EP2344182A1 - PRODUCTION AND USE OF HIGH-PURITY HEMOPARATIDE (hPTH-1-37) FOR THE TREATMENT OF INFLAMMATORY SCALING SKIN DISEASES - Google Patents

PRODUCTION AND USE OF HIGH-PURITY HEMOPARATIDE (hPTH-1-37) FOR THE TREATMENT OF INFLAMMATORY SCALING SKIN DISEASES

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Publication number
EP2344182A1
EP2344182A1 EP09783807A EP09783807A EP2344182A1 EP 2344182 A1 EP2344182 A1 EP 2344182A1 EP 09783807 A EP09783807 A EP 09783807A EP 09783807 A EP09783807 A EP 09783807A EP 2344182 A1 EP2344182 A1 EP 2344182A1
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EP
European Patent Office
Prior art keywords
hpth
seq
derivatives
ointment
treatment
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Legal status (The legal status 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 status listed.)
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EP09783807A
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German (de)
French (fr)
Inventor
Wolf-Georg Forssmann
Dirk TEICHMÜLLER
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Haemopep Pharma GmbH
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Haemopep Pharma GmbH
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Priority to EP09783807A priority Critical patent/EP2344182A1/en
Publication of EP2344182A1 publication Critical patent/EP2344182A1/en
Withdrawn legal-status Critical Current

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/22—Hormones
    • A61K38/29—Parathyroid hormone, i.e. parathormone; Parathyroid hormone-related peptides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A61P17/06—Antipsoriatics

