DOCSLIB.ORG

Equilibrium Assays Are Required to Accurately Characterize the Activity Profiles of Drugs Modulating Gq-Protein-Coupled Receptors S

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Equilibrium Assays Are Required to Accurately Characterize the Activity Profiles of Drugs Modulating Gq-Protein-Coupled Receptors S Supplemental material to this article can be found at: http://molpharm.aspetjournals.org/content/suppl/2018/06/28/mol.118.112573.DC1 1521-0111/94/3/992–1006$35.00 https://doi.org/10.1124/mol.118.112573 MOLECULAR PHARMACOLOGY Mol Pharmacol 94:992–1006, September 2018 Copyright ª 2018 by The American Society for Pharmacology and Experimental Therapeutics Equilibrium Assays Are Required to Accurately Characterize the Activity Profiles of Drugs Modulating Gq-Protein-Coupled Receptors s Sara Bdioui, Julien Verdi, Nicolas Pierre, Eric Trinquet, Thomas Roux, and Terry Kenakin Cisbio Bioassays, Codolet, France (S.B., J.V., N.P., E.T., T.R.); and Department of Pharmacology, University of North Carolina School of Medicine, Chapel Hill, North Carolina (T.K.) Received March 25, 2018; accepted June 11, 2018 Downloaded from ABSTRACT This paper discusses the process of determining the activity of or ghrelin receptors were tested using two functional assays candidate molecules targeting Gq-protein activation through (calcium release and IP1) and the results were compared with G-protein-coupled receptors for possible therapeutic applica- theoretical pharmacological models. The IP1 assay is an tion with two functional assays; calcium release and inositol equilibrium assay that is able to determine the correct (i.e., molpharm.aspetjournals.org phosphate metabolism [inositol monophosphate (IP1)]. While internally consistent) pharmacological profiles of all tested both are suitable for detecting ligand activity (screening), compounds. In contrast, the nonequilibrium nature of calcium differences are seen when these assays are used to quantita- assays yields misleading classification of most of the tested tively measure ligand parameters for therapeutic activity. compounds. Our study suggests that the use of an equilibrium Specifically, responses for Gq-related pathways present dif- assay, such as IP1, is mandatory for the optimal use of ferent and dissimulating patterns depending on the functional pharmacological models that can both identify mechanisms assay used to assess them. To investigate the impact of of action and also convert descriptive-to-predictive data for functional assays on the accuracy of compound pharmacolog- therapeutic systems. Such assays allow the identification of ical profiles, five exemplar molecules [partial agonist, antago- consistent and simple scales of activity that can guide nist, inverse agonist, positive allosteric modulator (PAM) medicinal chemistry in lead optimization of candidate mole- at ASPET Journals on September 27, 2021 agonist, and positive b-PAM] targeting either muscarinic M1 cules for therapeutic use. Introduction in all functional systems. A corollary to this approach is that these same models often provide internal checks, whereby The process of drug lead optimization involves the character- certain predictions must coincide with the observations to ization of the molecular profile of a drug candidate with system- correctly ascribe drug mode of action. For this procedure to be independent scales of drug activity, i.e., the profile ideally should successful, it is imperative that the assays used to determine not be linked only to the experimental system where it is drug function accurately reflect the molecular activity of the observed. This is important since drugs are almost always molecule at the receptor. studied and developed in test experimental systems and not There are two fundamentally different types of assay used the end therapeutic tissue. Therefore, a scale of activity must in drug discovery: detection of drug activity (screening) and bridge the gap between these two systems. Pharmacologically characterization of activity (lead optimization). Detection this is done by comparing experimental data to mathematical assays should be robust and sensitive; however, they only models of drug-receptor interaction and deriving universal need to reveal ligand-receptor interaction and not necessarily parameters such as equilibrium dissociation constants [equilib- quantitatively correct drug parameters. In contrast, lead rium dissociation constant of the agonist-receptor complex (K ) A optimization assays must provide predictive and accurate and equilibrium dissociation constant of antagonist (or allosteric measures of drug activity that can be used to characterize modulator)-receptor