Discovery and Optimization of Small-Molecule Ligands for the CBP/p300 Bromodomains

Abstract

Small-molecule inhibitors that target bromodomains outside of the bromodomain and extra-terminal (BET) sub-family are lacking. Here, we describe highly potent and selective ligands for the bromodomain module of the human lysine acetyl transferase CBP/p300, developed from a series of 5-isoxazolyl-benzimidazoles. Our starting point was a fragment hit, which was optimized into a more potent and selective lead using parallel synthesis employing Suzuki couplings, benzimidazole-forming reactions, and reductive aminations. The selectivity of the lead compound against other bromodomain family members was investigated using a thermal stability assay, which revealed some inhibition of the structurally related BET family members. To address the BET selectivity issue, X-ray crystal structures of the lead compound bound to the CREB binding protein (CBP) and the first bromodomain of BRD4 (BRD4(1)) were used to guide the design of more selective compounds. The crystal structures obtained revealed two distinct binding modes. By varying the aryl substitution pattern and developing conformationally constrained analogues, selectivity for CBP over BRD4(1) was increased. The optimized compound is highly potent (Kd = 21 nM) and selective, displaying 40-fold selectivity over BRD4(1). Cellular activity was demonstrated using fluorescence recovery after photo-bleaching (FRAP) and a p53 reporter assay. The optimized compounds are cell-active and have nanomolar affinity for CBP/p300; therefore, they should be useful in studies investigating the biological roles of CBP and p300 and to validate the CBP and p300 bromodomains as therapeutic targets.

CBP30, a selective CBP/p300 bromodomain inhibitor, suppresses human Th17 responses

“…ITC data revealed K D s of 26 nM and 32 nM for CBP and p300, respectively (Fig. 1C), in good agreement with affinity data already reported (15). Within the BET family, the first bromodomain of BRD4 [BRD4 (1)] bound with highest affinity (K D of 885 nM), thus with 34-fold reduced selectivity compared with CBP (Table S2).…”

“…1A). Initial screening against 10 bromodomains suggested selectivity for CBP/ p300 bromodomains (15). Here, we evaluated the selectivity of this inhibitor using temperature shift assays against a total of 45 bromodomains (Fig.…”

“…A number of weak inhibitors, such as ischemin (K D = 25 μM), have been reported to target the CBP/p300 bromodomain (14). The synthesis and binding modes of more potent CBP and BET/CBP inhibitors have been reported in the last year (15,16). The most potent of these inhibitors, CBP30, has recently been shown to bind with low nanomolar affinity to the bromodomains of CBP/p300 (15).…”

“…The synthesis and binding modes of more potent CBP and BET/CBP inhibitors have been reported in the last year (15,16). The most potent of these inhibitors, CBP30, has recently been shown to bind with low nanomolar affinity to the bromodomains of CBP/p300 (15). However, this initial study did not examine the broader specificity of CBP30 for other bromodomain targets or look at the ability of CBP30 to inhibit physiological cellular processes.…”

The 2016 WHO Classification of Tumours of the Urinary System and Male Genital Organs—Part B: Prostate and Bladder Tumours

“…The most frequently altered pathways in bladder cancer include the PI3K/AKT/mammalian target of rapamycin pathway [96,[119][120][121], the FGFR3/RAF/RAS pathway, the TP53/RB1 pathway, immune response checkpoint modulators [122,123], and chromatin-regulating and -remodelling genes [124][125][126]. In general, mutations along a given pathway are mutually exclusive.…”