DOCSLIB.ORG

Unexpected Binding of an Octapeptide to the Angiotensin II Receptor (Renin-Angiotensin System/Hypertension/Angiotensin Antagonist) R

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Unexpected Binding of an Octapeptide to the Angiotensin II Receptor (Renin-Angiotensin System/Hypertension/Angiotensin Antagonist) R Proc. Natl. Acad. Sci. USA Vol. 84, pp. 9219-9222, December 1987 Medical Sciences Unexpected binding of an octapeptide to the angiotensin II receptor (renin-angiotensin system/hypertension/angiotensin antagonist) R. L. SOFFER*, S. BANDYOPADHYAY*, E. ROSENBERG*, P. HOEPRICHt, A. TEITELBAUMt, T. BRUNCKt, C. B. COLBYt, AND C. GLOFFtt *Department of Medicine, Cornell University Medical College, New York, NY 10021; and tTriton Biosciences Inc., 1501 Harbor Bay Parkway, Alameda, CA 94501 Communicated by William A. Goddard III, August 17, 1987 (receivedfor review April 24, 1987) ABSTRACT An octapeptide, TBI-22 (Lys-Gly-Val-Tyr- MATERIALS AND METHODS Ile-His-Ala-Leu), inhibited binding of angiotensin II by a III-labeled angiotensin II ('l25-angiotensin II; 1400 puCi/nmol; solubilized angiotensin receptor partially purified from rabbit 1 Ci = 37 GBq) was purchased from New England Nuclear. liver. This inhibition appears to result from competition for t-Butoxycarbonyl (Boc) derivatives of amino acids were from binding to the same receptor. Radioiodinated TBI-22, like Peninsula Laboratories (Belmont, CA). Diisopropylcarbodii- angiotensin I, bound to the solubilized receptor with an mide and 1-hydroxybenzotriazole were products of Aldrich. affinity such that the binding was inhibited 50% by unlabeled Peptides were synthesized by the solid-phase procedure TBI-22 or angiotensin HI at nanomolar concentrations. The (12), using an automated synthesizer, model 9500 from binding reaction, like that for angiotensin HI, required p- Biosearch (San Rafael, CA). The first amino acid was chloromercuriphenylsulfonic acid and was reversed in the esterified to chloromethylated polystyrene-divinylbenzene presence of dithiothreitol. TBI-22 and angiotensin II share the copolymer (Bio-Rad; 1% cross-linked, 1.34 meq/g) as de- sequence Val-Tyr-Ile-His; this tetrapeptide alone, however, did scribed (13). Subsequent amino acids were coupled twice by not inhibit binding of angiotensin II. Replacement of the using diisopropylcarbodiimide. In general, the Boc group was tyrosine residue by aspartic acid in TBI-22 greatly reduced the removed by treatment of the resin containing protected ability of the peptide to compete with angiotensin H for peptide for 20 min with 45% trifl uoroacetic acid in dichloro- binding, suggesting an important contribution of this residue to methane (vol/vol) followed by two neutralizations of 5 min the configuration required for revognition by the receptor. with 5% diisopropylethylamine in dichloromethane (vol/ vol). The resin was washed with appropriate solvents before The is involved in the and after each step of deprotection, neutralization, and renin-angiotensin system regulation of coupling. Removal of the Boc group and completeness of blood pressure (1). Angiotensin II is the biologically active coupling were monitored qualitatively by a ninhydrin color component of this physiological system (2). Angiotensin II is test (14). All peptides were cleaved from the resin with a potent vasopressor agent as a consequence of its contrac- simultaneous removal ofthe protecting group by exposure to tive effect on blood vessels (2), its stimulation of aldosterone anhydrous hydrogen fluoride/10% anisole (vol/vol) for 40 secretion by the adrenal cortex (3), and its action on the min at 0-40C. Hydrogen fluoride and anisole were removed central nervous system (4). These and other effects of from the resin by evaporation under reduced pressure pro- angiotensin II are mediated by a specific receptor(s) on the vided by a water