Changes of telomerase activity and proliferation by inhibition of reverse transcriptase activity in human cancer cell.

Abstract

Purpose: Activation of telomerase is proposed to be an essential step in cancer cell immortalization and cancer progression. 3'-azido-2',3'-dideoxythymidine (AZT), a reverse transcriptase inhibitor, was reported to be incorporated in telomeric sequences of immortalized cells in culture and to suppress the activity of telomerase and the cell proliferation. In this study, after induction of cancer cell senescence with long-term treatment of AZT, we investigated the dynamics of telomerase subunits (hTERT, hTR, TEP), transcription factors (c-Myc, Mad1), telomerase activity, and finally, telomere length in a human breast cancer cell line. Materials and Methods: Human breast cancer cell (MDA-MB-231) was treated with AZT. Senescence was measured by senescence-associated β-gal staining and apoptosis was counted by dTd enzyme assay. Telomerase activity (by TRAP assay), expression of telomerase subunit genes (by RT-PCR and real-time PCR) and

telomere length (by Southern blot analysis) were measured after the AZT treatment.

Results: We found evidences of senescence, apoptosis and growth delay after AZT treatment. In addition, AZT- treated cancer cells showed inhibition of telomerase activity and shortening of telomere length in a dose- and duration-dependent way. Among the telomerase subunits, hTERT and c-Myc were the first factors to change after AZT treatment, subsequently, followed by the changes of hTR, Mad1 and TEP.

Conclusion: The suppression of hTERT and c-Myc by AZT treatment was the initial genetic phenomenon, subsequently followed by the changes of hTR, Mad1 and TEP.