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. 2022 Sep 2;12(11):2684–2709. doi: 10.1158/2159-8290.CD-21-1307

Figure 1.

Figure 1. A potent, selective, and orally bioavailable inhibitor of the ENL/AF9 YEATS domains. A, Chemical structure for TDI-11055. B, Left, schematic depiction of the TR-FRET assay used to quantify the ability of a compound to disrupt YEATS domain and acylated histone H3 peptide interaction. Right, dose-dependent inhibition of the TR-FRET signal by TDI-11055. The IC50 shown represents the mean of four independent experiments. C, ITC experiment demonstrating direct binding of TDI-11055 to ENL YEATS with 1:1 stoichiometry. One of two independent replicates is shown. D, Top, immunoblots showing endogenous levels of ENL, GAS41, and YEATS2 protein after heat treatment in MOLM-13 leukemia cells at increasing temperatures. Bottom, quantification of immunoblot signals. One of two independent experiments is shown.E, Docking studies of TDI-11055 bound to ENL YEATS from cocrystal structures of SGC-iMLLT [Protein Data Bank (PDB): 6HT1] and H3K27ac (PDB: 5J9S). Ribbon representations indicate the protein, and stick representations show key residues involved in ENL and TDI-11055 interactions. TDI-11055 is depicted as sticks with colors corresponding to the atom type (green, carbon; blue, nitrogen; red, oxygen). F, Left, immunoblots showing levels of ectopically expressed FLAG-tagged wild-type (WT) or Y78A-mutant ENL proteins in HEK293 cells after heat treatment at increasing temperatures. Right, quantification of immunoblot signals. One of two independent experiments is shown. G, Rank-ordered heat map of FLAG-ENL ChIP-seq signals at ENL-bound peaks in MOLM-13 cells treated with DMSO or TDI-11055 (5 μmol/L) for 24 hours. H3K27ac and H3K9ac (GSE80779) ChIP-seq signals at ENL-bound peaks in untreated cells are also shown to demonstrate ENL localization to acetylated chromatin. See Supplementary Table S1. H, The genome browser view of indicated ChIP-seq signals at key ENL-bound genes (MYC, HOXA9/10) in parental [H3K27ac (gray), H3K9ac (black)] as well as DMSO (blue)- or TDI-11055 (red)–treated (FLAG-ENL) MOLM-13 cells. I, PK studies in mice (n = 3) performed for TDI-11055 and SGC-iMLLT, demonstrating unbound plasma concentration of indicated compounds after an oral dose of 30, 50, and 100 mg/kg.

A potent, selective, and orally bioavailable inhibitor of the ENL/AF9 YEATS domains. A, Chemical structure for TDI-11055. B, Left, schematic depiction of the TR-FRET assay used to quantify the ability of a compound to disrupt YEATS domain and acylated histone H3 peptide interaction. Right, dose-dependent inhibition of the TR-FRET signal by TDI-11055. The IC50 shown represents the mean of four independent experiments. C, ITC experiment demonstrating direct binding of TDI-11055 to ENL YEATS with 1:1 stoichiometry. One of two independent replicates is shown. D, Top, immunoblots showing endogenous levels of ENL, GAS41, and YEATS2 protein after heat treatment in MOLM-13 leukemia cells at increasing temperatures. Bottom, quantification of immunoblot signals. One of two independent experiments is shown.E, Docking studies of TDI-11055 bound to ENL YEATS from cocrystal structures of SGC-iMLLT [Protein Data Bank (PDB): 6HT1] and H3K27ac (PDB: 5J9S). Ribbon representations indicate the protein, and stick representations show key residues involved in ENL and TDI-11055 interactions. TDI-11055 is depicted as sticks with colors corresponding to the atom type (green, carbon; blue, nitrogen; red, oxygen). F, Left, immunoblots showing levels of ectopically expressed FLAG-tagged wild-type (WT) or Y78A-mutant ENL proteins in HEK293 cells after heat treatment at increasing temperatures. Right, quantification of immunoblot signals. One of two independent experiments is shown. G, Rank-ordered heat map of FLAG-ENL ChIP-seq signals at ENL-bound peaks in MOLM-13 cells treated with DMSO or TDI-11055 (5 μmol/L) for 24 hours. H3K27ac and H3K9ac (GSE80779) ChIP-seq signals at ENL-bound peaks in untreated cells are also shown to demonstrate ENL localization to acetylated chromatin. See Supplementary Table S1. H, The genome browser view of indicated ChIP-seq signals at key ENL-bound genes (MYC, HOXA9/10) in parental [H3K27ac (gray), H3K9ac (black)] as well as DMSO (blue)- or TDI-11055 (red)–treated (FLAG-ENL) MOLM-13 cells. I, PK studies in mice (n = 3) performed for TDI-11055 and SGC-iMLLT, demonstrating unbound plasma concentration of indicated compounds after an oral dose of 30, 50, and 100 mg/kg.