aspirator, followed by washing with anhy- surface of its target cells (reviewed in ref. 5). drous diethyl ether. The peptide was then obtained with Angiotensin-converting enzyme (reviewed in ref. 6) is a sequential washes ofdimethylformamide, 50% dimethylform- dipeptidyl carboxypeptidase that cleaves angiotensin I to amnide/10% acetic acid/40% water (vol/vol), 10% acetic angiotensin II. The use of drugs that inhibit angiotensin- acid/90% water (vol/vol), and distilled water. The combined converting enzyme (7, 8) has been shown to be an effective washings were finally lyophilized. antihypertensive therapy. Angiotensin-converting enzyme The peptides were then purified by reverse-phase high- possesses broad substrate specificity, and therefore its inhi- performance liquid chromatography. The various peptides bition is likely to influence the metabolism of peptides were chromatographed on a preparative column (Whatman; unrelated to the renin-angiotensin system. An antagonist Partisil 10 ODS-3 Magnum 20, 2.2 x 50 cm); the elution was targeted at the angiotensin receptor might represent a more 65 min and the gradient ranged from 83% solvent A (0.05% physiologically specific agent for antihypertensive therapy. trifluoroacetic acid/water) and 17% solvent B (0.05% triflu- Analogs of angiotensin II with substitutions of the carboxyl- oroacetic acid/acetonitrile) to 50% A and 50% B. A portion terminal residue have shown promise in this regard (9-11). of each purified peptide was hydrolyzed in 5.7 M HCO and its However, jt may be that identification of determinants amino acid composition was determined (15). actually recognized by the receptor will be necessary to TBI-22 was radiolabeled by a modification of the method expedite development of the desired antagonist. To obtain of Greenwood et al. (16). The peptide (10 nmol) was incu- this type of information, examination of the binding of bated at 20'C for 30 sec with 4 mCi of Na125I (Amersham) in putative ligands by isolated receptor may be useful. To date, 70 Al of0.5 M potassium phosphate buffer, pH 7.4, containing there have been few such studies, and virtually all of them 10 ,ug of chloramine T. The reaction was stopped by addition have been carried out with intact membrane fractions (5). of 1 mg of 2-mercaptoethanolamine (Sigma) followed by 0.5 Here, we report the binding of an octapeptide and some ofits ml of0.1 M potassium phosphate buffer, pH 7.4. The solution analogs and derivatives to a solubilized angiotensin II recep- was then transferred to a C8 Sep-pak column (J. T. Baker tor partially purified from rabbit liver. Chemical, Phillipsburg, NJ) and unincorporated 1251 was washed off with 5 ml of 0.1 M phosphate buffer. Radioiodi- The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" Abbreviation: PCMS, p-chloromercuriphenylsulfonic acid. in accordance with 18 U.S.C. §1734 solely to indicate this fact. fTo whom reprint requests should be addressed. 9219 9220 Medical Sciences: Soffer et al. Proc. Natl. Acad. Sci. USA 84 (1987) nated TBI-22 (125I-TBI-22; 120 ,uCi/nmol) was eluted with Table 1. Binding of radioiodinated TBI-22 by solubilized glacial acetic acid and lyophilized. angiotensin II receptor The angiotensin II receptor was isolated as described (17) Binding, Residual binding, except that the SE-Sephadex and heat treatment steps were System cpm % replaced by chromatography on DEAE-cellulose. The DEAE- cellulose was equilibrated in 20 mM Tris-HCl, pH 7.6, Complete 32,764 100 containing 1 mM K2EDTA and 0.5% Brij 99 (Emulsion + 10 ,uM TBI-22 212 1 Engineering, Sanford, FL). After application of protein (10 + 10,M angiotensin II 290 1 mg/ml of bed volume), the column was washed with 20 bed - PCMS 596 2 volumes of a linear gradient ranging from 0.0 to 0.3 M KCI. + 200 ,uM dithiothreitol* 1,162 4 The receptor emerged as a symmetrical peak at about 0.13 M The complete system is described in