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High-Throughput Screening of Antiviral Compounds Using a Recombinant Hepatitis B Virus and Identification of a Possible Infection Inhibitor, Skimmianine.

ABSTRACT: We developed a novel hepatitis B virus (HBV) infection-monitoring system using a luminescent, 11-amino acid reporter (HiBiT). We performed high-throughput antiviral screening using this system to identify anti-HBV compounds. After the infection of primary human hepatocytes with the recombinant virus HiBiT-HBV, which contains HiBiT at its preS1, 1262 compounds were tested in a first screening using extracellular HiBiT activity as an indicator of viral infection. Following a second screening, we focused on the compound skimmianine, which showed a potent antiviral effect. When skimmianine was added at the same time as HiBiT-HBV infection, skimmianine inhibited HiBiT activity with EC₅₀ of 0.36 pM, CC₅₀ of 1.67 μM and a selectivity index (CC₅₀:EC₅₀ ratio) of 5,100,000. When skimmianine was added 72 h after HiBiT-HBV infection, the EC₅₀, CC₅₀ and selectivity index were 0.19 μM, 1.87 μM and 8.79, respectively. Time-lapse fluorescence imaging analysis using another recombinant virus, ReAsH-TC155HBV, with the insertion of tetra-cysteine within viral capsid, revealed that skimmianine inhibited the accumulation of the capsid into hepatocytes. Furthermore, skimmianine did not inhibit either attachment or internalization. These results imply that skimmianine inhibits the retrograde trafficking of the virus after internalization. This study demonstrates the usefulness of the recombinant virus, HiBiT-HBV, for high-throughput screening to identify anti-HBV compounds.

SUBMITTER: Yoshita M

PROVIDER: S-EPMC11360121 | biostudies-literature | 2024 Aug

REPOSITORIES: biostudies-literature

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High-Throughput Screening of Antiviral Compounds Using a Recombinant Hepatitis B Virus and Identification of a Possible Infection Inhibitor, Skimmianine.

Yoshita Mika M Funaki Masaya M Shimakami Tetsuro T Kakuya Masaki M Murai Kazuhisa K Sugimoto Saiho S Kawase Shotaro S Matsumori Koji K Kawane Taro T Nishikawa Tomoki T Nakamura Asuka A Suzuki Reo R Ishida Atsuya A Kawasaki Narumi N Sato Yuga Y Li Ying-Yi YY Sumiyadorj Ariunaa A Nio Kouki K Takatori Hajime H Kawaguchi Kazunori K Kuroki Kazuyuki K Kato Takanobu T Honda Masao M Yamashita Taro T

Viruses 20240822 8

We developed a novel hepatitis B virus (HBV) infection-monitoring system using a luminescent, 11-amino acid reporter (HiBiT). We performed high-throughput antiviral screening using this system to identify anti-HBV compounds. After the infection of primary human hepatocytes with the recombinant virus HiBiT-HBV, which contains HiBiT at its preS1, 1262 compounds were tested in a first screening using extracellular HiBiT activity as an indicator of viral infection. Following a second screening, we f ...[more]

PMID: 39205320

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Project description:American elderberry (Sambucus nigra subsp. canadensis) is a rapidly emerging new perennial crop for Missouri, recognized for its high level of bioactive compounds with significant health benefits, including antibacterial, antiviral, and antioxidant properties. A high-throughput screening assay combined with untargeted metabolomics analysis was utilized on American elderberry juice from 21 genotypes to explore and characterize these bioactive compounds. Our metabolomics study has identified 32 putative bioactive compounds in the American Elderberry juices. An array of high-throughput screening bioassays was conducted to evaluate 1) total antioxidant capacity, 2) activation of antioxidant response elements (ARE), 3) antiviral activity, and 4) antibacterial activity of the putatively identified compounds. Our results revealed that 14 of the 32 American elderberry compounds exhibited strong antioxidant activity. Four compounds (isorhamnetin 3-O-glucoside, kaempferol, quercetin, and naringenin) activated ARE activity and were found to be non-cytotoxic to cells. Notably, six of the 32 compounds demonstrated significant antiviral activity in an in vitro TZM-bl assay against two strains of HIV-1 virus, CXCR4-dependent NL4-3 virus and CCR5-dependent BaL virus. Luteolin showed the most potent anti-HIV activity against the NL4-3 virus (IC50 = 1.49 μM), followed by isorhamnetin (IC50 = 1.67 μM). The most potent anti-HIV compound against the BaL virus was myricetin (IC50 = 1.14 μM), followed by luteolin (IC50 = 4.38 μM). Additionally, six compounds were found to have antibacterial activity against gram-positive bacteria S. aureus, with cyanidin 3-O-rutinoside having the most potent antibacterial activity in vitro (IC50 = 2.9 μM), followed by cyanidin 3-O-glucoside (IC50 = 3.7 μM). These findings support and validate the potential health benefits of compounds found in American elderberry juices and highlight their potential for use in dietary supplements as well as innovative applications in health and medicine.

Dataset Information

High-Throughput Screening of Antiviral Compounds Using a Recombinant Hepatitis B Virus and Identification of a Possible Infection Inhibitor, Skimmianine.

Publications

High-Throughput Screening of Antiviral Compounds Using a Recombinant Hepatitis B Virus and Identification of a Possible Infection Inhibitor, Skimmianine.

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