Discovery of small molecule guanylyl cyclase A receptor positive allosteric modulators.

The particulate guanylyl cyclase A receptor (GC-A), via activation by its endogenous ligands atrial natriuretic peptide (ANP) and b-type natriuretic peptide (BNP), possesses beneficial biological properties such as blood pressure regulation, natriuresis, suppression of adverse remodeling, inhibition of the renin-angiotensin-aldosterone system, and favorable metabolic actions through the generation of its second messenger cyclic guanosine monophosphate (cGMP). Thus, the GC-A represents an important molecular therapeutic target for cardiovascular disease and its associated risk factors. However, a small molecule that is orally bioavailable and directly targets the GC-A to potentiate cGMP has yet to be discovered.

Discovery of 1-[2-(1-methyl-1H-pyrazol-5-yl)-[1,2,4]triazolo[1,5-a]pyridin-6-yl]-3-(pyridin-4-ylmethyl)urea as a potent NAMPT (nicotinamide phosphoribosyltransferase) activator with attenuated CYP inhibition.

Nicotinamide phosphoribosyltransferase (NAMPT) catalyzes the rate-limiting step of the NAD salvage pathway. Since NAD plays a pivotal role in many biological processes including metabolism and aging, activation of NAMPT is an attractive therapeutic target for treatment of diverse array of diseases. Herein, we report the continued optimization of novel urea-containing derivatives which were identified as potent NAMPT activators.

Optimization of a urea-containing series of nicotinamide phosphoribosyltransferase (NAMPT) activators.

NAD is a crucial cellular factor that plays multifaceted roles in wide ranging biological processes. Low levels of NAD have been linked to numerous diseases including metabolic disorders, cardiovascular disease, neurodegeneration, and muscle wasting disorders. A novel strategy to boost NAD is to activate nicotinamide phosphoribosyltransferase (NAMPT), the putative rate-limiting step in the NAD salvage pathway.

Discovery of small molecule antagonists of chemokine receptor CXCR6 that arrest tumor growth in SK-HEP-1 mouse xenografts as a model of hepatocellular carcinoma.

The chemokine system plays an important role in mediating a proinflammatory microenvironment for tumor growth in hepatocellular carcinoma (HCC). The CXCR6 receptor and its natural ligand CXCL16 are expressed at high levels in HCC cell lines and tumor tissues and receptor expression correlates with increased neutrophils in these tissues contributing to poor prognosis in patients. Availability of pharmacologcal tools targeting the CXCR6/CXCL16 axis are needed to elucidate the mechanism whereby neutrophils are affected in the tumor environment.

Boosting NAD with a small molecule that activates NAMPT.

Pharmacological strategies that boost intracellular NAD are highly coveted for their therapeutic potential. One approach is activation of nicotinamide phosphoribosyltransferase (NAMPT) to increase production of nicotinamide mononucleotide (NMN), the predominant NAD precursor in mammalian cells. A high-throughput screen for NAMPT activators and hit-to-lead campaign yielded SBI-797812, a compound that is structurally similar to active-site directed NAMPT inhibitors and blocks binding of these inhibitors to NAMPT.

Measurement of Pyridine Nucleotides in Biological Samples Using LC-MS/MS.

Pyridine nucleotides which include NAD, NADH, NADP, and NADPH play vital roles in many different biological processes. These metabolites can be accurately quantified in a wide variety of biological samples using LC-MS/MS. The quality and precision of these measurements was enhanced using heavy isotope-labeled internal standards and carefully crafted protocols for sample processing.

Identification and characterization of small molecule inhibitors of the ubiquitin ligases Siah1/2 in melanoma and prostate cancer cells.

Inhibition of ubiquitin ligases with small molecule remains a very challenging task, given the lack of catalytic activity of the target and the requirement of disruption of its interactions with other proteins. Siah1/2, which are E3 ubiquitin ligases, are implicated in melanoma and prostate cancer and represent high-value drug targets. We utilized three independent screening approaches in our efforts to identify small-molecule Siah1/2 inhibitors: Affinity Selection-Mass Spectrometry, a protein thermal shift-based assay and an in silico based screen.