Materials and Methods. The KCl. The active fractions were pooled and dialyzed against value obtained in the absence of receptor (277 cpm) has been equilibration buffer. The binding activity of the partially subtracted from all data. purified receptor, estimated by Scatchard analysis, was 18 *Added after standard 60-min incubation and reaction mixture pmol of angiotensin II per mg of protein. processed after additional 3 min. Reaction mixtures (150 ,u1) for binding assays contained 30 mM Tris-HCl at pH 7.6, 2.5 mM K2EDTA, 0.2 mM p- II to the partially purified receptor in a dose-dependent chloromercuriphenylsulfonic acid (PCMS), 100 ,ug of bovine manner (Fig. lA). The IC50 is approximately 8 nM. By serum albumin, 30 ,ug of receptor preparation, 0.25% Brij 99, comparison, unlabeled angiotensin II has an IC50 equal to 12 and 100,000 cpm of 125I-angiotensin II (0.25 nM) or 115I-TBI-22 nM in the same system. To determine the biochemical (3 nM). After incubation at 20°C for 60 min, bound radioactive mechanism for this effect ofTBI-22, a radioiodinated sample peptide was estimated as described (18) except that 0.05% of this octapeptide was prepared and its binding by the dextran/0.5% charcoal was employed. ICso values (concentra- receptor preparation was examined (Table 1). The receptor- tion yielding 50% inhibition) for competing unlabeled peptides dependent binding was completely blocked by 10 ,uM un- were estimated from inhibition dose-response curves com- labeled TBI-22 or arigiotensin II. The binding reaction re- posed of at least four points. Results were corrected for quired the presence of PCMS, and the receptor-ligand "nonspecific" binding-i.e., cpm bound in the presence of 10 complex was dissociated by dithiothreitol. These unusual ,uM unlabeled angiotensin II, which was essentially identical to characteristics are identical to those we previously described that measured in the absence of receptor and less than 2% of for the binding of angiotensin II to this solubilized rabbit that obtained with the complete system. Peptides that did not hepatic receptor (17). The data on binding of radioiodinated inhibit at 10 p.M were not further examined. TBI-22 thus provide strong evidence that this peptide is Rabbit antiserum to angiotensin II was developed as bound by the angiotensin II receptor and that it inhibits the described by Gocke et al. (19).
Load more

Recommended publications
  • ACE2–Angiotensin-(1–7)–Mas Axis and Oxidative Stress in Cardiovascular Disease
    Hypertension Research (2011) 34, 154–160 & 2011 The Japanese Society of Hypertension All rights reserved 0916-9636/11 $32.00 www.nature.com/hr REVIEW SERIES ACE2–angiotensin-(1–7)–Mas axis and oxidative stress in cardiovascular disease Luiza A Rabelo1,2, Natalia Alenina1 and Michael Bader1 The renin–angiotensin–aldosterone system (RAAS) is a pivotal regulator of physiological homeostasis and diseases of the cardiovascular system. Recently, new factors have been discovered, such as angiotensin-converting enzyme 2 (ACE2), angiotensin-(1–7) and Mas. This newly defined ACE2–angiotensin-(1–7)–Mas axis was shown to have a critical role in the vasculature and in the heart, exerting mainly protective effects. One important mechanism of the classic and the new RAAS regulate vascular function is through the regulation of redox signaling. Angiotensin II is a classic prooxidant peptide that increases superoxide production through the activation of NAD(P)H oxidases. This review summarizes the current knowledge about the ACE2–angiotensin-(1–7)–Mas axis and redox signaling in the context of cardiovascular regulation and disease. By interacting with its receptor Mas, angiotensin-(1–7) induces the release of nitric oxide from endothelial cells and thereby counteracts the effects of angiotensin II. ACE2 converts angiotensin II to angiotensin-(1–7) and, thus, is a pivotal regulator of the local effects of the RAAS on the vessel wall. Taken together, the ACE2–angiotensin-(1–7)–Mas axis emerges as a novel therapeutic target in the context of cardiovascular