Definitions

  • the invention relates to the use of hPTH-1-37 for the manufacture of a medicament for the treatment of inflammatory-scaly diseases, the use of its derivatives for the manufacture of a medicament for the treatment of inflammatory-scaly diseases, a process for the preparation of hPTH-1-37 and a drug comprising hPTH-1-37 or its derivatives.
  • Hemoparatide is the naturally occurring form of the parathyroid hormone parathyroid hormone (hPTH) processed from human blood (Hock et al., 1997).
  • PTH is a vital, parathyroid-derived peptide that plays a key role in many metabolic functions. In particular, it regulates cell proliferation and differentiation of keratinocytes in the skin.
  • Hemoparatide is the most important bioactive form of PTH in the human body, as evidenced by the efficacy and concentration of this peptide in blood plasma.
  • the effects of hPTH on the Skin does not develop in the organism therefore not by the complete molecule, but by the processed form hPTH-1-37.
  • US-B-6066618 discloses a method for inhibiting the proliferation of mammalian skin cells, wherein hPTH 1-34 is used as the antiproliferative peptide. Furthermore, in DE-A-19508672 cyclic parathyroid hormone fragments of z. B. hPTH (I-34) discloses which u. a. can be used to treat psoriasis. Both disclosures have foreign derivatives of human parathyroid hormone in the content, which are therefore expected to be more adverse effects and a poorer bioavailability.
  • WO-A-2004/024758 discloses parathyroid hormone peptides from fish. These are u. a. for the treatment of psoriasis.
  • US-A-2007/0117157 describes the treatment of psoriasis with parathyroid hormone peptides from fish.
  • the PTH derivatives described therein have an amino acid homology of only 53% to human PTH and have a markedly altered biological activity.
  • WO 02/28420 relates to methods of regulating cell differentiation and proliferation, e.g. for the treatment of psoriasis by administration of nucleic acid molecules encoding PTH.
  • WO-A-89/03873 relates to the regulation of cell proliferation by using peptides such as PTH (1-34). However, this application does not relate to PTH peptides 1-37.
  • WO 2005-A-007184 relates to cyclic analogs of hPTH.
  • WO-A-2008/150929 relates to topically administrable composition comprising hPTH 1-37 for the treatment of psoriasis in certain galenic preparations.
  • the chemical synthesis of Hemoparatide has been carried out by several producers, providing the product for preclinical and clinical investigations and studies.
  • the synthesis of hPTH is not trivial, so even products with high levels of impurities in incompletely synthesized peptides have been marketed.
  • An object of the present invention is to provide a high purity form of hemoparatide for use as a therapeutic.
  • galenic forms In addition to the purity requirements of hemoparatides, galenic forms must be found that optimize the use of the high-purity peptide, are adapted to the status and purpose of the treatment, and allow Hemoparatide to be used locally in inflammatory scaly (erythemato-squamous) skin conditions . Such formulations for Hemoparatide, even in highly pure form, are not yet available. Therefore, for today's regulations on the one hand, a sufficiently pure active substance must be available, which on the other hand can be provided in appropriate galenic forms, moreover can be produced commercially, and thus can be regarded as a highly pure form with the highest demands. A further object can thus be seen to provide the high-purity hPTH to be provided in a suitable pharmaceutical dosage form.
  • An object of the present invention is the use of hPTH-1-37 with the amino acid sequence SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVAL (SEQ ID NO: 1) for the manufacture of a medicament for the treatment of inflammatory-scaly (erythemato-squamous) diseases, in particular psoriasis.
  • hPTH-1-37 has a molecular weight of 4401.13 Da.
  • Another object of the present invention are derivatives of hPTH-1-37, namely their natural and pharmacologically acceptable derivatives, in particular amidated, acylated, phosphorylated and glycosylated derivatives for the manufacture of a medicament for the treatment of inflammatory scaly (erythemato-squamous) diseases, in particular psoriasis ,
  • Another aspect of the invention is a process for the preparation of hPTH-1-37 or its derivatives, characterized in that they are synthesized by chemical synthesis from the partial sequences SVSEIQLMHNL (SEQ ID No. 2) and GKHLNSMERVEWLRKKLQDVHNFVAL (SEQ ID No. 3) or SVSEIQLMHNL (SEQ ID No. 2), GKHLNSMERVEW (SEQ ID No. 4) and LRKKLQDVH N FVAL (SEQ ID No. 5) are prepared and purified by chromatography.
  • the hemoparatides (hPTH-1-37) or its derivatives are used for the preparation of a medicament in various pharmaceutical application forms, in particular as a lyophilisate.
  • a hydrophilic ointment is used in particular according to the German Pharmacopoeia as a galenic basis of application. It has surprisingly been found that the active substance is very stable except for methionine-oxidized metabolites in this formulation. These oxidized metabolites also occur as natural PTH forms in the blood plasma, but can be avoided by working in a nitrogen atmosphere and are free from side effects as naturally occurring endogenous derivatives.
  • a further aspect of the invention is a pharmaceutical composition containing from 300 micrograms to 30 milligrams of hPTH-1-37 per gram of preparation (SEQ ID No. 1) and / or from 300 micrograms to 30 milligrams of its derivatives.
  • the formulation in one embodiment contains typical ointment bases into which the peptide hPTH-1-37 is incorporated.
  • a typical ointment contains:
  • Emulsifying cetylstearyl alcohol type A 15-25 g, in particular 21 g
  • Anhydrous citric acid 0.01 - 1.5 g, especially 0.07 g
  • Benzalkonium chloride 50-200 mg, especially 100 mg
  • the amount of hPTH 1-37 may be 0.01 to 1.0% by weight based on the ointment base.
  • Figure 1 discloses an HPLC chromatogram of the high purity hemoparatide
  • FIG. 2 shows a MALDI (matrix assisted laser desorption / ionization) MS spectrum of hPTH (l-37) end product.
  • Figure 3 discloses a chromatogram of capillary zone electrophoresis (CZE) of hPTH (l-37) end product.
  • Figures 4A and 4B disclose HPLC chromatograms of hPTH- (1-37).
  • Figure 4A shows a chromatogram of freshly prepared hPTH- (1-37).
  • Figure 4B is a chromatogram of nine months of stored hPTH- (1-37).
  • Figure 5 discloses the comparison between the HPLC chromatograms of freshly prepared hPTH-1-37
  • the peptides of the invention are prepared by chemical synthesis in solution or on the solid support. This can be different
  • the peptides according to the invention can be obtained from the protected peptide fragment by convergent synthesis.
  • the temporary Fmoc protecting groups were cleaved with 20% piperidine in N-methyl-2-pyrrolidinone within 2-10 minutes. After removal of the Fmoc protecting group, the peptidyl resin was washed thoroughly and repeatedly with NMP followed by dichloromethane and dried. The dry resin was then suspended to cleave the peptide in a freshly prepared mixture of trifluoroacetic acid, ethanedithiol and water (94: 3: 3, vol, 20 ml per 1 g peptidyl resin) and shaken for three hours. The mixture was filtered, the residue was washed with further elimination mixture and the combined filtrates are added slowly with cooling to 10 times the volume of cold tert-butyl methyl ether. The precipitated deprotected peptidic material was stored overnight at + 4 ° C and then isolated by filtration or centrifugation and dried in vacuo.
  • the crude peptide was dissolved in 10% acetic acid and purified by chromatography (Waters Prep-Pak C18, 47 x 300 mm, eluent A: 0.7% trifluoroacetic acid (TFA) in water, eluant B: 0.7% TFA in acetonitrile / water 4 : 1 (vol), gradient: 35-55% Eluent B in 40 minutes, detection: UV at 215 nm, flow rate: 40 ml / min). Fractions containing the product in sufficient purity (determined by analytical HPLC) were combined and freeze-dried.
  • TFA trifluoroacetic acid
  • the dry product was taken up in 10% acetic acid and chromatographed in the presence of acetic acid / acetate (Waters Prep-Pak C18, 47 x 300 mm, equilibrated with 0.1 M ammonium acetate solution, eluent A: 10% acetic acid in Water: eluent B: 2% acetic acid in acetonitrile / water 4: 1 (vol), gradient: 10-60% eluent B in 40 minutes, detection: UV at 215 nm, flow rate: 40 ml / min). Fractions of sufficient purity were combined and freeze-dried.
  • Tab. 1 The essential specifications that are used for the shape of the peptide as a high-purity product.
  • the galenic application are preferably creams on an oil-in-water basis, in the aqueous phase, the water-soluble hemoparatide is incorporated.
  • Hemoparatide was incorporated into this base in concentrations of 0.03 wt%, 0.1 wt% and 0.3 wt%.
  • the active substance is very stable except for methionine-oxidized metabolites in this formulation.
  • These oxidized metabolites also occur as natural PTH forms in the blood plasma, but can be avoided by working in a nitrogen atmosphere and are free from side effects as naturally occurring endogenous derivatives.
  • To use the cream formulation it is applied as usual in galenic units preferably twice a day thinly on the skin.
  • the hPTH-1-37 cream was applied twice a day thinly on the lateral calf area to the right, dorsal to the arrows:
  • the treated area has an area of approx. 4 x 10 cm.
  • the ointment per application contains a total of> 20 ⁇ g hPTH-1-37, ie 40 ⁇ g per day.
  • Left picture shows the right lower leg before treatment, right picture after 14 days of treatment.