complex (KB)], measures of efficacy (t), or activity in therapeutic systems. The activation of Gq-protein- allosteric parameters describing effects on affinity (a)and coupled receptors results in increases in inositol triphosphate efficacy (b), which then can be used to predict agonist activity metabolism and calcium release in cells (Berridge et al., 2003). There are two main and well-described fluorescent-based https://doi.org/10.1124/mol.118.112573. s This article has supplemental material available at molpharm. assays that can be used to monitor the activation of the Gq aspetjournals.org. pathway. On the one hand, calcium assays measure the release ABBREVIATIONS: ACh, acetylcholine; BQCA, benzyl quinolone carboxylic acid [1-(4-methoxybenzyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid]; CHO, Chinese hamster ovary; DR, dose ratio; GHSR1a, growth hormone secretagogue receptor type 1a; HEK, human embryonic kidney; HR, heart rate; IP1, inositol monophosphate; KA, equilibrium dissociation constant of the agonist-receptor complex; KB, equilibrium dissociation constant of antagonist (or allosteric modulator)-receptor complex; NECA, 59-N-ethylcarboxamidoadenosine; PAM, positive allosteric modulator; SPA, [D-Arg1,D-Phe5,DTrp7,9,Leu11]-substance P. 992 Functional Assays for Gq-Protein–Activating Receptor Ligands 993 of calcium flux into the cells using permeant dyes, in which complemented culture medium at the desired concentration. Cells fluorescence emission increases upon binding to calcium were seeded into 96-well culture-treated plates for IP1 and calcium (Chambers et al., 2003). On the other hand, the inositol mono- experiments (Greiner Bio-One, Courtabœuf, France) at 100 ml/well phosphate (IP1) assay measures the accumulation of IP1, which is and incubated overnight at 37°C with 5% CO2 before being used in a metabolite of inositol triphosphate, through a competition assay functional assays. based on homogeneous time-resolved fluorescence technology (Trinquet et al., 2011). It will be seen that both IP1 and calcium Measurement of the Intracellular Accumulation of IP1 assays function well in screening mode but differ considerably in The homogeneous time-resolved fluorescence IP-One-Gq assay (IP1) lead optimization mode. This paper discusses whether these was used to measure intracellular IP1 accumulation according to the functional readouts deliver accurate data for ligand characteriza- manufacturer’s instructions (Cisbio Bioassays, Codolet, France). In tion or if they lead to dissimulating and misleading profiles. brief, on the day of the experiment, serial dilutions of compounds were It is well known that calcium transients emanate from prepared in the stimulation buffer of the kit (containing 50 mM LiCl to prevent IP1 degradation) at the desired working concentration. Cell hemi-equilibrium assays that do not furnish sustained agonist culture medium supernatant was removed from the plates. After a responses; therefore, pharmacological procedures normally washing step with phosphate-buffered saline, cells were treated by requiring these, such as detecting inverse, partial, and addition of 70 ml/well of the different compounds and incubated for the positive allosteric modulator (PAM) agonism, are precluded. indicated time at 37°C, 5% CO2. Then, IP1 detection was allowed by Downloaded from However, there are published procedures that have been used addition of the detection reagents previously diluted in the appropriate to furnish mass action pharmacological profiles for these types kit lysis and detection buffer (30 ml total). Plates were incubated for of ligands through coaddition formats, which apparently yield 1 hour at room temperature and the homogeneous time-resolved the correct profiles for some of these types of ligands. It can be fluorescence signal was recorded using a PHERAstar FS reader (BMG shown that while these procedures apparently provide correct LABTECH, Ortenberg, Germany) with flash lamp excitation. qualitative patterns of drug effect, they in fact can yield Assay parameters, such as the cellular density required to work within the assay dynamic range or the determination of the appro- molpharm.aspetjournals.org erroneous drug parameters that lead to misclassification of priate stimulation time for each compound (i.e., ACh, Alvameline, drug activity. The key to identifying such dissimulations is the Atropine), were properly optimized with the cellular models used application of internal checks within the mathematical models (CHO-M1 and HEK293-GHSR1a). of drug receptor pharmacodynamics. These concepts are illustrated in the present paper with examples of five general Measurement of Calcium types of ligands [agonists, antagonists, inverse agonists, PAM agonists, and PAMs increasing agonist efficacy (b-PAMs)]. After cell culture medium removal, cells were washed once with phosphate-buffered saline and incubated for 1 hour at 37°C, 5% CO Specifically, the results obtained suggest that the use of an 2 with 2 mM of Fluo-4 AM (ThermoFisher Scientific, Villebon sur Yvette, equilibrium assay based on the determination of the intracel- France) previously diluted in Hanks’ balanced salt solution (Thermo-