    [Show full text]
  • New Components of the Renin-Angiotensin System: Alamandine and the Mas-Related G Protein-Coupled Receptor D
    Curr Hypertens Rep (2014) 16:433 DOI 10.1007/s11906-014-0433-0 HYPERTENSION AND THE KIDNEY (R CAREY, SECTION EDITOR) New Components of the Renin-Angiotensin System: Alamandine and the Mas-Related G Protein-Coupled Receptor D Gisele Maia Etelvino & Antônio Augusto Bastos Peluso & Robson Augusto Souza Santos Published online: 24 April 2014 # Springer Science+Business Media New York 2014 Abstract The renin-angiotensin system is an important com- this novel component of the RAS opens new venues for ponent of the central and humoral mechanisms of blood understanding its physiological role and opens new putative pressure and hydro-electrolytic balance control. Angiotensin therapeutic possibilities for treating cardiovascular diseases. II is a key player of this system. Twenty-five years ago the first manuscripts describing the formation and actions of another Keywords Mas1 . Angiotensin II . Angiotensin-(1-7) . peptide of the RAS, angiotensin-(1-7), were published. Since Hypertension . Angiotensin-(1-12) . Angiotensin A then several publications have shown that angiotensin-(1-7) is as pleiotropic as angiotensin II, influencing the functions of many organs and systems. The identification of the ACE Introduction homologue ACE2 and, a few years later, Mas, as a receptor for angiotensin-(1-7) contributed a great deal to establish this The renin angiotensin system (RAS) has been described as an peptide as a key player of the RAS. Most of the actions of endocrine and tissular system involved in cardiovascular and angiotensin-(1-7) are opposite to those described for angio- renal control. Classically, it is comprised by angiotensinogen, tensin II. This has led to the concept of two arms of the RAS: an α-2-globulin produced mainly by the liver, which is one comprising ACE/AngII/AT1R and the other cleaved in the amino-terminal portion by renin, an aspartyl ACE2/Ang-(1-7)/Mas.
    [Show full text]
  • Angiotensin-I-Converting Enzyme and Prolyl Endopeptidase Inhibitory Peptides from Marine Processing By-Products
    instlttiild lctt«rkenny Talcneolaiochta institute Lyit Lelttr Ccanalnn of Technology Angiotensin-I-converting enzyme and prolyl endopeptidase inhibitory peptides from marine processing by-products Julia Wilson Supervisor: Dr. B. Camey, Letterkenny Institute of Technology External Supervisor: Dr, M. Hayes, Teagasc. Ash town, Dublin Submitted to the Higher Education and Training Awards Council in fulfilment of the requirements for the degree of Master of Science by research. Table of Contents Declaration 3 Abstract 4 List of Abbreviations 6 List of Figures 8 List of Tables 10 Publications 11 Acknowledgements 12 Chapter 1: Literature review 1.1 Introduction A 13 1.2 Mackerel and Whelk life history and habitats 14 1.3 Function of ACE-I and PEP inhibitory peptides 16 1.4 Sources of ACE-I and PEP inhibitory peptides 20 1.5 Derivatisation of ACE-I and PEP inhibitory peptides 22 1.5.1 Principles of capillary electrophoresis (CE) 28 1.5.2 Principles of high performance liquid chromatography (HPLC) 31 1.5.3 Principles of mass spectrometry (MS) 33 1.6 Structural properties involved in ACE-I and PEP inhibitory 33 activities of peptides 1.7 Bioactive peptides as functional foods 35 1.7.1 Survival of bioactive peptide inhibitors in vivo 36 1.8 Aims and objectives 38 Chapter 2: Materials and Methods 2.1 General materials and methods 40 2.1.1 Chemicals and reagents 40 2.1.2 Buffer preparation 41 2.1.3 Bradford protein assay 42 1 2.2 Enzyme hydrolytic studies 43 2.2.1 Sample pre-treatments 43 2.2.2 Hydrolytic enzymes and hydrolytic reactions 45 2.3 Colorimetric
