Luciferase assay for modulation of the human telomerase RNA component promoter

Immortalisation is a hallmark of cancer commonly achieved by transcriptional reactivation of the telomerase reverse transcriptase and RNA component genes TERT and TERC, leading to inappropriate telomere maintenance. Multiple transcription factors modulate the telomerase gene promoters, orchestrated by the activities of a range of upstream signalling pathways. Previous studies have identified many of the pathways which individually contribute to activate or repress telomerase levels in cancer cells, resulting in a highly complex picture of telomerase regulation. The pathways of telomerase activation or suppression could be appropriate therapeutic targets in cancer or in telomeropathies such as dyskeratosis which are associated with loss of telomere stability. To identify small molecule regulators of TERC transcriptional regulation, activity of the TERC core gene promoter region was monitored using firefly luciferase. Reporter pGL3-TERC contains the TERC promoter cloned in the Promega vector PGL3-basic. Telomerase positive A2780 ovarian carcinoma cells are transiently transfected with the reporter construct and subsequently treated with small molecules to detect up- or down-regulation of luciferase activity. Derivatives of compound C430-0073 identified in assay TELO-01 were tested