siRNA inhibition of telomerase enhances the anti-cancer effect of doxorubicin in breast cancer cells.
BACKGROUND
Doxorubicin is an effective breast cancer drug but is hampered by a severe, dose-dependent toxicity. Concomitant administration of doxorubicin and another cancer drug may be able to sensitize tumor cells to the cytotoxicity of doxorubicin and lowers the therapeutic dosage. In this study, we examined the combined effect of low-dose doxorubicin and siRNA inhibition of telomerase on breast cancer cells. We found that when used individually, both treatments were rapid and potent apoptosis inducers; and when the two treatments were combined, we observed an enhanced and sustained apoptosis induction in breast cancer cells.


METHODS
siRNA targeting the mRNA of the protein component of telomerase, the telomerase reverse transcriptase (hTERT), was transfected into two breast cancer cell lines. The siRNA inhibition was confirmed by RT-PCR and western blot on hTERT mRNA and protein levels, respectively, and by measuring the activity level of telomerase using the TRAP assay. The effect of the hTERT siRNA on the tumorigenicity of the breast cancer cells was also studied in vivo by injection of the siRNA-transfected breast cancer cells into nude mice. The effects on cell viability, apoptosis and senescence of cells treated with hTERT siRNA, doxorubicin, and the combined treatment of doxorubicin and hTERT siRNA, were examined in vitro by MTT assay, FACS and SA-beta-galactosidase staining.


RESULTS
The hTERT siRNA effectively knocked down the mRNA and protein levels of hTERT, and reduced the telomerase activity to 30% of the untreated control. In vivo, the tumors induced by the hTERT siRNA-transfected cells were of reduced sizes, indicating that the hTERT siRNA also reduced the tumorigenic potential of the breast cancer cells. The siRNA treatment reduced cell viability by 50% in breast cancer cells within two days after transfection, while 0.5 microM doxorubicin treatment had a comparable effect but with a slower kinetics. The combination of hTERT siRNA and 0.5 microM doxorubicin killed twice as many cancer cells, showing a cumulative effect of the two treatments.


CONCLUSIONS
The study demonstrated the potential of telomerase inhibition as an effective treatment for breast cancer. When used in conjunction to doxorubicin, it could potentiate the cytotoxic effect of the drug to breast cancer cells.
The Molecular Medicine Center of Shaoxing People's Hospital, The First Affiliate Hospital of Shaoxing University, Shaoxing, PR China. dxj9666@163.com
Xuejun
X
Dong
Anding
A
Liu
Cindy
C
Zer
Jianguo
J
Feng
Zhuan
Z
Zhen
Mingfeng
M
Yang
Li
L
Zhong
D000818:Animals; D000970:Antineoplastic Agents / Q000008:administration & dosage*; D017209:Apoptosis / Q000187:drug effects; D001943:Breast Neoplasms / Q000188:drug therapy* / Q000201:enzymology* / Q000235:genetics / Q000628:therapy; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D003131:Combined Modality Therapy; D004317:Doxorubicin / Q000008:administration & dosage*; D005260:Female; D015316:Genetic Therapy; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D034741:RNA, Small Interfering / Q000235:genetics / Q000627:therapeutic use*; D019098:Telomerase / Q000235:genetics* / Q000378:metabolism
D000970:Antineoplastic Agents; D034741:RNA, Small Interfering; D004317:Doxorubicin; D019098:Telomerase