PI4KIII3β inhibition reduces rhinovirus associated cell shedding and ciliary dysfunction

Velkova, S.; Petris, A.; Lee, D.; Cardinale, D.; Moulding, D.; Williamson, R.; Beinke, S.; Grace, K.; Hessel, E.; Belyaev, N.; Hoegg, T.; Hurst, J.; Smyth, R.; Smith, C.; Freestone, P.; O'Callaghan, C.

doi:10.1101/2023.12.14.571686

bioRxiv · 10.1101/2023.12.14.571686

PI4KIII3β inhibition reduces rhinovirus associated cell shedding and ciliary dysfunction

Velkova, S.·Petris, A.·Lee, D.·Cardinale, D.·Moulding, D.·Williamson, R.·Beinke, S.·Grace, K.·Hessel, E.·Belyaev, N.·Hoegg, T.·Hurst, J.·Smyth, R.·Smith, C.·Freestone, P.·O'Callaghan, C.

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Abstract

BackgroundPatients with chronic obstructive pulmonary disease (COPD) experience respiratory exacerbations, many of which are associated with rhinoviruses. Current treatment strategies do not target the pathogenic rhinovirus trigger. Research questionWhat is the immediate effect of rhinovirus on the ciliated respiratory epithelium and can viral replication and epithelial toxicity be reduced by targeted PI4KIII{beta} inhibition. MethodsShort (24h) and longer (7 days) rhinovirus infection were explored in primary ciliated airway epithelial cultures from multiple healthy and COPD patients using high-speed video microscopy, viral titration assays and immunofluorescence studies. Ciliated epithelial cultures were pre-treated with a PI4KIII{beta} (GSK533) blocker prior to infection to assess efficacy against rhinovirus. Cytokine and chemokine production were assessed by multiplex immunoassays. ResultsWithin hours of infection rhinovirus co-localised with ciliated cells causing extensive apoptosis-associated shedding of predominantly ciliated cells within 24 hours. Viral replication that peaked at day 1 and cleared by day 7, was associated with dramatic loss of ciliated cells confirmed by reduced ciliary activity and ciliary DNAI2 protein expression. Ciliary beat frequency (CBF) of remaining cilia was significantly reduced by day 7 in cultures from COPD. Infection was partly dependent on PI4KIII{beta} with the GSK533 blocker reducing viral replication while preserving ciliary activity. High levels of pro-inflammatory mediators were secreted by infected cells. ConclusionDecreased ciliation due to rhinovirus infection is likely to impair mucociliary clearance in healthy individuals and COPD patients, contributing to the pathophysiology of COPD exacerbations. PI4KIII{beta} inhibition blocks viral replication, helping to preserve ciliary activity. Take home messageRhinovirus replication in the healthy and COPD respiratory epithelium is mediated by PI4KIII{beta} and intracellular PI4P platform formation. Inhibition of PI4KIII{beta} reduced viral replication and ciliated cell loss.

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Velkova, S., Petris, A., Lee, D., Cardinale, D., Moulding, D., Williamson, R., Beinke, S., Grace, K., Hessel, E., Belyaev, N., Hoegg, T., Hurst, J., Smyth, R., Smith, C., Freestone, P., O'Callaghan, C.. 2023-12-14. PI4KIII3β inhibition reduces rhinovirus associated cell shedding and ciliary dysfunction. https://doi.org/10.1101/2023.12.14.571686

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