WO2004099245A1 - Interferon-$g(y) stabilise - Google Patents

Interferon-$g(y) stabilise Download PDF

Info

Publication number
WO2004099245A1
WO2004099245A1 PCT/EP2004/004677 EP2004004677W WO2004099245A1 WO 2004099245 A1 WO2004099245 A1 WO 2004099245A1 EP 2004004677 W EP2004004677 W EP 2004004677W WO 2004099245 A1 WO2004099245 A1 WO 2004099245A1
Authority
WO
WIPO (PCT)
Prior art keywords
interferon
amino acid
polyethylene glycol
homologous
cysteine
Prior art date
Application number
PCT/EP2004/004677
Other languages
German (de)
English (en)
Inventor
Herwig Brunner
Hayssam Zakaria
Bernd Otto
Tobias Thomas
Florian Battermann
Marco Kresin
Andreas Busche
Christian Schmalz
Original Assignee
Fraunhofer-Gesellschaft zur Förderung der angewandten Forschung e.V.
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Fraunhofer-Gesellschaft zur Förderung der angewandten Forschung e.V. filed Critical Fraunhofer-Gesellschaft zur Förderung der angewandten Forschung e.V.
Publication of WO2004099245A1 publication Critical patent/WO2004099245A1/fr

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/52—Cytokines; Lymphokines; Interferons
    • C07K14/555—Interferons [IFN]
    • C07K14/57—IFN-gamma

Definitions

  • the present invention relates to a stabilized interferon- ⁇ , in particular to a recombinantly produced, human interferon- ⁇ .
  • Human interferon- ⁇ is genetically derived from bacterial cells and is already used in two different indications. Clinical trials on pulmonary fibrosis patients are also so successful that therapy success and therefore a large market can generally be expected for fibrosis diseases.
  • Natural interferon- ⁇ is glycosylated twice. In the case of genetically engineered interferon- ⁇ , however, this glycosylation is absent in the production by means of bacterial systems. Although the biological activity in in vitro systems is not impaired by recombinant interferon- ⁇ , the unmodified interferon- ⁇ is pharmacokinetically very much less
  • glycosylated interferon- ⁇ is very complex and costly.
  • the interferon- ⁇ according to the invention is now modified by means of polyethylene glycol (PEG).
  • PEG polyethylene glycol
  • the polyethylene glycol is coupled to certain individual amino acids of an interferon- ⁇ .
  • Such coupling can take place via an amino or thiol group.
  • the amino acids cysteine, asparagine, glutamine, lysine, arginine and / or histidine are therefore particularly suitable for this purpose.
  • the interferon- ⁇ to be modified can be a recombinantly produced interferon- ⁇ , in particular human interferon- ⁇ .
  • Interferon- ⁇ is understood here to mean a protein which has an amino acid sequence which is essentially homologous to the native, in particular human, interferon- ⁇ .
  • the amino acid sequence can be one or more amino acid exchanges, deletions, shortenings at their ends or extensions at their ends have as long as the biological function of an interferon- ⁇ , in particular its pharmacological effects, are maintained.
  • interferon- ⁇ advantageously have the following sequences or are homologous to them:
  • IFN- ⁇ D63C in which aspartic acid D63 is replaced by cysteine and PEGylated:
  • IFN- ⁇ S99C in which serine S99 is replaced by cysteine and PEGylated:
  • IFN- ⁇ 7/69 D63C in which aspartic acid D63 has been replaced by cysteine and PEGylated and amino acids 7 and 69 have also been replaced by cysteine and bridged with a disulfide bridge:
  • IFN- ⁇ 7/69 S99C in which serine 99 is exchanged for cysteine and PEGylated, and furthermore amino acids 7 and 69 are exchanged for cysteine and bridged with a disulfide bridge:
  • Interferon- ⁇ contains many lysines and is strongly basic. It can therefore only be heterogeneously amino-PEGylated. PEGylation is therefore advantageously carried out via the thiol groups of free cysteines.
  • the amino acids aspartic acid 63 or serine 99 were exchanged for cysteines and the corresponding variants were produced by genetic engineering. Polyethylene glycol was then coupled to the thiol groups of these cysteines. It was found that these variants in both PEGylated and non-PEGy- form, regardless of the chain length of the polyethylene glycol, had largely the same biological activity as unchanged and unmodified interferon- ⁇ .
  • the interferon- ⁇ modified with polyethylene glycol is pharmacokinetically much more stable ' .
  • a PEG can be used which has a molecular weight between 5,000 mol and 40,000 Da.
  • interferon- ⁇ In addition to native interferon- ⁇ , various modified variants of interferon- ⁇ can also be used, for example with or without a disulfide bridge.
  • FIG. 1 shows the course of the activity of various forms of interferon- ⁇ and their decay over time in an animal experiment.
  • the solid line denotes human recombinant interferon- ⁇ that has not been PEGylated. This shows a high activity at the beginning (logarithmic scale of FIG. 1!). This activity then quickly subsides within a few hours.
  • the other three curves show human recombinant interferon- ⁇ which has been modified with various polyethylene glycols.
  • the modification was carried out, as indicated in the legend of FIG. 1, with polyethylene glycol with a molecular weight of 5,000 Da, 20,000 Da and 40,000 Da, respectively.
  • IFN- ⁇ 7/69 was used as the non-PEGylated Ifn- ⁇ and IFN- ⁇ 7/69 S99C was used as the PEGylated IFN- ⁇ , in which the amino acid serine 99 was replaced by cysteine and PEGylated.
  • a blood sample was taken retrobulously, a serum preparation was carried out and the IFN- ⁇ - Serum levels determined using an antiviral assay.

