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Topics {✒️}

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         headline:Brequinar inhibits African swine fever virus replication in vitro by activating ferroptosis
         description:African swine fever virus (ASFV) is one of the most fatal swine etiological agents and has a huge economic impact on the global pork industry. Given that no effective vaccines or anti-ASFV drugs are available, there remains a pressing need for novel anti-ASFV drugs. This study aimed to investigate the anti-African swine fever virus (ASFV) activity of brequinar, a DHODH inhibitor. The anti-ASFV activity of brequinar was investigated using IFA, HAD, HAD50, qRT-PCR, and western blotting assays. The western blotting assay was used to investigate whether brequinar inhibits ASFV replication by killing ASFV particles directly or by acting on cell factors. The confocal microscopy and western blotting assays were used to investigate whether brequinar inhibits ASFV replication by activating ferroptosis. In this study, brequinar was found to effectively inhibit ASFV replication ex vivo in porcine alveolar macrophages (PAMs) in a dose-dependent manner. In kinetic studies, brequinar was found to maintain ASFV inhibition from 24 to 72 hpi. Mechanistically, the time-of-addition assay showed that brequinar exerted anti-ASFV activity in all treatment modes, including pre-, co-, and post-treatment rather than directly killing ASFV particles. Notably, FerroOrange, Mito-FerroGreen, and Liperfluo staining experiments showed that brequinar increased the accumulation of intracellular iron, mitochondrial iron, and lipid peroxides, respectively. Furthermore, we also found that ferroptosis agonist cisplatin treatment inhibited ASFV replication in a dose-dependent manner and the inhibitory effect of brequinar on ASFV was partially reversed by the ferroptosis inhibitor ferrostatin-1, suggesting that brequinar activates ferroptosis to inhibit ASFV replication. Interestingly, exogenous uridine supplementation attenuated the anti-ASFV activity of brequinar, indicating that brequinar inhibits ASFV replication by inhibiting DHODH activity and the depletion of intracellular pyrimidine pools; however, the induction of ferroptosis by brequinar treatment was not reversed by exogenous uridine supplementation, suggesting that brequinar activation of ferroptosis is not related to the metabolic function of pyrimidines. Our data confirm that brequinar displays potent antiviral activity against ASFV in vitro and reveal the mechanism by which brequinar inhibits ASFV replication by activating ferroptosis, independent of inhibiting pyrimidine synthesis, providing novel targets for the development of anti-ASFV drugs.
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      headline:Brequinar inhibits African swine fever virus replication in vitro by activating ferroptosis
      description:African swine fever virus (ASFV) is one of the most fatal swine etiological agents and has a huge economic impact on the global pork industry. Given that no effective vaccines or anti-ASFV drugs are available, there remains a pressing need for novel anti-ASFV drugs. This study aimed to investigate the anti-African swine fever virus (ASFV) activity of brequinar, a DHODH inhibitor. The anti-ASFV activity of brequinar was investigated using IFA, HAD, HAD50, qRT-PCR, and western blotting assays. The western blotting assay was used to investigate whether brequinar inhibits ASFV replication by killing ASFV particles directly or by acting on cell factors. The confocal microscopy and western blotting assays were used to investigate whether brequinar inhibits ASFV replication by activating ferroptosis. In this study, brequinar was found to effectively inhibit ASFV replication ex vivo in porcine alveolar macrophages (PAMs) in a dose-dependent manner. In kinetic studies, brequinar was found to maintain ASFV inhibition from 24 to 72 hpi. Mechanistically, the time-of-addition assay showed that brequinar exerted anti-ASFV activity in all treatment modes, including pre-, co-, and post-treatment rather than directly killing ASFV particles. Notably, FerroOrange, Mito-FerroGreen, and Liperfluo staining experiments showed that brequinar increased the accumulation of intracellular iron, mitochondrial iron, and lipid peroxides, respectively. Furthermore, we also found that ferroptosis agonist cisplatin treatment inhibited ASFV replication in a dose-dependent manner and the inhibitory effect of brequinar on ASFV was partially reversed by the ferroptosis inhibitor ferrostatin-1, suggesting that brequinar activates ferroptosis to inhibit ASFV replication. Interestingly, exogenous uridine supplementation attenuated the anti-ASFV activity of brequinar, indicating that brequinar inhibits ASFV replication by inhibiting DHODH activity and the depletion of intracellular pyrimidine pools; however, the induction of ferroptosis by brequinar treatment was not reversed by exogenous uridine supplementation, suggesting that brequinar activation of ferroptosis is not related to the metabolic function of pyrimidines. Our data confirm that brequinar displays potent antiviral activity against ASFV in vitro and reveal the mechanism by which brequinar inhibits ASFV replication by activating ferroptosis, independent of inhibiting pyrimidine synthesis, providing novel targets for the development of anti-ASFV drugs.