Load more

Recommended publications
  • E30 SEM. O.C. Disclosed Is a Compound Represented by the Formula (1) (51) Int
    USOO9453000B2 (12) United States Patent (10) Patent No.: US 9.453,000 B2 Kimura et al. (45) Date of Patent: *Sep. 27, 2016 (54) POLYCYCLIC COMPOUND (56) References Cited (75) Inventors: Teiji Kimura, Tsukuba (JP); Noritaka U.S. PATENT DOCUMENTS Kitazawa, Tsukuba (JP); Toshihiko 3,470,167 A 9, 1969 Sarkar Kaneko, Tsukuba (JP); Nobuaki Sato, 3,989,816 A 1 1/1976 Rajadhyaksha Tsukuba (JP); Koki Kawano, Tsukuba 4,910,200 A 3, 1990 Curtze et al. (JP): Koichi Ito, Tsukuba (JP); 5,281,626 A 1/1994 Oinuma et al. M s Tak ishi Tsukub JP 5,563,162 A 10, 1996 Oku et al. amoru Takaishi Tsukuba (JP); 5,804,577 A 9, 1998 Hebeisen et al. Takeo Sasaki, Tsukuba (JP); Yu 5,985,856 A 11/1999 Stella et al. Yoshida, Tsukuba (JP); Toshiyuki 6,235,728 B1 5, 2001 Golik et al. Uemura, Tsukuba (JP); Takashi Doko, g R 1939. E. al. Its SE E. Shinmyo, 7,138.414 B2 11/2006 Schoenafingereatch et al. et al. sukuba (JP); Daiju Hasegawa, 7,300,936 B2 11/2007 Parker et al. Tsukuba (JP); Takehiko Miyagawa, 7,314,940 B2 1/2008 Graczyk et al. Hatfield (GB); Hiroaki Hagiwara, 7,618,960 B2 11/2009 Kimura et al. Tsukuba (JP) 7,667,041 B2 2/2010 Kimura et al. 7,687,640 B2 3/2010 Kimura et al. 7,713,993 B2 5/2010 Kimura et al. (73) Assignee: EISAI R&D MANAGEMENT CO., 7,737,141 B2 6/2010 Kimura et al. LTD., Tokyo (JP) 7,880,009 B2 2/2011 Kimura et al.
    [Show full text]
  • PHARMACEUTICAL APPENDIX to the TARIFF SCHEDULE 2 Table 1
    Harmonized Tariff Schedule of the United States (2020) Revision 19 Annotated for Statistical Reporting Purposes PHARMACEUTICAL APPENDIX TO THE HARMONIZED TARIFF SCHEDULE Harmonized Tariff Schedule of the United States (2020) Revision 19 Annotated for Statistical Reporting Purposes PHARMACEUTICAL APPENDIX TO THE TARIFF SCHEDULE 2 Table 1. This table enumerates products described by International Non-proprietary Names INN which shall be entered free of duty under general note 13 to the tariff schedule. The Chemical Abstracts Service CAS registry numbers also set forth in this table are included to assist in the identification of the products concerned. For purposes of the tariff schedule, any references to a product enumerated in this table includes such product by whatever name known.
    [Show full text]
  • Alzheimer's Disease Clinical Trials
    Clinical Trial Perspective 5 Clinical Trial Perspective Alzheimer’s disease clinical trials: past failures and future opportunities Clin. Invest. (Lond.) Over a decade has elapsed since the US FDA has approved a medication for Alzheimer’s Roy Yaari*,1,2 & Ann Hake1,2 disease (AD) despite clinical trials of numerous agents over a wide array of mechanisms 1Eli Lilly & Company, Lilly Corporate including neurotransmitter modulation and disease modifying therapy targeting Center, Indianapolis, IN 46285, USA 2Indiana University School of Medicine, amyloid and tau. The failures of clinical trials in AD may be due to inadequate Department of Neurology, Indianapolis, understanding of mechanisms of action and/or poor target engagement; however, IN 46202, USA other factors could include inadequate study design, stage of AD along the continuum *Author for correspondence: studied, inclusion of participants without Alzheimer’s pathology into clinical trials Tel.: +1 317 651 6163 and limited power of endpoint measures. Future studies will need to carefully assess [email protected] these possible shortcomings in design of upcoming trials, especially as the field moves toward studies of disease modifying agents (as opposed to symptomatic treatment) of AD and to patients that are very early in the disease spectrum. Keywords: Alzheimer’s disease • Alzheimer’s disease biomarkers • amyloid • clinical trials • preclinical Alzheimer’s disease • tau US FDA approved medications continue to provide significant, but modest More than three decades ago, the choliner- symptomatic benefit[5–7] . gic hypothesis proposed that degeneration The compound memantine introduced a of cholinergic neurons in the basal fore- second mechanism for symptomatic treat- brain and the associated loss of cholinergic ment of AD into clinical practice.