Landscapes

  • Chemical & Material Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Organic Chemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Biochemistry (AREA)
  • Biophysics (AREA)
  • Zoology (AREA)
  • Genetics & Genomics (AREA)
  • Medicinal Chemistry (AREA)
  • Molecular Biology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Toxicology (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Peptides Or Proteins (AREA)

Abstract

La présente invention concerne un interféron-Ω stabilisé. L'interféron-Ω selon l'invention est modifié sur un ou plusieurs de ses acides aminés comportant des groupes latéraux thiol ou amino, au moyen de polyéthylène glycol. Les acides aminés cystéine, asparagine, glutamine, lysine, arginine et/ou histidine sont particulièrement adaptés à une telle modification.
PCT/EP2004/004677 2003-05-05 2004-05-03 Interferon-$g(y) stabilise WO2004099245A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
DE10320223 2003-05-05
DE10320223.4 2003-05-05

Publications (1)

Publication Number Publication Date
WO2004099245A1 true WO2004099245A1 (fr) 2004-11-18

Family

ID=33426684

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/EP2004/004677 WO2004099245A1 (fr) 2003-05-05 2004-05-03 Interferon-$g(y) stabilise

Country Status (1)

Country Link
WO (1) WO2004099245A1 (fr)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EA020220B1 (ru) * 2011-12-21 2014-09-30 Общество С Ограниченной Ответственностью "Форт" (Ооо "Форт") Ковалентный конъюгат полиэтиленгликоля с полипептидом, имеющим активность интерферона-гамма

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1991001758A1 (fr) * 1989-08-07 1991-02-21 Debiopharm S.A. Derives polymeres de subtances medicamenteuses biologiquement actifs
US5567422A (en) * 1993-02-02 1996-10-22 Enzon, Inc. Azlactone activated polyalkylene oxides conjugated to biologically active nucleophiles
WO1999003887A1 (fr) * 1997-07-14 1999-01-28 Bolder Biotechnology, Inc. Derives d'hormone de croissance et proteines associees
WO2001036001A2 (fr) * 1999-11-12 2001-05-25 Maxygen Holdings Ltd Conjugues d'interferon gamma

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1991001758A1 (fr) * 1989-08-07 1991-02-21 Debiopharm S.A. Derives polymeres de subtances medicamenteuses biologiquement actifs
US5567422A (en) * 1993-02-02 1996-10-22 Enzon, Inc. Azlactone activated polyalkylene oxides conjugated to biologically active nucleophiles
WO1999003887A1 (fr) * 1997-07-14 1999-01-28 Bolder Biotechnology, Inc. Derives d'hormone de croissance et proteines associees
WO2001036001A2 (fr) * 1999-11-12 2001-05-25 Maxygen Holdings Ltd Conjugues d'interferon gamma

