8 research outputs found

    Localization and Quantification of Drugs in Animal Tissues by Use of Desorption Electrospray Ionization Mass Spectrometry Imaging

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    Mass spectrometric imaging (MSI) has emerged as a powerful technique to obtain spatial arrangement of individual molecular ions in animal tissues. Ambient desorption electrospray ionization (DESI) technique is uniquely suited for such imaging experiments, as it can be performed on animal tissues in their native environment without prior treatments. Although MSI has become a rapid growing technique for localization of proteins, lipids, drugs, and endogenous compounds in different tissues, quantification of imaged targets has not been explored extensively. Here we present a novel MSI approach for localization and quantification of drugs in animal thin tissue sections. DESI-MSI using an Orbitrap mass analyzer in full scan mode was performed on 6 μm coronal brain sections from rats that were administered 2.5 mg/kg clozapine. Clozapine was localized and quantified in individual brain sections 45 min postdose. External calibration curves were prepared by micropipetting standards with internal standard (IS) on top of the tissues, and average response factors were calculated for the scans in which both clozapine and IS were detected. All response factors were normalized to area units. Quantifications from DESI-MSI revealed 0.2–1.2 ng of clozapine in individual brain sections, results that were further confirmed by extraction and liquid chromatography/tandem mass spectrometry (LC/MS/MS) analysis

    Optimization of a Novel Quinazolinone-Based Series of Transient Receptor Potential A1 (TRPA1) Antagonists Demonstrating Potent in Vivo Activity

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    There has been significant interest in developing a transient receptor potential A1 (TRPA1) antagonist for the treatment of pain due to a wealth of data implicating its role in pain pathways. Despite this, identification of a potent small molecule tool possessing pharmacokinetic properties allowing for robust in vivo target coverage has been challenging. Here we describe the optimization of a potent, selective series of quinazolinone-based TRPA1 antagonists. High-throughput screening identified <b>4</b>, which possessed promising potency and selectivity. A strategy focused on optimizing potency while increasing polarity in order to improve intrinisic clearance culminated with the discovery of purinone <b>27</b> (AM-0902), which is a potent, selective antagonist of TRPA1 with pharmacokinetic properties allowing for >30-fold coverage of the rat TRPA1 IC<sub>50</sub> in vivo. Compound <b>27</b> demonstrated dose-dependent inhibition of AITC-induced flinching in rats, validating its utility as a tool for interrogating the role of TRPA1 in in vivo pain models

    Discovery of Novel, Induced-Pocket Binding Oxazolidinones as Potent, Selective, and Orally Bioavailable Tankyrase Inhibitors

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    Tankyrase (TNKS) is a poly-ADP-ribosylating protein (PARP) whose activity suppresses cellular axin protein levels and elevates β-catenin concentrations, resulting in increased oncogene expression. The inhibition of tankyrase (TNKS1 and 2) may reduce the levels of β-catenin-mediated transcription and inhibit tumorigenesis. Compound <b>1</b> is a previously described moderately potent tankyrase inhibitor that suffers from poor pharmacokinetic properties. Herein, we describe the utilization of structure-based design and molecular modeling toward novel, potent, and selective tankyrase inhibitors with improved pharmacokinetic properties (<b>39</b>, <b>40</b>)

    The Discovery and Optimization of a Novel Class of Potent, Selective, and Orally Bioavailable Anaplastic Lymphoma Kinase (ALK) Inhibitors with Potential Utility for the Treatment of Cancer

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    A class of 2-acyliminobenzimidazoles has been developed as potent and selective inhibitors of anaplastic lymphoma kinase (ALK). Structure based design facilitated the rapid development of structure–activity relationships (SAR) and the optimization of kinase selectivity. Introduction of an optimally placed polar substituent was key to solving issues of metabolic stability and led to the development of potent, selective, orally bioavailable ALK inhibitors. Compound <b>49</b> achieved substantial tumor regression in an NPM-ALK driven murine tumor xenograft model when dosed qd. Compounds <b>36</b> and <b>49</b> show favorable potency and PK characteristics in preclinical species indicative of suitability for further development

