Discovery of CREBBP Bromodomain Inhibitors by High-Throughput Docking and Hit Optimization Guided by Molecular Dynamics

Abstract

We have identified two chemotypes of CREBBP bromodomain ligands by fragment-based high-throughput docking. Only 17 molecules from the original library of two-million compounds were tested in vitro. Optimization of the two low-micromolar hits, the 4-acylpyrrole 1 and acylbenzene 9, was driven by molecular dynamics results which suggested improvement of the polar interactions with the Arg1173 side chain at the rim of the binding site. The synthesis of only two derivatives of 1 yielded the 4-acylpyrrole 6 which shows a single-digit micromolar affinity for the CREBBP bromodomain and a ligand efficiency of 0.34 kcal/mol per non-hydrogen atom. Optimization of the acylbenzene hit 9 resulted in a series of derivatives with nanomolar potencies, good ligand efficiency and selectivity (see Unzue, A.; Xu, M.; Dong, J.; Wiedmer, L.; Spiliotopoulos, D.; Caflisch, A.; Nevado, C.Fragment-Based Design of Selective Nanomolar Ligands of the CREBBP Bromodomain. J. Med. Chem. 2015, DOI: 10.1021/acs.jmedchem.5b00172). The in silico predicted binding mode of the acylbenzene derivative 10 was validated by solving the structure of the complex with the CREBBP bromodomain.

Using Fragment Based Drug Discovery to Target Epigenetic Regulators in Cancer

“…Some research groups directly dock the available online fragment libraries onto protein structures [40,45] while others, as we will describe in detail below, use a fragment-based in silico procedure called anchor-based library tailoring (ALTA) to select subsets of compounds from online compound library such as ZINC library for high throughput docking [39,41,42]. For the ALTA procedure, the molecules from online library are decomposed into fragments, screened against target proteins, and the fragment hits will serve as anchors to retrieve molecules from the original library for another run of high throughput docking [39,41,42]. The advantage of the ALTA procedure is that the rigid fragments usually dock more accurately than compounds with many rotatable bonds, and the computational cost can be reduced from the smaller subset of retrieved molecules for high throughput docking [41].…”

“…Four out of the 24 purchased hits are confirmed by positive Tm shifts in TSA assay and the inhibition of AcK histone peptide binding to BRD4(I) in an AlphaScreen ® assay [39]. This research group also used a similar ALTA strategy to perform an in silico screen against the bromodomain of CREBBP and 17 hits were purchased for verification [42]. Two out of the 17 selected hits inhibited the interaction between AcK peptide and CREBBP bromodomain at low µM range.…”

“…In silico screens were successfully deployed by several groups to target the bromodomains of BRD4 and CREBBP [39,[41][42][43][44][45]54]. Zhao et al [39] at the University of Zurich (Switzerland) used a fragment-based in silico method called Anchor-based Library Tailoring (ALTA) to select compounds from the large (~9 million molecules) ZINC all-now library (version of 2012) for high throughput docking [39].…”

“…Two out of the 17 selected hits inhibited the interaction between AcK peptide and CREBBP bromodomain at low µM range. Guided by extensive MD simulations and high resolution X-ray crystallography, they optimized both active fragment hits into nanomolar inhibitors of CREBBP bromodomain [42,58].…”

Discovery of New Bromodomain Scaffolds by Biosensor Fragment Screening

“…1,2 Currently, relatively selective and potent inhibitors have been identified for 17 of the 46 bromodomain proteins encoded by the human genome. Since the first bromodomain inhibitor was discovered by phenotypic screening, 3 a variety of methods have been successfully employed to discover bromodomain inhibitors including phenotypic screening, 4 analogue-based design, 5 structure-based screening, 6 focused-library screening, 7 and fragment screening. 8 Surface plasmon resonance (SPR) has become a primary fragment screening method in recent years.…”