Inhibitor Chemotypes Research Articles

Analysis of Botulinum Neurotoxin Serotype A Metalloprotease Inhibitors: Analogs of a Chemotype for Therapeutic Development in the Context of a Three-Zone Pharmacophore.

Botulinum neurotoxins (BoNTs), and in particular serotype A, are the most poisonous of known biological substances, and are responsible for the flaccid paralysis of the disease state botulism. Because of the extreme toxicity of these enzymes, BoNTs are considered highest priority biothreat agents. To counter BoNT serotype A (BoNT/A) poisoning, the discovery and development of small molecule, drug-like inhibitors as post-intoxication therapeutic agents has been/is being pursued. Specifically, we are focusing on inhibitors of the BoNT/A light chain (LC) (ie, a metalloprotease) subunit, since such compounds can enter neurons and provide post-intoxication protection of the enzyme target substrate. To aid/facilitate this drug development effort, a pharmacophore for inhibition of the BoNT/A LC subunit was previously developed, and is continually being refined via the incorporation of novel and diverse inhibitor chemotypes. Here, we describe several analogs of a promising therapeutic chemotype in the context of the pharmacophore for BoNT/A LC inhibition. Specifically, we describe: 1) the pharmacophoric 'fits' of the analogs and how these 'fits' rationalize the in vitro inhibitory potencies of the analogs and 2) pharmacophore refinement via the inclusion of new components from the most potent of the presented analogs.

Inhibitors of cathepsin C (dipeptidyl peptidase I)

Importance to the field: Cathepsin C (dipeptidyl peptidase I) plays a key role in the activation of several degradative enzymes linked to tissue destruction in inflammatory diseases. Thus, cathepsin C inhibitors could potentially be effective therapeutics for the treatment of such diseases as chronic obstructive pulmonary disease and cystic fibrosis.Areas covered in this review: Although this article focuses on cathepsin C inhibitor patents, the journal literature concerning small molecule inhibitors of the enzyme is also covered comprehensively (1981 – 2009).What the reader will gain: It is our aim to give the reader a complete overview of the cathepsin C inhibitor chemotypes that have been disclosed to date. In addition, key biological data have been included for both irreversible and reversible inhibitors.Take home message: All known cathepsin C inhibitors are believed to have a covalent interaction with the Cys-234 residue of the enzyme. The electrophilic and sometimes peptidic nature of these molecules is associated with poor metabolic stability and is also a potential safety concern. Thus, overcoming developability issues is a serious hurdle for these compounds and there can be little doubt that this is the principal reason why no cathepsin C inhibitors appear to have reached clinical development so far.

Design of two new chemotypes for inhibiting the Janus kinase 2 by scaffold morphing

JAK2 is a target of high interest in chronic myeloproliferative disorders drug research. Starting from a screening hit, two new JAK2 inhibitor chemotypes were designed by scaffold morphing. The prototype compounds of these new series showed nanomolar inhibition of the kinase.

A novel chemotype of kinase inhibitors: Discovery of 3,4-ring fused 7-azaindoles and deazapurines as potent JAK2 inhibitors

Pictet-Spengler condensation of aldehydes or alpha-keto-esters with 4-(2-anilinophenyl)-7-azaindole (11) or deazapurine (12) gave high yields of the 3,4-fused cyclic compounds. SAR studies, by varying the substituted benzaldehyde components, lead to the discovery of a series of potent JAK2 kinase inhibitors.

Structure-Based Design of a Benzodiazepine Scaffold Yields a Potent Allosteric Inhibitor of Hepatitis C NS5B RNA Polymerase

HCV NS5B polymerase, an essential and virus-specific enzyme, is an important target for drug discovery. Using structure-based design, we optimized a 1,5-benzodiazepine NS5B polymerase inhibitor chemotype into a new sulfone-containing scaffold. The design yielded potent inhibitor (S)-4c (K(D) = 0.79 nM), which has approximately 20-fold greater affinity for NS5B than its carbonyl analogue (R)-2c.