    The Discovery and Hit-to-Lead Optimization of Tricyclic Sulfonamides as Potent and Efficacious Potentiators of Glycine Receptors

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    Current pain therapeutics suffer from undesirable psychotropic and sedative side effects, as well as abuse potential. Glycine receptors (GlyRs) are inhibitory ligand-gated ion channels expressed in nerves of the spinal dorsal horn, where their activation is believed to reduce transmission of painful stimuli. Herein, we describe the identification and hit-to-lead optimization of a novel class of tricyclic sulfonamides as allosteric GlyR potentiators. Initial optimization of high-throughput screening (HTS) hit <b>1</b> led to the identification of <b>3</b>, which demonstrated ex vivo potentiation of glycine-activated current in mouse dorsal horn neurons from spinal cord slices. Further improvement of potency and pharmacokinetics produced in vivo proof-of-concept tool molecule <b>20</b> (AM-1488), which reversed tactile allodynia in a mouse spared-nerve injury (SNI) model. Additional structural optimization provided highly potent potentiator <b>32</b> (AM-3607), which was cocrystallized with human GlyRα3<sub>cryst</sub> to afford the first described potentiator-bound X-ray cocrystal structure within this class of ligand-gated ion channels (LGICs)

    Discovery of Potent and Selective 8‑Fluorotriazolopyridine c‑Met Inhibitors

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    The overexpression of c-Met and/or hepatocyte growth factor (HGF), the amplification of the MET gene, and mutations in the c-Met kinase domain can activate signaling pathways that contribute to cancer progression by enabling tumor cell proliferation, survival, invasion, and metastasis. Herein, we report the discovery of 8-fluorotriazolopyridines as inhibitors of c-Met activity. Optimization of the 8-fluorotriazolopyridine scaffold through the combination of structure-based drug design, SAR studies, and metabolite identification provided potent (cellular IC<sub>50</sub> < 10 nM), selective inhibitors of c-Met with desirable pharmacokinetic properties that demonstrate potent inhibition of HGF-mediated c-Met phosphorylation in a mouse liver pharmacodynamic model

    Discovery of (<i>R</i>)‑6-(1-(8-Fluoro-6-(1-methyl‑1<i>H</i>‑pyrazol-4-yl)-[1,2,4]triazolo[4,3‑<i>a</i>]pyridin-3-yl)ethyl)-3-(2-methoxyethoxy)-1,6-naphthyridin-5(6<i>H</i>)‑one (AMG 337), a Potent and Selective Inhibitor of MET with High Unbound Target Coverage and Robust In Vivo Antitumor Activity

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    Deregulation of the receptor tyrosine kinase mesenchymal epithelial transition factor (MET) has been implicated in several human cancers and is an attractive target for small molecule drug discovery. Herein, we report the discovery of compound <b>23</b> (AMG 337), which demonstrates nanomolar inhibition of MET kinase activity, desirable preclinical pharmacokinetics, significant inhibition of MET phosphorylation in mice, and robust tumor growth inhibition in a MET-dependent mouse efficacy model

    Development of Novel Dual Binders as Potent, Selective, and Orally Bioavailable Tankyrase Inhibitors

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    Tankyrases (TNKS1 and TNKS2) are proteins in the poly ADP-ribose polymerase (PARP) family. They have been shown to directly bind to axin proteins, which negatively regulate the Wnt pathway by promoting β-catenin degradation. Inhibition of tankyrases may offer a novel approach to the treatment of <i>APC</i>-mutant colorectal cancer. Hit compound <b>8</b> was identified as an inhibitor of tankyrases through a combination of substructure searching of the Amgen compound collection based on a minimal binding pharmacophore hypothesis and high-throughput screening. Herein we report the structure- and property-based optimization of compound <b>8</b> leading to the identification of more potent and selective tankyrase inhibitors <b>22</b> and <b>49</b> with improved pharmacokinetic properties in rodents, which are well suited as tool compounds for further in vivo validation studies
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