The usefulness of human telomerase reverse transcriptase (hTERT) gene promoter has been proposed in cancer-targeted gene therapy. However, this promoter may not be strong enough to achieve therapeutic levels of transgene expression. In this study, we tested an 'indirected-activator' strategy that utilizes radiation to increase the activity of the hTERT gene promoter. We demonstrated that hTERT may participate in the process of DNA repair induced by irradiation. We found that Zidovudine (AZT, an hTERT inhibitor) can decrease the telomerase activity in human HEp-2 larynx squamous carcinoma cells and lower the survival fraction of HEp-2 cells exposed to radiation. In HEp-2 cells exposed to 6 Gy-radiation, the hTERT promoter showed 2.9-fold higher activity compared with unirradiated cells. Importantly, an increased expression of enzyme horseradish peroxidase (HRP) controlled by the hTERT promoter was found in the transfected cells after irradiation, which coincided with a higher killing rate for HEp-2 cells after prodrug indole-3-acetic acid (IAA; converted by HRP into a cytotoxin) incubation combined with irradiation or not. Our observations suggest that hTERT promoter-mediated gene therapy could be improved in combination with radiotherapy, which may be due to cellular DNA damage responses.