelvir) demonstrates that AORBs efficiently predict human efficacy, comparable to gold-standard air-liquid interface cultures, but with higher throughput ([~]10-fold) and fewer cells ([~]100-fold). This combination of throughput and relevance allows AORBs to robustly detect false negative results in efficacy, preventing irretrievable loss of promising lead compounds. While this work leverages the SARS-CoV-2 study as a proof-of-concept application, the standardization capacity of AORB holds broader implications in line with regulatory efforts to push alternatives to animal studies.

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Lee, J.-H., LeCher, J. C., Parigoris, E., Shinagawa, N., Sentosa, J., Manfredi, C., Goh, S. L., De, R., Tao, S., Zandi, K., Amblard, F., Sorscher, E. J., Spence, J. R., Tirouvanziam, R., Schinazi, R. F., Takayama, S.. 2024-01-04. Stably-Inverted Apical-Out Human Upper Airway Organoids for SARS-CoV-2 Infection and Therapeutic Testing. https://doi.org/10.1101/2024.01.02.573939

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