Human bocavirus 1(HBoV1;family:Parvoviridae)causes a wide spectrum of respiratory diseases in children and gastroenteritis in adults.A lack of sensitive cell lines and efficient animal models hinders research on HBoV,...Human bocavirus 1(HBoV1;family:Parvoviridae)causes a wide spectrum of respiratory diseases in children and gastroenteritis in adults.A lack of sensitive cell lines and efficient animal models hinders research on HBoV,including the development of anti-HBoV drugs or vaccines.Although the construction of a wild-type HBoV1 infectious clone has been reported,generating HBoV1 infectious clone carrying foreign reporter genes with suitable insertion sites in its genome while retaining replicative ability remains challenging.Here,HBoV1 infectious clones harboring the 11-amino-acid HiBiT tag at five distinct insertion sites were constructed and evaluated.Only the recombinant HBoV1 carrying the HiBiT tag in the N-terminus of the NS1 protein(HBoV1-HiBiTNS1)displayed comparable characteristics to wild-type HBoV1 as determined via the analysis of viral DNA copy number,NanoLuc activity,viral protein expression,and the formation of replication intermediates.Notably,the replication kinetics of HBoV1-HiBiTNS1 could be examined by monitoring NanoLuc activity,which was noted to be correlated with the viral DNA level.Additionally,we successfully applied HiBiT-tagged HBoV1 for the evaluation of antiviral drug activity and identified ivermectin(EC50=2.27μM)as a potent anti-HBoV1 replication drug.Overall,our study demonstrated that the HBoV1-HiBiTNS1 reporter can serve as a convenient platform for screening candidate drugs targeting HBoV1 replication and may also be useful for investigating the life cycle of the virus.展开更多
Due to the limitations of current anti-HBV therapies, the HBV core (HBc or HBcAg) protein assembly modulators (CpAMs) are believed to be potential anti-HBV agents. Therefore, discovering safe and efficient CpAMs is of...Due to the limitations of current anti-HBV therapies, the HBV core (HBc or HBcAg) protein assembly modulators (CpAMs) are believed to be potential anti-HBV agents. Therefore, discovering safe and efficient CpAMs is of great value. In this study, we established a HiBiT-based high-throughput screening system targeting HBc and screened novel CpAMs from an in-house marine chemicals library. A novel lead compound 8a, a derivative of the marine natural product naamidine J, has been successfully screened for potential anti-HBV activity. Bioactivity-driven synthesis was then conducted, and the structure‒activity relationship was analyzed, resulting in the discovery of the most effective compound 11a (IC50 = 0.24 μmol/L). Furthermore, 11a was found to significantly inhibit HBV replication in multiple cell models and exhibit a synergistic effect with tenofovir disoproxil fumarate (TDF) and IFNa2 in vitro for anti-HBV activity. Treatment with 11a in a hydrodynamic-injection mouse model demonstrated significant anti-HBV activity without apparent hepatotoxicity. These findings suggest that the naamidine J derivative 11a could be used as the HBV core protein assembly modulator to develop safe and effective anti-HBV therapies.展开更多
基金supported by the Guangzhou National Laboratory(SRPG22-002 to X.W.Chen)the Pearl River Talent Recruitment Program(2019CX01Y422 to X.W.Chen)+1 种基金the Natural Science Foundation of Guangdong Province(2025A1515011018 to J.L.Tang)the Basic and Applied Basic Research Projects of Guangzhou Basic Research Program(2025A04J5492 to J.L.Tang,2023A04J0161 to Q.Yang).
文摘Human bocavirus 1(HBoV1;family:Parvoviridae)causes a wide spectrum of respiratory diseases in children and gastroenteritis in adults.A lack of sensitive cell lines and efficient animal models hinders research on HBoV,including the development of anti-HBoV drugs or vaccines.Although the construction of a wild-type HBoV1 infectious clone has been reported,generating HBoV1 infectious clone carrying foreign reporter genes with suitable insertion sites in its genome while retaining replicative ability remains challenging.Here,HBoV1 infectious clones harboring the 11-amino-acid HiBiT tag at five distinct insertion sites were constructed and evaluated.Only the recombinant HBoV1 carrying the HiBiT tag in the N-terminus of the NS1 protein(HBoV1-HiBiTNS1)displayed comparable characteristics to wild-type HBoV1 as determined via the analysis of viral DNA copy number,NanoLuc activity,viral protein expression,and the formation of replication intermediates.Notably,the replication kinetics of HBoV1-HiBiTNS1 could be examined by monitoring NanoLuc activity,which was noted to be correlated with the viral DNA level.Additionally,we successfully applied HiBiT-tagged HBoV1 for the evaluation of antiviral drug activity and identified ivermectin(EC50=2.27μM)as a potent anti-HBoV1 replication drug.Overall,our study demonstrated that the HBoV1-HiBiTNS1 reporter can serve as a convenient platform for screening candidate drugs targeting HBoV1 replication and may also be useful for investigating the life cycle of the virus.
基金National Key Research and Development Program of China(2022YFA1303600,2021YFF0502400)National Natural Science Foundation of China(U23A20474,82022069)+5 种基金Shanghai Municipal Health Commission(GWVI-11.2-XD27,China)Hainan Provincial Major Science and Technology Project(ZDKJ2021028,China)Non-profit Central Research Institute Fund of Chinese Academy of Medical Sciences(2023-PT310-02,China)CAMS Innovation Fund for Medical Sciences(2019-12M-5-040,China)Shandong Laboratory Program(SYS202205,China)CAS Youth Interdisciplinary Team,and Taishan Scholars Program(tsqn202312302,China).
文摘Due to the limitations of current anti-HBV therapies, the HBV core (HBc or HBcAg) protein assembly modulators (CpAMs) are believed to be potential anti-HBV agents. Therefore, discovering safe and efficient CpAMs is of great value. In this study, we established a HiBiT-based high-throughput screening system targeting HBc and screened novel CpAMs from an in-house marine chemicals library. A novel lead compound 8a, a derivative of the marine natural product naamidine J, has been successfully screened for potential anti-HBV activity. Bioactivity-driven synthesis was then conducted, and the structure‒activity relationship was analyzed, resulting in the discovery of the most effective compound 11a (IC50 = 0.24 μmol/L). Furthermore, 11a was found to significantly inhibit HBV replication in multiple cell models and exhibit a synergistic effect with tenofovir disoproxil fumarate (TDF) and IFNa2 in vitro for anti-HBV activity. Treatment with 11a in a hydrodynamic-injection mouse model demonstrated significant anti-HBV activity without apparent hepatotoxicity. These findings suggest that the naamidine J derivative 11a could be used as the HBV core protein assembly modulator to develop safe and effective anti-HBV therapies.
