Fig. 3

Identification of IRP2 inhibitors that dispersed the interaction between IRP2 and IREs. (A) Schematic diagram of the workflow implemented to identify small molecule inhibitors of IRP2. (B) Structures of KS-20073 and KS-20226. (C) Surface plasmon resonance (SPR) analysis revealed the inhibition of IRP2 binding to IRE by KS-20073 and KS-20226 in a dose-dependent manner. (D) Binding of IRP2 to biotin-FTH IRE was assessed using the RNA immunoprecipitation (IP) assay. HEK-293T cells were transfected with pCMV6-Myc (empty) and the pCMV6-Myc-IRP2 plasmid and were treated with KS-20073 and KS-20226 (10 µM) for 24 h. mRNA enrichment was achieved using qRT-PCR and normalized with input total RNA. Data are represented as the mean ± SEM (n = 4). **p < 0.01. (E) The RNA IP assay was used to assess the effect of endogenous IRP2 on IRP2 inhibitors. SW480 cells were treated with KS-20073 and KS-20226 (10 µM) for 24 h. Data are represented as the mean ± SEM (n = 4). ***p < 0.001