Endocrine Abstracts

Objective: The first orphan nuclear receptors discovered were Estrogen Related Receptors (ERRs), which were recognized due to their sequence closeness to the classical estrogen receptors. Countless impartial studies have linked ERRγ to the onset and progression of several malignancies, including thyroid cancer. Moreover, it is recognized that ERRγ has a unique function in controlling metabolic effectors, cell cycle progression, and apoptosis in cancers of different kinds. We assessed the potential of Estrogen Related Receptor Gamma (ERRγ) as a therapeutic target by analyzing its expression levels in different cell lines of thyroid cancer. Furthermore, the anti-tumor activity of DN200434 (an inverse agonist of ERRγ) in combination with MK2206 (an Akt inhibitor), was investigated.

Methods: ATC cells (HTh7, 8505c) and PTC cells (KTC-1, TPC-1) were cultivated in their corresponding medium. CCK-assay was used to measure the IC50 value of DN200434 and MK2206 in ATC/PTC cells after 24 hours. Cells were treated with drugs alone or in combination for 24h to assess their inhibitory effect on the viability, apoptosis and cell cycle using flow cytometry. Additionally, western blot was used to enumerate the proteins level implicated in the cell cycle and apoptotic pathways. Following combination treatment, the colony formation assay was carried out in six-well plates with 500 cells per well. Crystal violet staining was followed by a count of large colonies with up to 50 cells.

Results: We observed that the IC50 of DN200434 and MK2206 was 30-45 μM and 10-20 μM in ATC/PTC cells at 24h, respectively. The combination of DN200434 and MK2206 in ATC/PTC cells synergistically enhanced cell cytotoxicity up to 70-75% as compared to DN200434 (5-15%) and MK2206 (10-20%) alone. Flow cytometry results showed significant apoptosis and cell cycle arrest at G0/G1 phase in the combination drugs treated cells respectively. Western blot data confirmed significant apoptosis in combination treated cells with increased level of cleaved caspase-3 and cleaved-PARP. A significant reduction (70-80%) in the colony forming ability was observed in combination treated cells.

Conclusion: Our findings show that DN200434 and MK2206 together can be employed as a treatment combination to treat advanced papillary thyroid cancer and anaplastic thyroid cancer. Before this powerful combination is put through clinical trials, more assessment of combination therapy in animal models or 3D spheroid models is needed.