    [Show full text]
  • The Use of Stems in the Selection of International Nonproprietary Names (INN) for Pharmaceutical Substances
    WHO/PSM/QSM/2006.3 The use of stems in the selection of International Nonproprietary Names (INN) for pharmaceutical substances 2006 Programme on International Nonproprietary Names (INN) Quality Assurance and Safety: Medicines Medicines Policy and Standards The use of stems in the selection of International Nonproprietary Names (INN) for pharmaceutical substances FORMER DOCUMENT NUMBER: WHO/PHARM S/NOM 15 © World Health Organization 2006 All rights reserved. Publications of the World Health Organization can be obtained from WHO Press, World Health Organization, 20 Avenue Appia, 1211 Geneva 27, Switzerland (tel.: +41 22 791 3264; fax: +41 22 791 4857; e-mail: [email protected]). Requests for permission to reproduce or translate WHO publications – whether for sale or for noncommercial distribution – should be addressed to WHO Press, at the above address (fax: +41 22 791 4806; e-mail: [email protected]). The designations employed and the presentation of the material in this publication do not imply the expression of any opinion whatsoever on the part of the World Health Organization concerning the legal status of any country, territory, city or area or of its authorities, or concerning the delimitation of its frontiers or boundaries. Dotted lines on maps represent approximate border lines for which there may not yet be full agreement. The mention of specific companies or of certain manufacturers’ products does not imply that they are endorsed or recommended by the World Health Organization in preference to others of a similar nature that are not mentioned. Errors and omissions excepted, the names of proprietary products are distinguished by initial capital letters.
    [Show full text]
  • Equilibrium Assays Are Required to Accurately Characterize the Activity Profiles of Drugs
    Molecular Pharmacology Fast Forward. Published on June 28, 2018 as DOI: 10.1124/mol.118.112573 This article has not been copyedited and formatted. The final version may differ from this version. MOL112573 Equilibrium Assays are Required to Accurately Characterize the Activity Profiles of Drugs Modulating Gq-Coupled GPCRs Sara Bdioui, Julien Verdi, Nicolas Pierre, Eric Trinque, Thomas Roux, Terry Kenakin SB, JV, NP, ET, TR: Cisbio Bioassays, 30200 Codolet, France TK : Department of Pharmacology, University of North Carolina School of Medicine, Chapel Hill, North Carolina Downloaded from molpharm.aspetjournals.org at ASPET Journals on September 26, 2021 1 Molecular Pharmacology Fast Forward. Published on June 28, 2018 as DOI: 10.1124/mol.118.112573 This article has not been copyedited and formatted. The final version may differ from this version. MOL112573 RUNNING (SHORT) TITLE Functional assays for Gq Protein activating Receptor Ligands Corresponding author: Terry Kenakin Ph.D. Professor, Department of Pharmacology University of North Carolina School of Medicine Downloaded from 120 Mason Farm Road Room 4042 Genetic Medicine Building, CB# 7365 molpharm.aspetjournals.org Chapel Hill, NC 27599-7365 Phone: 919-962-7863 Fax: 919-966-7242 or 5640 Email: [email protected] at ASPET Journals on September 26, 2021 Number of text pages: 56 Number of tables: 0 Number of figures: 13 Number of references: 33 Number of words in the Abstract: 233 Number of words in the Introduction: 599 Number of words in the Discussion: 1497 2 Molecular Pharmacology Fast Forward. Published on June 28, 2018 as DOI: 10.1124/mol.118.112573 This article has not been copyedited and formatted.