    [Show full text]
  • Amino Acid Side Chain Conformation in Angiotensin II and Analogs
    Proc. Nati. Acad. Sci. USA Vol. 77, No. 1, pp. 82-86, January 1980 Biochemistry Amino acid side chain conformation in angiotensin II and analogs: Correlated results of circular dichroism and 1H nuclear magnetic resonance (peptide hormones/competitive inhibitor/N-methylation/pH titration) F. PIRIou*, K. LINTNER*, S. FERMANDJIAN*, P. FROMAGEOT*, M. C. KHOSLAt, R. R. SMEBYt, AND F. M. BUMPUSt *Service de Biochimie, D)partement de Biologie, Centre d'Etudes Nucl&aires de Saclay, P.B. No. 2, F-91190 Gif-sur-Yvette, France; and tThe Clinic Center, Cleveland Clinic, 9500 Euclid Avenue, Cleveland, Ohio 44106 Communicated by Irvine H. Page, September 4,1979 ABSTRACT [1-Sarcosine,S-isoleucinelangiotensin II (Sar- [Sarl,11e8]Angiotensin II was shown to be a potent antagonist Arg-Val-Tyr-Ile-His-Pro-Ile) has been shown to be a potent an- of the pressor action of antiotensin II (Asp-Arg-Val-Tyr-Ile- tagonist of the pressor action of angiotensin II. With a view to increase half-life in vivo of this peptide, the amino acid residue His-Pro-Phe). However, this and other similar antagonistic at position 4 (tyrosine) or position 5 (isoleucine) was replaced peptides have short half-lives in vivo; the shortness is presum- with the corresponding N-methylated residue. This change ably due to rapid degradation by peptidases (for detailed re- drastically reduced the antagonistic properties of this analog. views, see refs. 1 and 2). With a view to render these peptides The present work was therefore undertaken to investigate the more resistant to enzymatic degradation, the residue at position effect of N-methylation on overall conformation of these pep- tides and to determine the conformational requirements for 4 (tyrosine) or position 5 (isoleucine) was replaced with the maximum agonistic or antagonistic pro rties.
    [Show full text]
  • An Update on the Tissue Renin Angiotensin System and Its Role in Physiology and Pathology
    Journal of Cardiovascular Development and Disease Review An Update on the Tissue Renin Angiotensin System and Its Role in Physiology and Pathology Ali Nehme 1 , Fouad A. Zouein 2 , Zeinab Deris Zayeri 3 and Kazem Zibara 4,* 1 EA4173, Functional genomics of arterial hypertension, Univeristy Claude Bernard Lyon-1 (UCBL-1), 69008 Lyon, France; [email protected] 2 Department of Pharmacology and Toxicology, Heart Repair Division, Faculty of Medicine, American University of Beirut, Beirut 11-0236, Lebanon; [email protected] 3 Thalassemia & Hemoglobinopathy Research Center, Health Research Institute, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran; [email protected] 4 PRASE, Biology Department, Faculty of Sciences-I, Lebanese University, Beirut, Lebanon * Correspondence: [email protected] Received: 10 February 2019; Accepted: 26 March 2019; Published: 29 March 2019 Abstract: In its classical view, the renin angiotensin system (RAS) was defined as an endocrine system involved in blood pressure regulation and body electrolyte balance. However, the emerging concept of tissue RAS, along with the discovery of new RAS components, increased the physiological and clinical relevance of the system. Indeed, RAS has been shown to be expressed in various tissues where alterations in its expression were shown to be involved in multiple diseases including atherosclerosis, cardiac hypertrophy, type 2 diabetes (T2D) and renal fibrosis. In this chapter, we describe the new components of RAS, their tissue-specific expression, and their alterations under pathological conditions, which will help achieve more tissue- and condition-specific treatments. Keywords: renin-angiotensin-aldosterone system; tissue; expression; physiology 1. Introduction In its classical view, RAS was defined as an endocrine system involved in blood pressure regulation and body electrolyte balance.