Molecular mechanism of a covalent allosteric inhibitor of SUMO E1 activating enzyme.

E1 enzymes activate ubiquitin (Ub) and ubiquitin-like modifiers (Ubls) in the first step of Ub/Ubl conjugation cascades and represent potential targets for therapeutic intervention in cancer and other life-threatening diseases. Here, we report the crystal structure of the E1 enzyme for the Ubl SUMO in complex with a recently discovered and highly specific covalent allosteric inhibitor (COH000). The structure reveals that COH000 targets a cryptic pocket distinct from the active site that is completely buried in all previous SUMO E1 structures and that COH000 binding to SUMO E1 is accompanied by a network of structural changes that altogether lock the enzyme in a previously unobserved inactive conformation.

Discovery of Novel Small-Molecule Inducers of Heme Oxygenase-1 That Protect Human iPSC-Derived Cardiomyocytes from Oxidative Stress.

Oxidative injury to cardiomyocytes plays a critical role in cardiac pathogenesis following myocardial infarction. Transplantation of stem cell-derived cardiomyocytes has recently progressed as a novel treatment to repair damaged cardiac tissue but its efficacy has been limited by poor survival of transplanted cells owing to oxidative stress in the post-transplantation environment. Identification of small molecules that activate cardioprotective pathways to prevent oxidative damage and increase survival of stem cells post-transplantation is therefore of great interest for improving the efficacy of stem cell therapies.

Structure-Guided Strategy for the Development of Potent Bivalent ERK Inhibitors.

ERK is the effector kinase of the RAS-RAF-MEK-ERK signaling cascade, which promotes cell transformation and malignancy in many cancers and is thus a major drug target in oncology. Kinase inhibitors targeting RAF or MEK are already used for the treatment of certain cancers, such as melanoma. Although the initial response to these drugs can be dramatic, development of drug resistance is a major challenge, even with combination therapies targeting both RAF and MEK.

MondoA coordinately regulates skeletal myocyte lipid homeostasis and insulin signaling.

Intramuscular lipid accumulation is a common manifestation of chronic caloric excess and obesity that is strongly associated with insulin resistance. The mechanistic links between lipid accumulation in myocytes and insulin resistance are not completely understood. In this work, we used a high-throughput chemical biology screen to identify a small-molecule probe, SBI-477, that coordinately inhibited triacylglyceride (TAG) synthesis and enhanced basal glucose uptake in human skeletal myocytes.

Synthesis and physicochemical characterization of novel phenotypic probes targeting the nuclear factor-kappa B signaling pathway.

Activation of nuclear factor-kappa B (NF-κB) and related upstream signal transduction pathways have long been associated with the pathogenesis of a variety of inflammatory diseases and has recently been implicated in the onset of cancer. This report provides a synthetic and compound-based property summary of five pathway-related small-molecule chemical probes identified and optimized within the National Institutes of Health-Molecular Libraries Probe Center Network (NIH-MLPCN) initiative. The chemical probes discussed herein represent first-in-class, non-kinase-based modulators of the NF-κB signaling pathway, which were identified and optimized through either cellular phenotypic or specific protein-target-based screening strategies.

Discovery and optimization of indole and 7-azaindoles as Rho kinase (ROCK) inhibitors (part-II).

Therapeutic interventions with Rho kinase (ROCK) inhibitors may effectively treat several disorders such as hypertension, stroke, cancer, and glaucoma. Herein we disclose the optimization and biological evaluation of potent novel ROCK inhibitors based on substituted indole and 7-azaindole core scaffolds. Substitutions on the indole C3 position and on the indole NH and/or amide NH positions all yielded potent and selective ROCK inhibitors (25, 42, and 50).

