Structure- and Property-Based Optimization of Efficient Pan-Bromodomain and Extra Terminal Inhibitors to Identify Oral and Intravenous Candidate I-BET787.

The bromodomain and extra terminal (BET) family of bromodomain-containing proteins are important epigenetic regulators that elicit their effect through binding histone tail N-acetyl lysine (KAc) post-translational modifications. Recognition of such markers has been implicated in a range of oncology and immune diseases and, as such, small-molecule inhibition of the BET family bromodomain-KAc protein-protein interaction has received significant interest as a therapeutic strategy, with several potential medicines under clinical evaluation. This work describes the structure- and property-based optimization of a ligand and lipophilic efficient pan-BET bromodomain inhibitor series to deliver candidate I-BET787 (70) that demonstrates efficacy in a mouse model of inflammation and suitable properties for both oral and intravenous (IV) administration. This focused two-phase explore-exploit medicinal chemistry effort delivered the candidate molecule in 3 months with less than 100 final compounds synthesized.

Structure-Guided Design and Optimization of Covalent VHL-Targeted Sulfonyl Fluoride PROTACs.

Proteolysis-targeting chimeras (PROTACs) are heterobifunctional molecules that have emerged as a therapeutic modality to induce targeted protein degradation (TPD) by harnessing cellular proteolytic degradation machinery. PROTACs which ligand the E3 ligase in a covalent manner have attracted intense interest; however, covalent PROTACs with a broad protein of interest (POI) scope have proven challenging to discover by design. Here, we report the structure-guided design and optimization of Von Hippel-Lindau (VHL) protein-targeted sulfonyl fluorides which covalently bind Ser110 in the HIF1α binding site. We demonstrate that their incorporation in bifunctional degraders induces targeted protein degradation of BRD4 or the androgen receptor without further linker optimization. Our study discloses the first covalent VHL ligands which can be implemented directly in bifunctional degrader design, expanding the substrate scope of covalent E3 ligase PROTACs.

Structure-Guided Design of a Domain-Selective Bromodomain and Extra Terminal N-Terminal Bromodomain Chemical Probe.

Small-molecule-mediated disruption of the protein-protein interactions between acetylated histone tails and the tandem bromodomains of the bromodomain and extra-terminal (BET) family of proteins is an important mechanism of action for the potential modulation of immuno-inflammatory and oncology disease. High-quality chemical probes have proven invaluable in elucidating profound BET bromodomain biology, with seminal publications of both pan- and domain-selective BET family bromodomain inhibitors enabling academic and industrial research. To enrich the toolbox of structurally differentiated N-terminal bromodomain (BD1) BET family chemical probes, this work describes an analysis of the GSK BRD4 bromodomain data set through a lipophilic efficiency lens, which enabled identification of a BD1 domain-biased benzimidazole series. Structure-guided growth targeting a key Asp/His BD1/BD2 switch enabled delivery of GSK023, a high-quality chemical probe with 300-1000-fold BET BD1 domain selectivity and a phenotypic cellular fingerprint consistent with BET bromodomain inhibition.

Identification and Optimization of a Ligand-Efficient Benzoazepinone Bromodomain and Extra Terminal (BET) Family Acetyl-Lysine Mimetic into the Oral Candidate Quality Molecule I-BET432.

The bromodomain and extra terminal (BET) family of proteins are an integral part of human epigenome regulation, the dysregulation of which is implicated in multiple oncology and inflammatory diseases. Disrupting the BET family bromodomain acetyl-lysine (KAc) histone protein-protein interaction with small-molecule KAc mimetics has proven to be a disease-relevant mechanism of action, and multiple molecules are currently undergoing oncology clinical trials. This work describes an efficiency analysis of published GSK pan-BET bromodomain inhibitors, which drove a strategic choice to focus on the identification of a ligand-efficient KAc mimetic with the hypothesis that lipophilic efficiency could be drastically improved during optimization. This focus drove the discovery of the highly ligand-efficient and structurally distinct benzoazepinone KAc mimetic. Following crystallography to identify suitable growth vectors, the benzoazepinone core was optimized through an explore-exploit structure-activity relationship (SAR) approach while carefully monitoring lipophilic efficiency to deliver I-BET432 (41) as an oral candidate quality molecule.

Investigation of Janus Kinase (JAK) Inhibitors for Lung Delivery and the Importance of Aldehyde Oxidase Metabolism.

The Janus family of tyrosine kinases (JAK1, JAK2, JAK3, and TYK2) play an essential role in the receptor signaling of cytokines that have been implicated in the pathogenesis of severe asthma, and there is emerging interest in the development of small-molecule-inhaled JAK inhibitors as treatments. Here, we describe the optimization of a quinazoline series of JAK inhibitors and the results of mouse lung pharmacokinetic (PK) studies where only low concentrations of parent compound were observed. Subsequent investigations revealed that the low exposure was due to metabolism by aldehyde oxidase (AO), so we sought to identify quinazolines that were not metabolized by AO. We found that specific substituents at the quinazoline 2-position prevented AO metabolism and this was rationalized through computational docking studies in the AO binding site, but they compromised kinome selectivity. Results presented here highlight that AO metabolism is a potential issue in the lung.

Hi-JAK-ing the ubiquitin system: The design and physicochemical optimisation of JAK PROTACs.

PROTACs have recently emerged as a novel paradigm in drug discovery. They can hijack existing biological machinery to selectively degrade proteins of interest, in a catalytic fashion. Here we describe the design, optimisation and biological activity of a set of novel PROTACs targeting the Janus kinase family (JAK1, JAK2, JAK3 and TYK2) of proximal membrane-bound proteins. The JAK family proteins display membrane localisation by virtue of their association with cytoplasmic tails of cytokine receptors, and there are no reports of a successful PROTAC strategy being deployed against this class of proteins. JAK PROTACs from two distinct JAK chemotypes were designed, optimising the physicochemical properties for each template to enhance cell permeation. These PROTACs are capable of inducing JAK1 and JAK2 degradation, demonstrating an extension of the PROTAC methodology to an unprecedented class of protein targets. A number of known ligase binders were explored, and it was found that PROTACs bearing an inhibitor of apoptosis protein (IAP) ligand induced significantly more JAK degradation over Von Hippel-Lindau (VHL) and Cereblon (CRBN) PROTACs. In addition, the mechanism of action of the JAK PROTACs was elucidated, and it was confirmed that JAK degradation was both IAP- and proteasome-dependent.

Discovery of a Bromodomain and Extraterminal Inhibitor with a Low Predicted Human Dose through Synergistic Use of Encoded Library Technology and Fragment Screening.

The bromodomain and extraterminal (BET) family of bromodomain-containing proteins are important regulators of the epigenome through their ability to recognize N-acetyl lysine (KAc) post-translational modifications on histone tails. These interactions have been implicated in various disease states and, consequently, disruption of BET-KAc binding has emerged as an attractive therapeutic strategy with a number of small molecule inhibitors now under investigation in the clinic. However, until the utility of these advanced candidates is fully assessed by these trials, there remains scope for the discovery of inhibitors from new chemotypes with alternative physicochemical, pharmacokinetic, and pharmacodynamic profiles. Herein, we describe the discovery of a candidate-quality dimethylpyridone benzimidazole compound which originated from the hybridization of a dimethylphenol benzimidazole series, identified using encoded library technology, with an N-methyl pyridone series identified through fragment screening. Optimization via structure- and property-based design led to I-BET469, which possesses favorable oral pharmacokinetic properties, displays activity in vivo, and is projected to have a low human efficacious dose.