    [Show full text]
  • Angiotensin-Converting Enzyme 2- and Prolyl Carboxypeptidase-Independent Conversion of Angiotensin II to Angiotensin-(1-7) in Circulation and Peripheral Tissues
    Aus dem Max-Delbrück-Centrum für Molekulare Medizin sowie der Abteilung für Nephrologie der Feinberg School of Medicine der Northwestern University DISSERTATION Angiotensin-Converting Enzyme 2- and Prolyl Carboxypeptidase-Independent Conversion of Angiotensin II to Angiotensin-(1-7) in Circulation and Peripheral Tissues zur Erlangung des akademischen Grades Doctor medicinae (Dr. med.) vorgelegt der Medizinischen Fakultät Charité – Universitätsmedizin Berlin von Peter Daniel Serfözö aus Budapest, Ungarn Datum der Promotion: 18.09.2020 Table of contents 1. List of abbreviations ........................................................................................................... 3 2. Abstract (Deutsch) .............................................................................................................. 4 3. Abstract (English) .............................................................................................................. 5 4. Synopsis ............................................................................................................................. 6 Introduction .................................................................................................................... 6 Methods ........................................................................................................................ 10 Results .......................................................................................................................... 14 Discussion ....................................................................................................................
    [Show full text]
  • Histidine in Health and Disease: Metabolism, Physiological Importance, and Use As a Supplement
    nutrients Review Histidine in Health and Disease: Metabolism, Physiological Importance, and Use as a Supplement Milan Holeˇcek Department of Physiology, Faculty of Medicine in Hradec Králové, Charles University, Šimkova 870, 500 38 Hradec Kralove, Czech Republic; [email protected] Received: 7 February 2020; Accepted: 20 March 2020; Published: 22 March 2020 Abstract: L-histidine (HIS) is an essential amino acid with unique roles in proton buffering, metal ion chelation, scavenging of reactive oxygen and nitrogen species, erythropoiesis, and the histaminergic system. Several HIS-rich proteins (e.g., haemoproteins, HIS-rich glycoproteins, histatins, HIS-rich calcium-binding protein, and filaggrin), HIS-containing dipeptides (particularly carnosine), and methyl- and sulphur-containing derivatives of HIS (3-methylhistidine, 1-methylhistidine, and ergothioneine) have specific functions. The unique chemical properties and physiological functions are the basis of the theoretical rationale to suggest HIS supplementation in a wide range of conditions. Several decades of experience have confirmed the effectiveness of HIS as a component of solutions used for organ preservation and myocardial protection in cardiac surgery. Further studies are needed to elucidate the effects of HIS supplementation on neurological disorders, atopic dermatitis, metabolic syndrome, diabetes, uraemic anaemia, ulcers, inflammatory bowel diseases, malignancies, and muscle performance during strenuous exercise. Signs of toxicity, mutagenic activity, and allergic reactions or peptic ulcers have not been reported, although HIS is a histamine precursor. Of concern should be findings of hepatic enlargement and increases in ammonia and glutamine and of decrease in branched-chain amino acids (valine, leucine, and isoleucine) in blood plasma indicating that HIS supplementation is inappropriate in patients with liver disease.
    [Show full text]
  • Dependent and ACE2 (Angiotensin-Converting Enzyme 2)
    Renin-Angiotensin System Ang II (Angiotensin II) Conversion to Angiotensin-(1-7) in the Circulation Is POP (Prolyloligopeptidase)-Dependent and ACE2 (Angiotensin-Converting Enzyme 2)-Independent Peter Serfozo, Jan Wysocki, Gvantca Gulua, Arndt Schulze, Minghao Ye, Pan Liu, Jing Jin, Michael Bader, Timo Myöhänen, J. Arturo García-Horsman, Daniel Batlle Abstract—The Ang II (Angiotensin II)-Angiotensin-(1-7) axis of the Renin Angiotensin System encompasses 3 enzymes that form Angiotensin-(1-7) [Ang-(1-7)] directly from Ang II: ACE2 (angiotensin-converting enzyme 2), PRCP (prolylcarboxypeptidase), and POP (prolyloligopeptidase). We investigated their relative contribution to Ang- (1-7) formation in vivo and also ex vivo in serum, lungs, and kidneys using models of genetic ablation coupled with pharmacological inhibitors. In wild-type (WT) mice, infusion of Ang II resulted in a rapid increase of plasma Ang-(1-7). In ACE2−/−/PRCP−/− mice, Ang II infusion resulted in a similar increase in Ang-(1-7) as in WT (563±48 versus 537±70 fmol/mL, respectively), showing that the bulk of Ang-(1-7) formation in circulation is essentially independent of ACE2 and PRCP. By contrast, a POP inhibitor, Z-Pro-Prolinal reduced the rise in plasma Ang-(1-7) after infusing Ang II to control WT mice. In POP−/− mice, the increase in Ang-(1-7) was also blunted as compared with WT mice (309±46 and 472±28 fmol/mL, respectively P=0.01), and moreover, the rate of recovery from acute Ang II-induced hypertension was delayed (P=0.016). In ex vivo studies, POP inhibition with ZZP reduced Ang-(1-7) formation from Ang II markedly in serum and in lung lysates.