Discovery and optimization of indoles and 7-azaindoles as Rho kinase (ROCK) inhibitors (part-I).

Rho kinase (ROCK) inhibitors are potential therapeutic agents to treat disorders such as hypertension, multiple sclerosis, cancers, and glaucoma. Here, we disclose the synthesis, optimization, biological evaluation of potent indole and 7-azaindole based ROCK inhibitors that have high potency on ROCK (IC(50)=1 nM) with 740-fold selectivity over PKA (47). Moreover, 47 showed very good DMPK properties making it a good candidate for further development.

Identification of Inhibitors of NOD1-Induced Nuclear Factor-κB Activation.

NOD1 (nucleotide-binding oligomerization domain 1) protein is a member of the NLR (NACHT and leucine rich repeat domain containing proteins) protein family, which plays a key role in innate immunity as a sensor of specific microbial components derived from bacterial peptidoglycans and induction of inflammatory responses. Mutations in NOD proteins have been associated with various inflammatory diseases that affect NF-κB (nuclear factor κB) activity, a major signaling pathway involved in apoptosis, inflammation, and immune response. A luciferase-based reporter gene assay was utilized in a high-throughput screening program conducted under the NIH-sponsored Molecular Libraries Probe Production Center Network program to identify the active scaffolds.

Synthesis and biological evaluation of 4-quinazolinones as Rho kinase inhibitors.

Rho kinase (ROCK) is an attractive therapeutic target for various diseases including glaucoma, hypertension, and spinal cord injury. Herein, we report the development of a series of ROCK-II inhibitors based on 4-quinazolinone and quinazoline scaffolds. SAR studies at three positions of the quinazoline core led to the identification of analogs with high potency against ROCK-II and good selectivity over protein kinase A (PKA).

The development of benzimidazoles as selective rho kinase inhibitors.

Rho Kinase (ROCK) is a serine/threonine kinase whose inhibition could prove beneficial in numerous therapeutic areas. We have developed a promising class of ATP-competitive inhibitors based upon a benzimidazole scaffold, which show excellent potency toward ROCK (IC(50)<10nM). This report details the optimization of selectivity for ROCK over other related kinases such as Protein kinase A (PKA).

Benzimidazole- and benzoxazole-based inhibitors of Rho kinase.

Inhibitors of Rho kinase have been developed based on two distinct scaffolds, benzimidazoles, and benzoxazoles. SAR studies and efforts to optimize the initial lead compounds are described. Novel selective inhibitors of ROCK-II with excellent potency in both enzyme and cell-based assays were obtained.

Furanosteroid studies. Stereoselective synthesis of the A,B,E-ring core of wortmannin.

Alkyne oxazole 11c is converted in three steps, and approximately 45% overall yield, to furanolactone 21alpha having the A,B,E-ring core of the wortmannin (2) family of furanosteroids. The TiCl4-catalyzed insertion of EtO2C-CH=O between C3 and C10 in furanoacid 14d is >98% stereoselective via a pathway involving chemoselective lactonization of equilibrating aldol intermediates 23alpha,beta (dynamic kinetic resolution).

Studies on the synthesis of furanosteroids. I. Viridin models.

Alkyne oxazoles of general structure I are transformed directly to furo[2,3-b]phenol derivatives II by a sequence involving intramolecular Diels-Alder/retro-Diels-Alder reaction followed by tautomerization. Suitably functionalized phenols II undergo an intramolecular phenol-dienone-aldol condensation, generating the A,B,E-ring skeleton III characteristic of the viridin (1) class of furanosteroids.

Two-step electrochemical annulation for the assembly of polycyclic systems.

[reaction: see text] A two-step electrochemical annulation has been developed for the preparation of fused furans. The process involves an initial conjugate addition of a furyethyl cuprate and trapping of the enolate as the corresponding silyl enolether. The second step of the annulation involves the anodic coupling of the furan and the silyl enol ether to form a six-membered ring.