Published online Sep 15, 2003. doi: 10.3748/wjg.v9.i9.2030
Revised: April 23, 2003
Accepted: May 24, 2003
Published online: September 15, 2003
AIM: To investigate the anti-tumor mechanism of antisense oligodeoxynucleotide cantide against hTERT.
METHODS: Tumor cells were cultured overnight and grown to 50%-60% confluence. HepG2 and SMMC-7721 were treated with cantide mixed with lipofectin, or lipofectin alone. After inducted for 6 h at 37 °C, 10% FCS in DMEM was replaced in each well. After the treatment repeated twice to three times in each concentration of cantide, hTERT mRNA and protein expression were measured by RT-PCR and Western blot analysis, respectively. Telomerase activity was determined by TRAP-ELISA assay. CPP32- and ICE-like activity was also investigated using CasPACE assay system at 48 h after cantide treatment, and apoptosis was evaluated using the DeadEnd assay at 24, 48 and 72 h after cantide treatment.
RESULTS: Compared to the control cells, the cells treated with cantide showed a dose-dependent decrease in hTERT mRNA levels at 24 h and in protein levels at 48 h respectively. The telomerase activity was decreased as the concentration of cantide increased at 48 h. At the concentration of 800 nM, the telomerase activity in the treated HepG2 and SMMC-7721 cells was only 17.1% (P < 0.01) and 20.3% (P < 0.01) of that in untreated cells. The levels of CPP32-like protease activity in HepG2 and SMMC-7721 increased by 2.8- and 3.0-fold (P < 0.05) at 48 h, and the levels of ICE-like protease activity also increased by 2.6- and 3.2-fold (P < 0.05) respectively. The percentage of apoptosis in HepG2 and SMMC-7721 cells treated with 800 nM cantide at 72 h was 63% and 52% (P < 0.01), respectively. By contrast, 8% and 9% of the cells were apoptosis after 72 h treatment with lipofectin alone.
CONCLUSION: Cantide can decrease telomerase activity by inhibiting the expression of hTERT gene and has a rapid anti-tumor effect through inducing the Caspase-dependent apoptosis. The rapid inhibitory effect of cantide on tumor growth demonstrates its feasibility in cancer treatment.