    [Show full text]
  • Angiotensin II
    Proc. Nati Acad. Sci. USA Vol. 78, No. 2, pp. 757-760, February 1981 Biochemistry Synthesis of [a-methyltyrosine-4]angiotensin II: Studies of its conformation, pressor activity, and mode of enzymatic degradation (angiotensin II analog/resistance to a-chymotrypsin degradation/full pressor activity/NMR and circular dichroism spectra similar to those of angiotensin II) M. C. KHOSLA*, K. STACHOWIAK, R. R. SMEBY*, F. M. BUMPUS*, F. PIRIOUt, K. LINTNERt, AND S. FERMANDJIANt *Research Division, The Cleveland Clinic Foundation, Cleveland, Ohio 44106; and tService de Biochimie, Department de Biologie, Centre d'Etudes Nucleaires de Saclay, P. B. No. 2, F-91190 Gif-sur-Yvette, France Communicated by Irvine H. Page, October 15, 1980 ABSTRACT Modifications in angiotensin II and its antago- full biological activity (agonist or antagonist), the backbone and nistic peptides that should have increased in vivo half-lives but not side-chain structure of the analog should resemble that of the reduced biological activity were studied by determining the effect hormone, All (2, 4). These observations may be helpful in the of a-methylation of the tyrosine in position 4. [a-Methyltyrosine- 4]angiotensin II, synthesized by the solid-phase procedure, design of new potent analogs. showed 92.6 ± 5.3% pressor activity of angiotensin H. Incubation with a-chymotrypsin for 1 hr indicated absence of degradation EXPERIMENTAL although, under the same conditions, angiotensin H was com- O-Benzyl-a-Methyl-L-Tyrosine. The procedure used for the pletely degraded to two components. Comparison of the 'H NMR synthesis ofthis compound is a modification ofthe one used by spectra in aqueous solution and the circular dichroism spectra in trifluoroethanol of angiotensin II and [a-methyltyrosine- Khosla et aL (2) for the synthesis of 0-(2,6-dichlorobenzyl)-L- 4]angiotensin II suggested that a methylation of the tyrosine res- tyrosine.
    [Show full text]
  • 209360Orig1s000
    CENTER FOR DRUG EVALUATION AND RESEARCH APPLICATION NUMBER: 209360Orig1s000 CLINICAL PHARMACOLOGY AND BIOPHARMACEUTICS REVIEW(S) Office of Clinical Pharmacology Review NDA Number 209360 Link to EDR \\cdsesub1\evsprod\nda209360 Submission Date 06-29-2017 Submission Type Priority review Brand Name (b) (4) Generic Name Angiotensin II Dosage Form and Strengths Solution for injection in normal saline (0.9% sodium chloride); 2.5 mg/vial (2.5 mg/mL) and 5 mg/vial (2.5 mg/mL) Route of Administration Intravenous infusion Proposed Indication Treatment of hypotension in adults with distributive or vasodilatory shock who remain hypotensive despite fluid and vasopressor therapy Applicant La Jolla Pharmaceutical Company Associated INDs IND (b) (4) and IND 122708 OCP Review Team Venkateswaran Chithambaram Pillai, MS(Pharm), PhD; Sudharshan Hariharan PhD; 1 Reference ID: 4188162 Table of Contents 1. EXECUTIVE SUMMARY..............................................................................................................................3 1.1 Recommendations..............................................................................................................................3 1.2 Post-Marketing Requirements and Commitments.............................................................................3 2. SUMMARY OF CLINICAL PHARMACOLOGY FINDINGS ..............................................................................4 2.1 Clinical Pharmacokinetics ...................................................................................................................4
    [Show full text]
  • Effect of UVC Irradiation on the Oxidation of Histidine In
