To identify novel small molecules against the TSH receptor, we developed a sensitive transcription-based luciferase high-throughput screening (HTS) system named the TSHR-Glo Assay (TSHR-Glo).
This assay uses double-transfected Chinese hamster ovary cells stably expressing the human TSHR and a cAMP-response element (CRE) construct fused to an improved luciferase reporter gene.
The assay was highly responsive toward TSH in a dose-dependent manner with a TSH sensitivity of 10−10M (10 ± 1.12 μU/ml) and thyroid-stimulating antibodies, a hallmark of Graves’ disease, could also be detected. The assay was validated against the standard indicator of HTS performance – the
Using this approach, we were able to identify specific agonists against the TSH receptor leading to the characterization of several TSH agonist molecules. Hence, the TSHR-Glo assay was a one-step cell-based HTS assay, which was successful in the discovery of novel small molecular agonists and for the detection of stimulating antibodies to the TSH receptor.