Discovery of electrophilic degraders that exploit SAr chemistry.

Targeted covalent inhibition (TCI) and targeted protein degradation (TPD) have proven effective in pharmacologically addressing formerly 'undruggable' targets. Integration of both methodologies has resulted in the development of electrophilic degraders where recruitment of a suitable E3 ubiquitin ligase is achieved through formation of a covalent bond with a cysteine nucleophile. Expanding the scope of electrophilic degraders requires the development of electrophiles with tempered reactivity that enable selective ligase recruitment and reduce cross-reactivity with other cellular nucleophiles.

Relocalizing transcriptional kinases to activate apoptosis.

Kinases are critical regulators of cellular function that are commonly implicated in the mechanisms underlying disease. Most drugs that target kinases are molecules that inhibit their catalytic activity, but here we used chemically induced proximity to convert kinase inhibitors into activators of therapeutic genes. We synthesized bivalent molecules that link ligands of the transcription factor B cell lymphoma 6 (BCL6) to inhibitors of cyclin-dependent kinases (CDKs).

Borrowing Transcriptional Kinases to Activate Apoptosis.

Protein kinases are disease drivers whose therapeutic targeting traditionally centers on inhibition of enzymatic activity. Here chemically induced proximity is leveraged to convert kinase inhibitors into context-specific activators of therapeutic genes. Bivalent molecules that link ligands of the transcription factor B-cell lymphoma 6 (BCL6) to ATP-competitive inhibitors of cyclin-dependent kinases (CDKs) were developed to re-localize CDK to BCL6-bound loci on chromatin and direct phosphorylation of RNA Pol II.

Proteomics-Based Discovery of First-in-Class Chemical Probes for Programmed Cell Death Protein 2 (PDCD2).

Chemical probes are essential tools for understanding biological systems and for credentialing potential biomedical targets. Programmed cell death 2 (PDCD2) is a member of the B-cell lymphoma 2 (Bcl-2) family of proteins, which are critical regulators of apoptosis. Here we report the discovery and characterization of 10 e, a first-in-class small molecule degrader of PDCD2.

Total Synthesis and Target Identification of the Curcusone Diterpenes.

The curcusone natural products are complex diterpenes featuring a characteristic [6-7-5] tricyclic carbon skeleton similar to the and diterpenes. Among them, curcusones A-D demonstrated potent anticancer activity against a broad spectrum of human cancer cell lines. Prior to this study, no total synthesis of the curcusones was achieved and their anticancer mode of action remained unknown.

Chemoproteomics-Enabled De Novo Discovery of Photoswitchable Carboxylesterase Inhibitors for Optically Controlled Drug Metabolism.

Herein, we report arylazopyrazole ureas and sulfones as a novel class of photoswitchable serine hydrolase inhibitors and present a chemoproteomic platform for rapid discovery of optically controlled serine hydrolase targets in complex proteomes. Specifically, we identify highly potent and selective photoswitchable inhibitors of the drug-metabolizing enzymes carboxylesterases 1 and 2 and demonstrate their pharmacological application by optically controlling the metabolism of the immunosuppressant drug mycophenolate mofetil. Collectively, this proof-of-concept study provides a first example of photopharmacological tools to optically control drug metabolism by modulating the activity of a metabolizing enzyme.

Combined Omics Approach Identifies Gambogic Acid and Related Xanthones as Covalent Inhibitors of the Serine Palmitoyltransferase Complex.

In this study, we identify the natural product gambogic acid as well as structurally related synthetic xanthones as first-in-class covalent inhibitors of the de novo sphingolipid biosynthesis. We apply chemoproteomics to determine that gambogic acid binds to the regulatory small subunit B of the serine palmitoyltransferase complex (SPTSSB). We then test structurally related synthetic xanthones to identify 18 as an equally potent but more selective binder of SPTSSB and show that 18 reduces sphingolipid levels in situ and in vivo.

Ethynylation of Cysteine Residues: From Peptides to Proteins in Vitro and in Living Cells.

Current approaches to introduce terminal alkynes for bioorthogonal reactions into biomolecules still present limitations in terms of either reactivity, selectivity, or adduct stability. We present a method for the ethynylation of cysteine residues based on the use of ethynylbenziodoxolone (EBX) reagents. The acetylene group is directly introduced onto the thiol group of cysteine and can be used for copper-catalyzed alkyne-azide cycloaddition (CuAAC) without further processing.

Discovery and Evaluation of New Activity-Based Probes for Serine Hydrolases.

Serine hydrolases play crucial biological roles and are important therapeutic targets in many clinical applications. Activity-based protein profiling of serine hydrolases by using fluorophosphonate probes, pioneered by Cravatt and co-workers, has been a powerful tool for interrogating serine hydrolases in various biological systems. Herein, we present new phenyl phosphonate probes with an azide handle for click chemistry that offer remarkable improvements over the classical fluorophosphonate serine hydrolase activity-based probes including ease of preparation, excellent cell permeability, and distinct reactivity profiles, as controlled by the phenolate leaving group.

The Hairpin Form of r(GC) in c9ALS/FTD Is Repeat-Associated Non-ATG Translated and a Target for Bioactive Small Molecules.

The most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) is an expanded GC repeat [(GC)] in C9ORF72. ALS/FTD-associated toxicity has been traced to the RNA transcribed from the repeat expansion [r(GC)], which sequesters RNA-binding proteins (RBPs) and undergoes repeat-associated non-ATG (RAN) translation to generate toxic dipeptide repeats. Using in vitro and cell-based assays, we identified a small molecule (4) that selectively bound r(GC), prevented sequestration of an RBP, and inhibited RAN translation.

Drugging the RNA World.

Although we live in the remnants of an RNA world, the world of drug discovery and chemical probes is firmly protein-centric. Developing highly selective small molecules targeting RNA is often considered to be an insurmountable challenge. Our goal is to demystify the design of such compounds.

Ruthenium anticancer agent KP1019 binds more tightly than NAMI-A to tRNA.

The ruthenium-based anticancer agent NAMI-A (ImH[trans-RuCl(dmso)(Im)], where Im = imidazole) has been shown to interact with RNA in vivo and in vitro. We hypothesized that the similarly structured drug KP1019 (IndH[trans-RuCl(Ind)], where Ind = indazole) binds to RNA as well. Fluorescence spectroscopy was employed to assay the interactions between either NAMI-A or KP1019 and tRNA through an intrinsic fluorophore wybutosine (Y) base and by extrinsic displacement of the intercalating agent ethidium bromide.