Targeted adenovirus containing r-caspase-3 gene induces apoptosis of hepatocellular carcinoma cells in vitro

Objective To investigate the effect and specificity of adenovirus containing r-caspase-3 gene on apoptosis of hepatocellular carcinoma cells. Methods The vector α-fetoprotein enhancer-albumin promotor （pAdTraek-EArv-PALB） was constructed and the r-caspase-3 gene was subcloned into the vector. The linearized shuttle plasmid was homogenously recombined with AdEasy-l in BJ5183 cells. The candidate clone was analyzed by restriction endonuclease digestion and sequencing, and then pAdeasy-EAFP-PALS/r- caspase-3 vector was digested with Pac I and transfected into AD293 cells for packaging and amplifying. Infection titer and rate of recombinant virus was monitored by green fluorescent protein （GFP） expression. The expression of r-caspase-3 was detected by RT-PCR and Western-blot. The apoptosis of HepG2, 7721, L- 02 and MDA- MB- 231 cells was detected by FCM method, Results The sequence of shuttle vector pAdTrack-EArp-PALB/r-caspase-3 was correct after digestion by restriction endonuclease. Vector pAdeasy-EAvv- PALB/r-caspase- 3 was identificated by Pac I restriction digestion and PCR. The expression of GFP was observed in the transfected AD293 cells. The expression of r-caspase-3 gene was detected in the infected HepG2 cells by RT-PCR and Western-blot. 3＇he cells were infected with recombinant r-caspase-3 after 24 hours, and their apoptotic index were as follows： HepG ceils, 48.2%; 7721 cells, 17.7%; L-02 ceils, 7.3%; MDA- MB- 231 cells, 0%. Conclusion The recombinant of hepatocellular carcinoma- targeting adenovirus containing r- caspase- 3 gene is constructed successfully and can induce the targeted apoptosis of hepatocellular carcinoma cells, which provides the evidences for future research in hepatocellular carcinoma. Key words: Adenovirus;  Caspase 3 ;  Liver heoplasms ;  Apoptosis;  Target