Cdc25B Dual-Specificity Phosphatase Inhibitors Identified in a High-Throughput Screen of the NIH Compound Library

The University of Pittsburgh Molecular Library Screening Center (Pittsburgh, PA) conducted a screen with the National Institutes of Health compound library for inhibitors of in vitro cell division cycle 25 protein (Cdc25) B activity during the pilot phase of the Molecular Library Screening Center Network. Seventy-nine (0.12%) of the 65,239 compounds screened at 10 muM met the active criterion of > or =50% inhibition of Cdc25B activity, and 25 (31.6%) of these were confirmed as Cdc25B inhibitors with 50% inhibitory concentration (IC(50)) values <50 microM. Thirteen of the Cdc25B inhibitors were represented by singleton chemical structures, and 12 were divided among four clusters of related structures. Thirteen (52%) of the Cdc25B inhibitor hits were quinone-based structures. The Cdc25B inhibitors were further characterized in a series of in vitro secondary assays to confirm their activity, to determine their phosphatase selectivity against two other dual-specificity phosphatases, mitogen-activated protein kinase phosphatase (MKP)-1 and MKP-3, and to examine if the mechanism of Cdc25B inhibition involved oxidation and inactivation. Nine Cdc25B inhibitors did not appear to affect Cdc25B through a mechanism involving oxidation because they did not generate detectable amounts of H(2)O(2) in the presence of dithiothreitol, and their Cdc25B IC(50) values were not significantly affected by exchanging the dithiothreitol for beta-mercaptoethanol or reduced glutathione or by adding catalase to the assay. Six of the nonoxidative hits were selective for Cdc25B inhibition versus MKP-1 and MKP-3, but only the two bisfuran-containing hits, PubChem substance identifiers 4258795 and 4260465, significantly inhibited the growth of human MBA-MD-435 breast and PC-3 prostate cancer cell lines. To confirm the structure and biological activity of 4260465, the compound was resynthesized along with two analogs. Neither of the substitutions to the two analogs was tolerated, and only the resynthesized hit 26683752 inhibited Cdc25B activity in vitro (IC(50) = 13.83 +/- 1.0 microM) and significantly inhibited the growth of the MBA-MD-435 breast and PC-3 prostate cancer cell lines (IC(50) = 20.16 +/- 2.0 microM and 24.87 +/- 2.25 microM, respectively). The two bis-furan-containing hits identified in the screen represent novel nonoxidative Cdc25B inhibitor chemotypes that block tumor cell proliferation. The availability of non-redox active Cdc25B inhibitors should provide valuable tools to explore the inhibition of the Cdc25 phosphatases as potential mono- or combination therapies for cancer.

Trypanothione Reductase High-Throughput Screening Campaign Identifies Novel Classes of Inhibitors with Antiparasitic Activity

High-throughput screening of 100,000 lead-like compounds led to the identification of nine novel chemical classes of trypanothione reductase (TR) inhibitors worthy of further investigation. Hits from five of these chemical classes have been developed further through different combinations of preliminary structure-activity relationship rate probing and assessment of antiparasitic activity, cytotoxicity, and chemical and in vitro metabolic properties. This has led to the identification of novel TR inhibitor chemotypes that are drug-like and display antiparasitic activity. For one class, a series of analogues have displayed a correlation between TR inhibition and antiparasitic activity. This paper explores the process of identifying, investigating, and evaluating a series of hits from a high-throughput screening campaign.

DNA gyrase (GyrB)/topoisomerase IV (ParE) inhibitors: Synthesis and antibacterial activity

The synthesis and antibacterial activities of three chemotypes of DNA supercoiling inhibitors based on imidazolo[1,2- a]pyridine and [1,2,4]triazolo[1,5- a]pyridine scaffolds that target the ATPase subunits of DNA gyrase and topoisomerase IV (GyrB/ParE) is reported. The most potent scaffold was selected for optimization leading to a series with potent Gram-positive antibacterial activity and a low resistance frequency.

Toward an Improved Clustering of Large Data Sets Using Maximum Common Substructures and Topological Fingerprints

A new clustering algorithm was developed that is able to group large data sets with more than 100,000 molecules according to their chemotypes. The algorithm preclusters a data set using a fingerprint version of the hierarchical k-means algorithm. Chemotypes are extracted from the terminal clusters via a maximum common substructure approach. Molecules forming a chemotype have to share a predefined number of rings, atoms, and non-carbon heavy atoms. In an iterative procedure, similar chemotypes and singletons are fused to larger chemotypes. Singletons that cannot be assigned to any chemotype are then grouped based on the proportion of overlap between the molecules. Representatives from each chemotype and the singletons are used in a second round of the hierarchical k-means algorithm to provide a final hierarchical grouping. Results are reported to an interactive graphical user interface which allows initial insights about the structure activity relationship (SAR) of the molecules. Example applications are shown for two chemotypes of reverse transcriptase inhibitors in the MDDR database and for the evaluation of descriptor-based similarity searching routines. A special focus was laid on the chemotype hopping potential of each individual routine. The algorithm will allow the analysis of high-throughput and virtual screening results with improved quality.

