关键词: 前列腺特异膜抗原, 腺病毒载体, 基因治疗

Abstract: Objective To construct secretary recombinant adenoviral vector carrying the mouse prostate-specific membrane antigen gene through AdEasy vector system and the immune result was evaluated. Methods mPSMA was amplified from plasmid pCR-BluntII-TOPO by PCR and subcloned into transfer vector pAdenoVator CMV5, The signal peptide DNA sequence of hIL-2 was fused to 5′terminal of mPSMA gene to construct a secretary Ad-mPSMA. pAdv-mPSMA was co-transformed with pAdenoVator ΔE1/E3 through homologous recombination. The recombinant adenoviruses were packaged ,amplified and purified in HEK293 cells. HeLa cell was infected by recombinant adenovirus Ad-mPSMA and the expression of mouse prostate-specific membrane antigen gene was detected by RT-PCR and Western blot. The recombinant adenovirus had been immuned mice, sera antibody against mPSMA from immunized mice was detected by ELISA.Results The secretary pAd-mPSMA was constructed successfully and typical cytopathic effect (CPE) was observed. The titer of the recombinant adenovirus was 1.32x1011IU/ml and expression of mPSMA was confirmed by RT-PCR and Western blot. The specific antibody against mPSMA had been found in serum of the immunized mice. Conclusion Secretary mPSMA gene recombinant adenovirus was constructed successfully, which provide a basis for further study on the anti-tumor immunotherapy role of PSMA.

Key words: prostate-specific membrane antigen, adenoviral vector, gene therapy