LAUSR

HEK293 cells stably expressing OATP1B1 or OATP1B3 or HepG2 cells were plated into 6-well plates at 3 X 105 cells/well. After overnight incubation, the cells were transfected with 0.5ug – 2ug Simplicon/well in the presence of 200ng/ml B18R IFN binding protein. The following day, cells were removed, counted and plated into 96-well plates at 1 X 105 cells/well in 200ul DMEM/F12 growth media containing 0.1ug/ml puromycin and 200ng/ml – 600ng/ml B18R protein. The next day, the cells were treated with substrate compounds (10uM, with the exception of phenacetin which was 40uM) for 2-3 hours in serum-free medium and assayed for metabolite production by mass spectrophotometry. Cells that were incubated for 144 hours were passaged and scaled into the same growth media supplemented with 0.1ug/ml puromycin and 200ng/ml B18R IFN binding protein. Introduction