    www.nature.com/scientificreports OPEN Efect of UVC Irradiation on the Oxidation of Histidine in Monoclonal Antibodies Yuya Miyahara 1, Koya Shintani2, Kayoko Hayashihara-Kakuhou2, Takehiro Zukawa3, Yukihiro Morita3, Takashi Nakazawa 4, Takuya Yoshida 1, Tadayasu Ohkubo1* & Susumu Uchiyama 2* We oxidized histidine residues in monoclonal antibody drugs of immunoglobulin gamma 1 (IgG1) using ultraviolet C irradiation (UVC: 200–280 nm), which is known to be potent for sterilization or disinfection. Among the reaction products, we identifed asparagine and aspartic acid by mass spectrometry. 18 18 In the photo-induced oxidation of histidine in angiotensin II, O atoms from H2 O in the solvent were incorporated only into aspartic acid but not into asparagine. This suggests that UVC irradiation generates singlet oxygen and induces [2 + 2] cycloaddition to form a dioxetane involving the imidazole Cγ − Cδ2 bond of histidine, followed by ring-opening in the manner of further photo-induced retro [2 + 2] cycloaddition. This yields an equilibrium mixture of two keto-imines, which can be the precursors to aspartic acid and asparagine. The photo-oxidation appears to occur preferentially for histidine residues with lower pKa values in IgG1. We thus conclude that the damage due to UVC photo-oxidation of histidine residues can be avoided in acidic conditions where the imidazole ring is protonated. Terapeutic antibodies are widely used in the treatment of diseases such as cancers and immunological disorders. Because the degradation of protein-based drugs during storage is inevitable, it is necessary to establish methods to characterize the degradation process and to assess the safety of the drugs1.
    [Show full text]
  • Targeting the Renin−Angiotensin Signaling Pathway in COVID-19: Unanswered Questions, Opportunities, and Challenges Krishna Srirama, Rohit Loombab, and Paul A
    PERSPECTIVE PERSPECTIVE Targeting the renin−angiotensin signaling pathway in COVID-19: Unanswered questions, opportunities, and challenges Krishna Srirama, Rohit Loombab, and Paul A. Insela,b,1 Edited by Susan G. Amara, National Institutes of Health, Bethesda, MD, and approved September 2, 2020 (received for review June 29, 2020) The role of the renin−angiotensin signaling (RAS) pathway in COVID-19 has received much attention. A central mechanism for COVID-19 pathophysiology has been proposed: imbalance of angiotensin convert- ing enzymes (ACE)1 and ACE2 (ACE2 being the severe acute respiratory syndrome coronavirus 2 [SARS-CoV-2] virus “receptor”) that results in tissue injury from angiotensin II (Ang II)-mediated signaling. This mechanism provides a rationale for multiple therapeutic approaches. In parallel, clinical data from retrospective analysis of COVID-19 cohorts has revealed that ACE inhibitors (ACEIs) or angiotensin recep- tor blockers (ARBs) may be beneficial in COVID-19. These findings have led to the initiation of clinical trials using approved drugs that target the generation (ACEIs) and actions (ARBs) of Ang II. However, treatment of COVID-19 with ACEIs/ARBs poses several challenges. These include choosing appropriate inclusion and exclusion criteria, dose optimization, risk of adverse effects and drug interactions, and verification of target engagement. Other approaches related to the RAS pathway might be considered, for example, inhalational administration of ACEIs/ARBs (to deliver drugs directly to the lungs) and use of compounds with other actions (e.g., activation of ACE2, agonism of MAS1 receptors, β-arrestin−based Angiotensin receptor agonists, and administration of soluble ACE2 or ACE2 peptides). Studies with animal models could test such approaches and assess therapeutic benefit.
    [Show full text]