Targeting RSK: An Overview of Small Molecule Inhibitors

Ribosomal S6 kinase (RSK) is a family of serine/threonine kinases that has been identified as a promising anti-cancer target. While a number of protein kinase inhibitors that have potent activity against other serine/threonine kinases were shown to also inactivate RSK, there is keen interest in the three different inhibitor chemotypes that were shown to be RSK specific, since these compounds have tremendous utility as chemical probes in elucidating the biochemistry of the RSK signaling cascade and unraveling the molecular basis of cancer. Because each compound may have therapeutic potential, the nonspecific kinase inhibitors as well as the RSK specific inhibitors will be discussed.

A Specific Mechanism for Nonspecific Activation in Reporter-Gene Assays

The importance of bioluminescence in enabling a broad range of high-throughput screening (HTS) assay formats is evidenced by widespread use in industry and academia. Therefore, understanding the mechanisms by which reporter enzyme activity can be modulated by small molecules is critical to the interpretation of HTS data. In this Perspective, we provide evidence for stabilization of luciferase by inhibitors in cell-based luciferase reporter-gene assays resulting in the counterintuitive phenomenon of signal activation. These data were derived from our analysis of luciferase inhibitor compound structures and their prevalence in the Molecular Libraries Small Molecule Repository using 100 HTS experiments available in PubChem. Accordingly, we found an enrichment of luciferase inhibitors in luciferase reporter-gene activation assays but not in assays using other reporters. In addition, for several luciferase inhibitor chemotypes, we measured reporter stabilization and signal activation in cells that paralleled the inhibition determined using purified luciferase to provide further experimental support for these contrasting effects.

Structure–Activity Studies on Splitomicin Derivatives as Sirtuin Inhibitors and Computational Prediction of Binding Mode

NAD (+)-dependent histone deacetylases (sirtuins) are enzymes that cleave acetyl groups from lysines in histones and other proteins. Potent selective sirtuin inhibitors are interesting tools for the investigation of the biological functions of those enzymes and may be future drugs for the treatment of cancer. Splitomicin was among the first two inhibitors that were discovered for yeast sirtuins but showed rather weak inhibition on human enzymes. We present detailed structure-activity relationships on splitomicin derivatives and their inhibition of recombinant Sirt2. To rationalize our experimental results, ligand docking followed by molecular mechanics Poisson-Boltzmann/surface area (MM-PBSA) calculations were carried out. These analyses suggested a molecular basis for the interaction of the beta-phenylsplitomicins with human Sirt2. Protein-based virtual screening resulted in the identification of a novel Sirt2 inhibitor chemotype. Selected inhibitors showed antiproliferative properties and tubulin hyperacetylation in MCF7 breast cancer cells and are promising candidates for further optimization as potential anticancer drugs.

Activation of Inhibitors by Sortase Triggers Irreversible Modification of the Active Site

Sortases anchor surface proteins to the cell wall of Gram-positive pathogens through recognition of specific motif sequences. Loss of sortase leads to large reductions in virulence, which identifies sortase as a target for the development of antibacterials. By screening 135,625 small molecules for inhibition, we report here that aryl (beta-amino)ethyl ketones inhibit sortase enzymes from staphylococci and bacilli. Inhibition of sortases occurs through an irreversible, covalent modification of their active site cysteine. Sortases specifically activate this class of molecules via beta-elimination, generating a reactive olefin intermediate that covalently modifies the cysteine thiol. Analysis of the three-dimensional structure of Bacillus anthracis sortase B with and without inhibitor provides insights into the mechanism of inhibition and reveals binding pockets that can be exploited for drug discovery.

Pyrrolo[3,4‐c]quinoline‐1,3‐diones as Potent Caspase‐3 Inhibitors. Synthesis and SAR of 2‐Substituted 4‐Methyl‐8‐(morpholine‐4‐sulfonyl)‐pyrrolo[3,4‐c]quinoline‐1,3‐diones.