    [Show full text]
  • I Regulations
    23.2.2007 EN Official Journal of the European Union L 56/1 I (Acts adopted under the EC Treaty/Euratom Treaty whose publication is obligatory) REGULATIONS COUNCIL REGULATION (EC) No 129/2007 of 12 February 2007 providing for duty-free treatment for specified pharmaceutical active ingredients bearing an ‘international non-proprietary name’ (INN) from the World Health Organisation and specified products used for the manufacture of finished pharmaceuticals and amending Annex I to Regulation (EEC) No 2658/87 THE COUNCIL OF THE EUROPEAN UNION, (4) In the course of three such reviews it was concluded that a certain number of additional INNs and intermediates used for production and manufacture of finished pharmaceu- ticals should be granted duty-free treatment, that certain of Having regard to the Treaty establishing the European Commu- these intermediates should be transferred to the list of INNs, nity, and in particular Article 133 thereof, and that the list of specified prefixes and suffixes for salts, esters or hydrates of INNs should be expanded. Having regard to the proposal from the Commission, (5) Council Regulation (EEC) No 2658/87 of 23 July 1987 on the tariff and statistical nomenclature and on the Common Customs Tariff (1) established the Combined Nomenclature Whereas: (CN) and set out the conventional duty rates of the Common Customs Tariff. (1) In the course of the Uruguay Round negotiations, the Community and a number of countries agreed that duty- (6) Regulation (EEC) No 2658/87 should therefore be amended free treatment should be granted to pharmaceutical accordingly, products falling within the Harmonised System (HS) Chapter 30 and HS headings 2936, 2937, 2939 and 2941 as well as to designated pharmaceutical active HAS ADOPTED THIS REGULATION: ingredients bearing an ‘international non-proprietary name’ (INN) from the World Health Organisation, specified salts, esters or hydrates of such INNs, and designated inter- Article 1 mediates used for the production and manufacture of finished products.
    [Show full text]
  • Advice Concerning the Addition of Certain Pharmaceutical Products
    U.S. International Trade Commission COMMISSIONERS Daniel R. Pearson, Chairman Shara L. Aranoff, Vice Chairman Jennifer A. Hillman Stephen Koplan Deanna Tanner Okun Charlotte R. Lane Robert A. Rogowsky Director of Operations Karen Laney-Cummings Director of Industries Address all communications to Secretary to the Commission United States International Trade Commission Washington, DC 20436 U.S. International Trade Commission Washington, DC 20436 www.usitc.gov Advice Concerning the Addition of Certain Pharmaceutical Products and Chemical Intermediates to the Pharmaceutical Appendix to the Harmonized Tariff Schedule of the United States Investigation No. 332--476 Publication 3883 September 2006 This report was prepared principally by Office of Industries Philip Stone, Project Leader With assistance from Elizabeth R. Nesbitt Primary Reviewers David G. Michels, Office of Tariff Affairs and Trde Agreements, John Benedetto, and Nannette Christ, Office of Economics Administrative Support Brenda F. Carroll Under the direction of Dennis Rapkins, Chief Chemicals and Textiles Division ABSTRACT Under the Pharmaceutical Zero-for-Zero Initiative, which entered into force in 1995, the United States and its major trading partners eliminated tariffs on many pharmaceuticals, their derivatives, and certain chemical intermediates used to make pharmaceuticals. The U.S. list of pharmaceutical products and chemical intermediates eligible for duty-free treatment under the agreement is given in the Pharmaceutical Appendix to the Harmonized Tariff Schedule of the United States. The Pharmaceutical Appendix is periodically updated to provide duty relief for additional such products, including newly developed pharmaceuticals. This report provides advice on the third update to the agreement, in which approximately 1,300 products are proposed to receive duty-free treatment.
    [Show full text]
  • (12) United States Patent (10) Patent No.: US 8,673,964 B2 Lautt (45) Date of Patent: Mar
    USOO8673964B2 (12) United States Patent (10) Patent No.: US 8,673,964 B2 Lautt (45) Date of Patent: Mar. 18, 2014 (54) USE OF DRUG COMBINATIONS FOR OTHER PUBLICATIONS TREATING INSULIN RESISTANCE Yki-Jarvinen (Cobination Therapies with Insulin in Type 2 diabetes, Deabetes care, vol. 24, No. 4, pp. 758-767).* (75) Inventor: Wilfred Wayne Lautt, Winnipeg (CA) Holz (Glucagol-Like Peptide-1 Synthetic Analogs: New Therapeutic Agents for Use in the Treatment of Diabetes Mellitus, Curr Med (73) Assignee: DiaMedica Inc. (CA) Chem, Nov. 2003, 10(22): pp. 2471-2483).* Nodari (Efficacy and tolerability of the long-term administration of (*) Notice: Subject to any disclaimer, the term of this carvedilol in patients with chronic heart failure with and without patent is extended or adjusted under 35 concomitant diabetes mellitus, The European Journal of Heart Fail ure, 2003, vol. 4, pp. 803-809).* U.S.C. 154(b) by 971 days. Beyer et al. "Assessment of Insulin Needs in Insulin-Dependent Diabetics and Healthy Volunteers under Fasting Conditions' Horm (21) Appl. No.: 11/597,032 Metab Res Suppl 1990; vol. 24 p.71-77. Brownlee “Biochemistry and molecular cell biology of diabetic com (22) PCT Filed: May 20, 2005 plications' Nature vol. 414, Dec. 2001 p. 813-820. Hsu et al. “Five Cysteine-Containing Compounds Delay Diabetic Deterioration in Balb/cA Mice' J. Nutr, 2004 134:3245-3249. (86). PCT No.: PCT/CA2005/000775 Lautt “Hepatic Parasympathetic Neuropathy as cause of Maturity S371 (c)(1), Onset Diabetes?” Gen. Pharmac. vol. 11 p. 343-345. Xie et al. “Induction of insulin resistance by cholinergic blockade (2), (4) Date: May 7, 2008 with atropine in the cat' J.
