In cancer gene therapies, it is ideal to target tumor-specific genes. Since telomerase is activated in almost all cancer cells but not most somatic cells, it is considered as one of the favorite targets for cancer gene therapies. Ribozymes are catalytic RNA molecules with site-specific endoribonuclease activity. In the present study, we designed hammerhead ribozymes against human telomerase RNA (hTR) and human reverse transcriptase subunit (hTERT) to evaluate their effect on the attenuation of telomerase in the pancreas cancer cell line, PK-8. Hammerhead ribozyme targeting hTR depressed the level of telomerase activity in PK-8 cells. pRc-hTR vector with ribozyme targeting hTR and pRc-hTERT vector with ribozyme targeting hTERT mRNA were transfected into PK-8 cells and depressed telomerase activity and target RNA, but in pRc-hTR transfectant, hTERT mRNA expression was slightly upregulated and in pRc-hTERT transfectant hTR expression was also slightly upregulated. These findings indicate the co-regulation of hTR and hTERT mRNA expression in cancer cells. Extrachromosomal replicational vector, pCEP4, containing ribozyme targeting hTERT mRNA showed the most effective inhibition of telomerase activity, suggesting that the continuous effect of ribozyme is necessary to inhibit telomerase activity. Since the level of hTERT mRNA expression is less than that of hTR expression in cancer cells, ribozyme targeting hTERT mRNA might be a more useful therapeutic strategy for cancer gene therapy. Moreover, the co-regulation of hTR and hTERT mRNA expression in cancer cells to maintain the levels of telomerase activity suggested that the strategy of inhibiting hTERT mRNA and hTR simultaneously has a powerful potential as a gene therapy for targeting human telomerase.