hTERT expression in response to telomerase inhibitor 1 (TI1) and cell viability of hTERT-expressing cells to cisplatin. (A) Telomerase activity following treatment with TI1. pLNCX and GSE24-2 cells were treated with TI1 (5 μM) for the indicated time periods (0, 3, and 7 days). Telomerase activity was detected with the TRAP assay kit. Different extract dilutions are presented for each TRAP assay (total protein amount indicated in triangles). Asterisks indicate the samples that showed important differences. The experiments were repeated 3 times, with similar results. (B) pLNCX and GSE24-2 cells were untreated (−) or treated (+) with c-DDP (3 μg/mL) for the indicated times (0, 3, and 7 days). Cells were harvested and mRNA levels for hTERT, hTR, and DKC were determined by RT-PCR. GAPDH mRNA was used as control. The experiments were repeated 3 times, with similar results. (C) pLNCX and GSE24-2 cells untreated (0) or treated with TI1 for the indicated times (0, 3, and 7 days) were harvested and mRNA levels for hTERT, hTR, and GAPDH were determined as in panel B. (D) 293T cells were transfected with pBABE or pBABE-flag-hTERT and stable cell lines. hTERT expression was detected by immunoblot using a specific antibody against the FLAG tag. (E) pBABE and pBABEhTERT cell lines were treated with increasing amounts of cisplatin. Survival was measured 72 hours after treatment. Data represent the means and standard deviations of 2 experiments performed in quadruplicate.