    [Show full text]
  • Alzheimer Syllabus
    Update on Alzheimer’s Disease June, 2006 Stephen S. Flitman, MD, Medical Director 21 st Century Neurology 3100 North Third Ave Suite 100, Phoenix, Arizona (602) 265-6500 www.neurozone.org Alzheimer’s Disease (AD), first described by Dr. Alois Alzheimer in 1907, is a disorder that 10 million Americans are thought to have, but less than 40% are diagnosed or receiving treatment now. It is very common, affecting people ages 40-90 with 2-4% prevalence at ages 65- 75 and increasing in subsequent decades. After heart disease, cancer, and stroke it is the fourth leading cause of death in the US. AD is a disorder of the gray matter of the cerebral cortex. It is characterized clinically by gradually progressive dementia and pathologically by extraneuronal plaques and intraneuronal neurofibrillary tangles by light microscopy. AD progresses anatomically via the connections made by affected neurons. It starts in the entorhinal cortex and then spreads to the hippocampus, basal nuclei, and then to the neocortex. It spares primary motor and sensory cortices, preferentially attacking association areas in the frontal, parietal, and temporal lobes. Affected regions have neurochemical deficits of acetylcholine, glutamate, and serotonin. The degree of dementia correlates with the decline in these neurotransmitters in biopsy or autopsy tissue. While short-term memory deficits predominate early in the progressive dementias, the term dementia is reserved for a clinical syndrome of memory disorder plus at least one other cognitive realm: language, motor or procedural memory (praxis), spatial function, and executive function (decision-making, planning, overall control of behavior). As an etiology for dementia, AD accounts for about two-thirds of cases, with metabolic causes and Lewy body disease making up the lion’s share of the remaining third.
    [Show full text]
  • (12) United States Patent (10) Patent No.: US 8,039,464 B2 Schrattenholz (45) Date of Patent: Oct
    US008O39464B2 (12) United States Patent (10) Patent No.: US 8,039,464 B2 Schrattenholz (45) Date of Patent: Oct. 18, 2011 (54) MUSCARINIC ANTAGONISTS WITH PARP 5,576.436 A 11/1996 McCabe et al. AND SIRMODULATING ACTIVITY AS 5,620,978 A 4/1997 Cai et al. 5,705.499 A 1/1998 Cywin et al. AGENTS FOR INFLAMMATORY DISEASES 5,712,269 A 1/1998 McCabe et al. 5,716,952 A 2f1998 WoldeMussie et al. (75) Inventor: André Schrattenholz, Mainz (DE) 5,817,679 A 10, 1998 Shen et al. 5,935,781 A 8, 1999 Poirier 6,022,683 A 2/2000 Poirier (73) Assignee: ProteoSys AG, Mainz (DE) 6,060,473 A 5, 2000 Shen et al. 6,077,846 A 6/2000 Qian et al. (*) Notice: Subject to any disclaimer, the term of this 6,117,889 A 9, 2000 Shen et al. patent is extended or adjusted under 35 6.255.490 B1 7/2001 Shen et al. U.S.C. 154(b) by 0 days. 6,306,861 B1 10/2001 Leonardi et al. 6,316,423 B1 1 1/2001 Von Lubitz et al. 6,365,592 B1 4/2002 Leblanc et al. (21) Appl. No.: 11/632,648 6,403,584 B1 6/2002 de Laszlo et al. 6,403,594 B1 6/2002 Leonardi et al. (22) PCT Filed: Jul.18, 2005 6.410,583 B1 6/2002 Labelle et al. 6,486,163 B1 1 1/2002 Leonardi et al. (86). PCT No.: PCT/EP2005/007805 6,528,529 B1 3/2003 Brann et al.
