Establishment of high-throughput drug screening platform for sodium/iodide symporter (NIS) enhancer using dual reporter gene system

Objectives Anaplastic thyroid cancer (ATC), is the most aggressive and resistant cancer to the radioiodine treatment, which leads to poorest prognosis among thyroid cancers. Although ATC has insufficient NIS expression, enhancement of NIS expression is able to make the ATC susceptible to radioiodine therapy. The aim of this study was to establish high-throughput drug screening (HTS) platform for sodium/iodide symporter (NIS) enhancer using dual reporter gene system using dual reporter gene system.

Methods 8505C cells (ATC) were transfected with a plasmid vector having NIS promoter driven one reporter gene and CMV promoter driven the other reporter gene system (pNIS-FL2-TurboFP635-pCMV-Rluc plasmid). Stably transfected cells were selected with geneticin (8505C-pNIS-pCMV cells) and expression of the reporter system was confirmed by FACS, bioluminescence and fluorescent imaging. 8505C-pNIS-pCMV cells were used as a screening platform cells for detecting strong NIS enhancers among thousands of tyrosine kinase inhibitor (TKI) candidates (0.025mM ± 0.28). RT-PCR and Western blotting of NIS and radioactive I-125 uptake assay was performed with 8505C cells before and after exposure to several screened TKI candidates.

Results 8505C-pNIS-pCMV cell line having the dual reporter gene system was stably established. The luciferase activities (F-luc and R-luc), TurboFP635 expression were confirmed by bioluminescent and fluorescent imaging, respectively. In vitro BLI signals of F-luc (R²=0.9245, p<0.0001) and R-luc (R²=0.9820, p<0.05) increased by increase of cell number and FACS analysis confirmed stable expression of TurboFP635. Four Chemicals which enhance NIS promoter activity more than 4 fold were selected using the HTS platform and the selected chemicals increased mRNA and protein of NIS and also increased radioactive I-125 uptake significantly.

Conclusions We successfully developed high-throughput NIS enhancer screening platform using optical dual reporter gene system and screened TKI chemicals which enhance NIS expression among numerous candidates. The chemicals can be applied to restore iodine avidity to the ATC and the modified ATC can be successfully treated with I-131 therapy.