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
TANG WEI ET AL: "Studies on the PEGylation of protein at a specific site: sulfhydryl-PEGylation of 97 Cys-IFN-.gamma", 1997, STN CAPLUS, XP002937792 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EA020220B1 (ru) * 2011-12-21 2014-09-30 Общество С Ограниченной Ответственностью "Форт" (Ооо "Форт") Ковалентный конъюгат полиэтиленгликоля с полипептидом, имеющим активность интерферона-гамма

Similar Documents

Publication Publication Date Title
DE69935345T3 (de) Methoden und zusammensetzungen zur prävention und behandlung der anämie
EP1161452B1 (fr) Dimeres d'insuline pontes de maniere covalente
DE3650101T2 (de) Analoga von Insulin und deren Herstellungsmethode.
DE69034179T2 (de) Expressionssysteme für ein amidierendes Enzym
DE69110101T2 (de) GIP-Analoge und deren Verwendung.
DE3650055T2 (de) Fibroblast-wachstumsfaktor.
DE3689845T2 (de) Rekombinanter fibroblast-wachstumsfaktor.
DE68912777T2 (de) Insulin-analoge.
DE60224415T2 (de) Pegyliertes und diglykosyliertes erythropoietin
DE60117919T2 (de) Fusionsprotein mit verbesserter in vivo erythropoietinwirkung
EP0885961A1 (fr) Nouveau dérivés de l'insuline avec déclenchement d'action rapide
EP0769020B1 (fr) Nouvelles proteines mutantes de l'il-4 humaine utilisees comme antagonistes ou agonistes partiels de l'interleukine humaine 4
DE69635675T2 (de) Verkürzter aus einer gliazelllinie gewonnener neurotropher faktor
DE102008003566A1 (de) Neue Insulinderivate mit extrem verzögertem Zeit-/ Wirkungsprofil
DE69434909T2 (de) Herstellung von humaninsulin
DE3780179T2 (de) Herstellung von menschlichem erythropoietin.
Gottschall et al. Interleukin-1 beta is more potent than interleukin-1 alpha in supressing follicle-stimulating hormone-induced differentiation of ovarian granulosa cells
EP1084248B1 (fr) Nouveaux analogues de l'insuline presentant un pouvoir de liaison accru avec le zinc
EP0427162A1 (fr) Nouveaux dérivés de l'insuline, leur préparation, leur usage et une composition pharmaceutique les contenant
DE69737766T2 (de) Zusammensetzungen von stammzellfaktor (scf)-analogen und auf diese bezogene verfahren
DE4036856C1 (fr)
DE69636052T2 (de) Mpl-liganden analoga
EP0089007B1 (fr) Procédé de transformation d'analogues de préproinsulines en insulines
WO2004099245A1 (fr) Interferon-$g(y) stabilise
EP0110294B1 (fr) Agent pour traitement d'ostéoporose

Legal Events

Date Code Title Description
AK Designated states

Kind code of ref document: A1

Designated state(s): AE AG AL AM AT AU AZ BA BB BG BR BW BY BZ CA CH CN CO CR CU CZ DK DM DZ EC EE EG ES FI GB GD GE GH GM HR HU ID IL IN IS JP KE KG KP KR KZ LC LK LR LS LT LU LV MA MD MG MK MN MW MX MZ NA NI NO NZ OM PG PH PL PT RO RU SC SD SE SG SK SL SY TJ TM TN TR TT TZ UA UG US UZ VC VN YU ZA ZM ZW

AL Designated countries for regional patents

Kind code of ref document: A1

Designated state(s): BW GH GM KE LS MW MZ NA SD SL SZ TZ UG ZM ZW AM AZ BY KG KZ MD RU TJ TM AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HU IE IT LU MC NL PL PT RO SE SI SK TR BF BJ CF CG CI CM GA GN GQ GW ML MR NE SN TD TG

121 Ep: the epo has been informed by wipo that ep was designated in this application
122 Ep: pct application non-entry in european phase