    [Show full text]
  • United States Patent (10) Patent No.: US 8,003,632 B2 Tracey Et Al
    US008.003632B2 (12) United States Patent (10) Patent No.: US 8,003,632 B2 Tracey et al. (45) Date of Patent: Aug. 23, 2011 (54) CHOLINESTERASE INHIBITORS FOR Sg: A t 3. Ryaninomiya et et alal. TREATING INFLAMMATION 5,536,728 A 7/1996 Ninomiya et al. 5,554,780 A 9, 1996 Wolf (75) Inventors: Kevin J. Tracey, Old Greenwich, CT 5,693,668 A 12, 1997 Sain et al. (US); Valentin A. Pavlov, Ridgewood, 5,760,267 A 6/1998 Gandolfi et al. NY (US) 5,861.411 A 1/1999 Ninomiya et al. 5,929,084 A 7, 1999 Zhu et al. 5,981,549 A 11, 1999 V (73) Assignee: The Feinstein Institute for Medical 6, 194403 B1 2, 2001 E. al. Research, Manhasset, NY (US) 6,433,173 B1 8, 2002 Omori 6,610,713 B2 * 8/2003 Tracey .......................... 514,343 (*) Notice: Subject to any disclaimer, the term of this 6,838,471 B2 ck 1/2005 Tracey ded or adiusted under 35 2003. O139391 A1 7/2003 Parys et al. .............. 514,214.03 patent 1s extende 2004/0214.863 A1 10, 2004 Pratt U.S.C. 154(b) by 0 days. FOREIGN PATENT DOCUMENTS (21) Appl. No.: 11/893,116 EP O 633 251 A1 1/1995 JP 571 14512 7, 1982 (22) Filed: Aug. 14, 2007 WO WO98/OO119 A 1, 1998 9 WO WO99,08672 2, 1999 O O WO WOO1,89526 A 11, 2001 (65) Prior Publication Data WO WO 2004/034963 A 4, 2004 US 2008/OO70901 A1 Mar. 20, 2008 OTHER PUBLICATIONS Rees et al., Cardiovascular Research (1993)27:453-458.* Related U.S.
    [Show full text]
  • CUSTOMS TARIFF - SCHEDULE 99 - I
    CUSTOMS TARIFF - SCHEDULE 99 - i Chapter 99 SPECIAL CLASSIFICATION PROVISIONS - COMMERCIAL Notes. 1. The provisions of this Chapter are not subject to the rule of specificity in General Interpretative Rule 3 (a). 2. Goods which may be classified under the provisions of Chapter 99, if also eligible for classification under the provisions of Chapter 98, shall be classified in Chapter 98. 3. Goods may be classified under a tariff item in this Chapter and be entitled to the Most-Favoured-Nation Tariff or a preferential tariff rate of customs duty under this Chapter that applies to those goods according to the tariff treatment applicable to their country of origin only after classification under a tariff item in Chapters 1 to 97 has been determined and the conditions of any Chapter 99 provision and any applicable regulations or orders in relation thereto have been met. 4. The words and expressions used in this Chapter have the same meaning as in Chapters 1 to 97. Issued January 1, 2016 99 - 1 CUSTOMS TARIFF - SCHEDULE Tariff Unit of MFN Applicable SS Description of Goods Item Meas. Tariff Preferential Tariffs 9901.00.00 Articles and materials for use in the manufacture or repair of the Free CCCT, LDCT, GPT, UST, following to be employed in commercial fishing or the commercial MT, MUST, CIAT, CT, harvesting of marine plants: CRT, IT, NT, SLT, PT, COLT, JT, PAT, HNT, Artificial bait; KRT: Free Carapace measures; Cordage, fishing lines (including marlines), rope and twine, of a circumference not exceeding 38 mm; Devices for keeping nets open; Fish hooks; Fishing nets and netting; Jiggers; Line floats; Lobster traps; Lures; Marker buoys of any material excluding wood; Net floats; Scallop drag nets; Spat collectors and collector holders; Swivels.
    [Show full text]