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In silico identification of novel EGFR inhibitors with antiproliferative activity against cancer cells

Inhibition of the epidermal growth factor receptor (EGFR) tyrosine kinase activity by small molecules has proved effective for the treatment of cancer. To the best of our knowledge, the crystal structure of EGFR has been used for the first time to identify novel inhibitor chemotypes by docking-based in silico screening of a large virtual chemical library followed up by experimental validation. We identified several compounds with antiproliferative effects on cancer cells. Amongst them, a C(4)-N(1)-substituted pyrazolo[3,4- d]pyrimidine MSK-039 ( 39) was discovered as a low-micromolar inhibitor of EGFR tyrosine kinase activity. The predicted binding mode of 39 opens a new avenue toward the optimization of novel chemical entities to develop potent and selective inhibitors of EGFR signaling.

Steroid sulfatase inhibitors

Steroid sulfatase (STS), a microsomal enzyme catalysing the hydrolysis of the sulfate esters of 3-hydroxy steroids, has received considerable attention as an attractive drug target. It is crucial for the local production of active oestrogens and androgens from their systemic circulating sulfated precursor, namely oestrone sulfate (E1S) and dehydroepiandrosterone sulfate (DHEAS), which are involved in the pathogenesis of a number of diseases. STS inhibitors have been proposed for the treatment of breast, endometrial and prostate cancer, and for androgen-dependent skin diseases such as alopecia, hirsutism and acne. In addition to various classes of potent irreversible arylsulfamate-based inhibitors, recently novel chemotypes of reversible inhibitors originating from high-throughput screening (HTS) were discovered. All STS inhibitors to date are in the preclinical stage, with the exception of 667COUMATE, which is presently in a Phase I clinical trial. In this review the latest patent disclosures and literature in the field of STS inhibitors are summarised.

Pyrrolo[3,4-c]quinoline-1,3-diones as potent caspase-3 inhibitors. Synthesis and SAR of 2-substituted 4-methyl-8-(morpholine-4-sulfonyl)-pyrrolo[3,4- c]quinoline-1,3-diones

Synthesis, biological evaluation and structure–activity relationships for a series of 2-substituted 4-methyl-8-(morpholine-4-sulfonyl)-1,3-dioxo-2,3-dihydro-1 H-pyrrolo[3,4- c]quinolines are described. These compounds represent a new chemotype of nonpeptide small molecule inhibitors of caspase-3. Among the studied compounds, several potent inhibitors with IC 50 in the range of 3–10 nM have been identified. The most active compound within this series, 7 {49} and 7 {58}, inhibited caspase-3 with IC 50 = 3 nM.

N-Acyl arylsulfonamides as novel, reversible inhibitors of human steroid sulfatase

Steroid sulfatase (STS) is an attractive target for a range of oestrogen- and androgen-dependent diseases. In search of novel chemotypes of STS inhibitors, we had previously identified nortropinyl–arylsulfonylureas 1; however, while these compounds were good inhibitors of purified STS (lowest K i = 76 nM), they showed only weak inhibition of STS activity in cells (lowest IC 50 around 2 μM). Extended structure–activity relationship studies involving modification of the phenylacetyl side chain and replacement of the nortropine element by simpler scaffolds led to the discovery of N-acyl arylsulfonamides, more specifically N-(Boc-piperidine-4-carbonyl)-benzenesulfonamides, as STS inhibitors, some of which exhibit improved cellular potency (best IC 50 = 270 nM).

Discovery and optimization of 2-aryl oxazolo-pyrimidines as adenosine kinase inhibitors using liquid phase parallel synthesis

Adenosine kinase inhibition is an attractive therapeutic approach for several conditions for example, neurodegeneration, seizures, ischemia, inflammation and pain. Several nucleosidic and non-nucleosidic inhibitors are available. Using a virtual screening approach, we have discovered that 2-aryl oxazolo-pyrimidines are adenosine kinase inhibitors. Subsequent high throughput derivatization enabled the optimization of this new inhibitor chemotype resulting in highly potent derivatives. A variety of analogues were produced by applying liquid phase parallel synthesis to vary the 7-amino residues as well as the 2-aryl moiety.

Total synthesis and stereochemistry of (-)-cacospongionolide F.

[reaction: see text] The most recently described member of the cacospongionolide class of marine natural products has been assembled using a diastereochemically divergent total synthesis strategy that independently establishes the complete stereochemistry of cacospongionolide F and provides a new entry toward expansion of this phospholipase A(2) inhibitor chemotype.