I-BET151 (GSK1210151A) (GSK1210151A) is a specific BET inhibitor for BRD2/3/4 (IC50: 0.5/0.25/0.79 μM, in cell-free assays).

BRD2:0.5 μM, BRD3:0.25 μM, BRD4:0.79 μM

I-BET151 administration (30 mg/kg/day) in mice significantly inhibits the growth of MLL-AF9 murine and MLL-AF4 human leukemia tumors and notably prolongs lifespan.

When interacting with HL60 nuclear extracts, I-BET151 at concentrations of 0.5 or 5 μM inhibits the binding of BETs (BRD2/3/4/9) to acetylated histone peptides without affecting 23 other bromodomain proteins. This compound demonstrates potent inhibitory effects on cell lines with various MLL fusions, such as MV4;11, RS4;11, MOLM13, and NOMO1 cells (IC50: 15-192 nM). I-BET151 shows broad selectivity in inhibiting multiple protein types, including COX-2, P450, Aurora B, GSK3β, PI3K-γ, GPCR, ion channels, and transporters. Similar to I-BET762 (GSK525762A), I-BET151 possesses a high binding affinity for BRD2/3/4 (Kd: 0.02-0.1 μM) and significantly inhibits the production of the IL-6 cytokine in response to lipopolysaccharide stimulation in human peripheral blood mononuclear cells, whole blood, and rat WB (IC50: 0.16/1.26/1.26 μM).

Fluorescence anisotropy (FP) ligand displacement assay: All components are dissolved in buffer of composition 50 mM HEPES pH 7.4, 150 mM NaCl and 0.5 mM CHAPS with final concentrations of BRD 2/3/4 75 nM, fluorescent ligand 5 nM. 10 μL of this reaction mixture is added using a micro multidrop to wells containing 100 nL of various concentrations of I-BET151 or DMSO vehicle (1% final) in Greiner 384 well Black low volume microtitre plate and equilibrated in the dark for 60 minutes at room temperature. Fluorescence anisotropy is read in Envision (lex = 485 nm, lEM = 530 nm; Dichroic = 505 nM).

Cells are exposed to various concentrations of I-BET151 for 24 or 72 hours in 384-well or 96-well plates. For cell growth inhibition assays, plates are added with CellTiter-Glo reagent using a volume equivalent to the cell culture volume in the wells, shaken for approximately 2 minutes and chemiluminescent signal is read on the Analyst GT or EnVision Plate Reader. For cell proliferation assays, CellTiter-Aqueous One is added to each well and plates are incubated for 4 hours at 37 °C. Absorbance is read at 490 nm on a SpectraMax Gemini reader (Only for Reference)