Objective: To construct a new gene therapy plasmid that can express both hepatitis B surface antibody(HBsAb) targeted interferon (dsFvα) and human interleukin 12 (hIL-12) genes for the immunotherapy of chronic hepatitis B.
Methods: The pEE14.1-dsFvα plasmid was digested to obtain dsFvα fragment, and then the fragment was cloned into the upstream of IRES sequence in vector pVAX-IRES-hIL-12 digested by the same enzyme to construct the recombinant expression plasmid pVAX-HBVE. The recombinant plasmid was transiently transfected into the HEK293T cells and the expression of target gene was detected by ELISA. The recombinant plasmid pVAX-HBVE was extracted and injected into the leg muscles of HBV transgenic mice with electroporation delivery. The HBV gene copies were detected by quantitative PCR before and after injection.
Results: Enzyme digestion and sequencing analysis showed that the recombinant plasmid pVAX-HBVE was constructed as expected before. ELISA showed that dsFvα gene and IL-12 gene were successfully expressed in the supernatant of transfected cells. The recombinant plasmid pVAX-HBVE (30 μg, once) reduced the HBV gene copy by two orders of magnitude after being injected into transgenic mice.
Conclusion: The neo-type immune gene therapy plasmid was successfully constructed, which would provide an alternative for immune gene therapy of chronic hepatitis B.