Small molecules targeting the MBM1 site efficiently disrupt bivalent menin-MLL interaction. (A) Fluorescence polarization experiments demonstrating displacement of MLL_4-43 from menin by MI-2 and MI-2-2. Data represent mean values from 2 experiments ± SD. (B) Coimmunoprecipitation experiment in HEK293 cells transfected with Flag-MLL-AF9 showing that MI-2 and MI-2-2 disrupt the interaction of menin with MLL-AF9 in human cells. Input shows the levels of menin and MLL. The amount of menin bound to Flag-MLL-AF9 was detected by coimmunoprecipitation using anti-Flag Ab followed by immunoblotting using menin Ab. (C) Model of the disruption of bivalent MLL-menin interaction by MI-2-2 via binding to MBM1 site in menin.