[Anti-tumor effect of adenovirus vector generated from homologous recombination of bacteria mediating TK gene on hepatic neoplasm cell].

OBJECTIVE

To test the anti-tumor effect of adenovirus vector generated from the homologous recombination of bacteria mediating TK gene on hepatic neoplasm cell.

METHODS

TK gene was liberated from plasmid and subcloned into shuttle plasmid, which formed the plasmid of pAdTrack-GFP-TK. The pAdTrack-GFP-TK plasmid was then linealized with Pme I and cotransformed into BJ5183 bacterial cells with adenovirus genomic DNA plasmid of pAdEasy-1. The recombinant adenovirus plasmid DNA was digested with Pac I and transfected to the 293 cells to package the recombinant adenovirus particles. The infection rate of the recombinant adenovirus, the target gene expression in the carcinoma cells, the survival rate of the GCV treated hepatic neoplasm cell SMMC-7721 infected with recombinant adenovirus, and the bystander effect were measured.

RESULTS

The titre of purified Ad was 2 x 10(12) efu/mL. More than 90% of the SMMC-7721 cells were infected when the MOI reached 100. Target gene expression was detected in the infected cells. The SMMC-7721 cells infected with AdEasy-GFP-TK were very sensitive to the prodrug GCV, with IC50 less than 0.2 pmol/L. Significant bystander effect was observed.

CONCLUSION

The constructed AdEasy-GFP-TK virus can efficiently infect the hepatic neoplasm cells and enable the SMMC-7721 cells to express the HSV-TK genes. The bystander effect has improved the anti-tumor effect of the TK gene, which makes the clinical application possible.