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Induction of the inflammasome by the SARS-CoV-2 accessory protein ORF9b, abrogated by small-molecule ORF9b homodimerization inhibitors
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  • Erika Zodda,
  • Mònica Pons,
  • Natàlia DeMoya-Valenzuela,
  • Cristina Calvo-González,
  • Cristina Benítez-Rodríguez,
  • Blanca D. López-Ayllón,
  • Achraf Hibot,
  • Marta Cascante,
  • María Montoya,
  • María Dolors Pujol,
  • Jaime Rubio-Martínez,
  • Timothy M. Thomson
Erika Zodda
Institut de Biologia Molecular de Barcelona
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Mònica Pons
Institut de Biologia Molecular de Barcelona
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Natàlia DeMoya-Valenzuela
Universitat de Barcelona Departament de Ciencia de Materials i Quimica Fisica
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Cristina Calvo-González
Institut de Biologia Molecular de Barcelona
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Cristina Benítez-Rodríguez
Institut de Biologia Molecular de Barcelona
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Blanca D. López-Ayllón
Centro de Investigaciones Biologicas Margarita Salas Biblioteca
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Achraf Hibot
Universitat de Barcelona Facultat de Farmacia i Ciencies de l'Alimentacio
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Marta Cascante
Universitat de Barcelona Departament de Bioquimica i Biologia Molecular
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María Montoya
Centro de Investigaciones Biologicas Margarita Salas Biblioteca
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María Dolors Pujol
Universitat de Barcelona Facultat de Farmacia i Ciencies de l'Alimentacio
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Jaime Rubio-Martínez
Universitat de Barcelona Departament de Ciencia de Materials i Quimica Fisica
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Timothy M. Thomson
Institut de Biologia Molecular de Barcelona

Corresponding Author:[email protected]

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Abstract

Viral accessory proteins play critical roles in viral escape form host innate immune responses and in viral inflammatory pathogenesis. Here we show that the SARS-CoV-2 accessory protein, ORF9b, but not other SARS-CoV-2 accessory proteins (ORF3a, ORF3b, ORF6, ORF7, ORF8, ORF9c, ORF10), strongly activates inflammasome-dependent caspase-1 in A549 lung carcinoma cells and THP-1 monocyte-macrophage cells. Exposure to lipopolysaccharide (LPS) and ATP additively enhanced the activation of caspase-1 by ORF9b, suggesting that ORF9b and LPS follow parallel pathways in the activation of the inflammasome and caspase-1. Following rational in silico approaches, we have designed small molecules capable of inhibiting the homodimerization of ORF9b, which experimentally inhibited ORF9b-ORF9b homotypic interactions, caused mitochondrial eviction of ORF9b, inhibited ORF9b-induced activation of caspase-1 in A549 and THP-1 cells, cytokine release in THP-1 cells, and restored type I interferon (IFN-I) signaling suppressed by ORF9b in both cell models. These small molecules are first-in-class compounds targeting a viral accessory protein critical for viral-induced exacerbated inflammation and escape from innate immune responses, with the potential of mitigating the severe immunopathogenic damage induced by highly pathogenic coronaviruses and restoring antiviral innate immune responses curtailed by viral infection.
Submitted to Journal of Medical Virology
12 Jun 2024Review(s) Completed, Editorial Evaluation Pending
15 Jun 2024Reviewer(s) Assigned
22 Jul 2024Editorial Decision: Revise Major