Down regulation of human telomerase reverse transcriptase (hTERT) expression by BIBR1532 in human glioblastoma LN18 cells
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Increased telomerase activity can be blocked by targeting the hTERT activity at both RNA and catalytic subunits. Various inhibitors had been used to regulate hTERT activity in glioblastoma cell lines and showed promising results. The present study hypothesized that the telomerase specific inhibitor BIBR1532 can effectively down-regulate the telomerase activity in LN18 glioblastoma cell line. LN18 glioblastoma cell line was treated with various concentrations of BIBR1532 at different time intervals. MTT assay was performed to determine cell viability after BIBR1532 treatment. hTERT mRNA and protein expression were determined by qRT-PCR and western blotting, respectively. Flow cytometry and TRAP assay was performed to detect the rate of apoptosis and telomerase activity in treated and control samples. One-way ANOVA was performed to compare the mean values of variables in control and BIBR1532 treated groups. LN18 cells showed a significant dose dependent cytotoxic effect after treatment with BIBR1532. hTERT mRNA expression in cells treated with 25, 100 and 200 μM BIBR1532 treated groups was decreased ~ 21, ~ 61.2, and ~ 77%, respectively (p < 0.05). We also observed that, BIBR1532 treatment reduced the expression of hTERT protein in LN18 cells in a dose dependent manner. The Flow cytometry data showed that, the drug induced significant increase in the total percentage of apoptotic cells with 200 μM concentration of BIBR1532 at all time points. BIBR1532 exhibited potent inhibition of telomerase activity in a dose-dependent manner in LN18 cells. BIBR1532 could induce apoptosis in LN18 cells through the downregulation of telomerase activity at transcriptional and translational level. We conclude that BIBR1532 may be a therapeutic agent to suppress telomerase activity, however, further efforts are necessary in order to explore this therapeutic strategy.
KeywordshTERT BIBR1532 Glioblastoma Regulation
Lavanya C was supported by a UGC RGNF-Senior research fellowship (University Grants Commission-Rajiv Gandhi National Fellowship), New Delhi, India. The study was financially supported by DST-SERB (Department of Science & Technology-Science and Engineering research Board), Government of India. We thank Dr. Vijay Kumar Kalia for providing chemicals and reagents.
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Conflict of interest
The authors declare that there are no conflicts of interest.
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