      datePublished:2023-10-24T00:00:00Z
      dateModified:2023-10-24T00:00:00Z
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         Brequinar
         Ferroptosis
         Uridine
         Virology
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         type:PostalAddress
      name:African Swine Fever Regional Laboratory of China (Guangzhou)
      address:
         name:African Swine Fever Regional Laboratory of China (Guangzhou), Guangzhou, China
         type:PostalAddress
      name:Maoming Branch, Guangdong Laboratory for Lingnan Modern Agriculture
      address:
         name:Maoming Branch, Guangdong Laboratory for Lingnan Modern Agriculture, Maoming, China
         type:PostalAddress
ImageObject:
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Person:
      name:Yang Chen
      affiliation:
            name:South China Agricultural University
            address:
               name:Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China
               type:PostalAddress
            type:Organization
            name:African Swine Fever Regional Laboratory of China (Guangzhou)
            address:
               name:African Swine Fever Regional Laboratory of China (Guangzhou), Guangzhou, China
               type:PostalAddress
            type:Organization
      name:Yanchen Guo
      affiliation:
            name:South China Agricultural University
            address:
               name:Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China
               type:PostalAddress
            type:Organization
            name:African Swine Fever Regional Laboratory of China (Guangzhou)
            address:
               name:African Swine Fever Regional Laboratory of China (Guangzhou), Guangzhou, China
               type:PostalAddress
            type:Organization
            name:Maoming Branch, Guangdong Laboratory for Lingnan Modern Agriculture
            address:
               name:Maoming Branch, Guangdong Laboratory for Lingnan Modern Agriculture, Maoming, China
               type:PostalAddress
            type:Organization
      name:Hao Chang
      affiliation:
            name:South China Agricultural University
            address:
               name:Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China
               type:PostalAddress
            type:Organization
            name:African Swine Fever Regional Laboratory of China (Guangzhou)
            address:
               name:African Swine Fever Regional Laboratory of China (Guangzhou), Guangzhou, China
               type:PostalAddress
            type:Organization
      name:Zebu Song
      affiliation:
            name:South China Agricultural University
            address:
               name:Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China
               type:PostalAddress
            type:Organization
            name:Maoming Branch, Guangdong Laboratory for Lingnan Modern Agriculture
            address:
               name:Maoming Branch, Guangdong Laboratory for Lingnan Modern Agriculture, Maoming, China
               type:PostalAddress
            type:Organization
      name:Zhi Wei
      affiliation:
            name:South China Agricultural University
            address:
               name:Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China
               type:PostalAddress
            type:Organization
            name:African Swine Fever Regional Laboratory of China (Guangzhou)
            address:
               name:African Swine Fever Regional Laboratory of China (Guangzhou), Guangzhou, China
               type:PostalAddress
            type:Organization
      name:Zhao Huang
      affiliation:
            name:South China Agricultural University
            address:
               name:Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China
               type:PostalAddress
            type:Organization
            name:African Swine Fever Regional Laboratory of China (Guangzhou)
            address:
               name:African Swine Fever Regional Laboratory of China (Guangzhou), Guangzhou, China
               type:PostalAddress
            type:Organization
      name:Zezhong Zheng
      affiliation:
            name:South China Agricultural University
            address:
               name:Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China
               type:PostalAddress
            type:Organization
            name:Maoming Branch, Guangdong Laboratory for Lingnan Modern Agriculture
            address:
               name:Maoming Branch, Guangdong Laboratory for Lingnan Modern Agriculture, Maoming, China
               type:PostalAddress
            type:Organization
      name:Guihong Zhang
      affiliation:
            name:South China Agricultural University
            address:
               name:Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China
               type:PostalAddress
            type:Organization
            name:African Swine Fever Regional Laboratory of China (Guangzhou)
            address:
               name:African Swine Fever Regional Laboratory of China (Guangzhou), Guangzhou, China
               type:PostalAddress
            type:Organization
            name:Maoming Branch, Guangdong Laboratory for Lingnan Modern Agriculture
            address:
               name:Maoming Branch, Guangdong Laboratory for Lingnan Modern Agriculture, Maoming, China
               type:PostalAddress
            type:Organization
            name:Key Laboratory of Animal Vaccine Development, Ministry of Agriculture and Rural Affairs
            address:
               name:Key Laboratory of Animal Vaccine Development, Ministry of Agriculture and Rural Affairs, Guangzhou, China
               type:PostalAddress
            type:Organization
      email:[email protected]
      name:Yankuo Sun
      affiliation:
            name:South China Agricultural University
            address:
               name:Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China
               type:PostalAddress
            type:Organization
            name:African Swine Fever Regional Laboratory of China (Guangzhou)
            address:
               name:African Swine Fever Regional Laboratory of China (Guangzhou), Guangzhou, China
               type:PostalAddress
            type:Organization
            name:Maoming Branch, Guangdong Laboratory for Lingnan Modern Agriculture
            address:
               name:Maoming Branch, Guangdong Laboratory for Lingnan Modern Agriculture, Maoming, China
               type:PostalAddress
            type:Organization
      email:[email protected]
PostalAddress:
      name:Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China
      name:African Swine Fever Regional Laboratory of China (Guangzhou), Guangzhou, China
      name:Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China
      name:African Swine Fever Regional Laboratory of China (Guangzhou), Guangzhou, China
      name:Maoming Branch, Guangdong Laboratory for Lingnan Modern Agriculture, Maoming, China
      name:Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China
      name:African Swine Fever Regional Laboratory of China (Guangzhou), Guangzhou, China
      name:Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China
      name:Maoming Branch, Guangdong Laboratory for Lingnan Modern Agriculture, Maoming, China
      name:Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China
      name:African Swine Fever Regional Laboratory of China (Guangzhou), Guangzhou, China
      name:Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China
      name:African Swine Fever Regional Laboratory of China (Guangzhou), Guangzhou, China
      name:Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China
      name:Maoming Branch, Guangdong Laboratory for Lingnan Modern Agriculture, Maoming, China
      name:Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China
      name:African Swine Fever Regional Laboratory of China (Guangzhou), Guangzhou, China
      name:Maoming Branch, Guangdong Laboratory for Lingnan Modern Agriculture, Maoming, China
      name:Key Laboratory of Animal Vaccine Development, Ministry of Agriculture and Rural Affairs, Guangzhou, China
      name:Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China
      name:African Swine Fever Regional Laboratory of China (Guangzhou), Guangzhou, China
      name:Maoming Branch, Guangdong Laboratory for Lingnan Modern Agriculture, Maoming, China

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