Objective: To study the use of preS1-tp fusion
protein as a novel vector to mediate the entry of
small interfering RNA (
siRNA) targeting the carboxy-terminal
nuclear localization signal (NLS) region of hepatitis B virus (HBV) core
protein into HBV-infected hepatocytes, and to further explore the HBV replication inhibition and covalently closed
circular DNA synthesis. Methods: HepG2.2.15 cells expressing the human
sodium taurocholate co-transporting
polypeptide were established on the basis of
lentivirus infection system.
siRNA against HBV NLS region was designed and synthesized. PreS1-tp fusion
protein expression and purification was observed to test its ability to cell entry and
DNA binding. NLS
siRNA were delivered into HepG2.2.15-
sodium taurocholate co-transporting
polypeptide cells by preS1-tp fusion
protein as a vector to observe the effects of NLS
siRNA on HBV replication and covalently closed
circular DNA levels. Analysis of variance was used for comparison between multiple groups, and the measurement data differences between groups were analyzed by t-test. Results: HepG2.2.15-sodium
taurocholate co-transporting
polypeptide cell line was successfully constructed. Screened synthetic HBV NLS
siRNA had significantly inhibited HBV replication. The preS1-tp fusion
protein was expressed and purified on a large-scale. The fusion
protein as a vector for HBV NLS
siRNA had targeted delivery. The result showed that the fusion
protein had effectively targeted
siRNA to Hepg2.2.15-sodium
taurocholate co-transporting
polypeptide cell, which not only had effectively inhibited the expression of HBV
mRNA,
HBsAg and
HBeAg, but also had significantly reduced the levels of HBV
DNA and covalently closed
circular DNA. Conclusion: The preS1-tp fusion
protein constructed in this study uses the dual functional characteristics of preS1 binding to hepatocyte HBV receptor, and tp binding to
nucleic acids, and targets HBV NLS
siRNA against HBV-infected cells and block rcDNA from being transported to nucleus.
siRNA plays a role in inhibiting HBV replication and covalently close
circular DNA synthesis, providing a new strategy for the treatment of
chronic hepatitis B caused by HBV
infection, and a new research perspective for the complete elimination of HBV from the body.