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  • Dermatology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
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Abstract

The invention relates to the use of hPTH-1-37 having the amino acid sequence SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVAL (SEQ ID No. 1) or its natural and pharmacologically acceptable derivatives, especially amidated, acylated, phosphorylated and glycosylated derivatives for producing a drug for the treatment of inflammatory scaling (erythematosquamous) diseases, especially psoriasis, the hPTH-1-37 (SEQ ID No. 1) being used in an amount of 300 µg to mg per gram of the drug.

Description

Herstellung und Verwendung von hochreinem Hemoparatide (hPTH-1-37') für die Behandlung von entzündlich-schuppenden Erkrankungen der HautManufacture and use of high purity hemoparatide (hPTH-1-37 ' ) for the treatment of inflammatory scaling disorders of the skin
Die Erfindung betrifft die Verwendung von hPTH-1-37 zur Herstellung eines Arzneimittels zur Behandlung von entzündlich-schuppenden Erkrankungen, die Verwendung seiner Derivate zur Herstellung eines Arzneimittels zur Behandlung von entzündlich-schuppenden Erkrankungen, ein Verfahren zur Herstellung von hPTH-1-37 sowie ein Arzneimittel, welches hPTH-1-37 oder dessen Derivate umfasst.The invention relates to the use of hPTH-1-37 for the manufacture of a medicament for the treatment of inflammatory-scaly diseases, the use of its derivatives for the manufacture of a medicament for the treatment of inflammatory-scaly diseases, a process for the preparation of hPTH-1-37 and a drug comprising hPTH-1-37 or its derivatives.
Hemoparatide (hPTH-1-37) ist die natürlich vorkommende Form des prozessierten Nebenschilddrüsenhormons Parathormon (hPTH), das aus menschlichem Blut dargestellt wurde (Hock et al., 1997). PTH ist ein lebenswichtiges, in der Nebenschilddrüse gebildetes Peptid, das bei zahlreichen Stoffwechselfunktionen eine Schlüsselrolle spielt. Insbesondere reguliert es in der Haut Zellproliferation und Differenzierung der Keratinozyten.Hemoparatide (hPTH-1-37) is the naturally occurring form of the parathyroid hormone parathyroid hormone (hPTH) processed from human blood (Hock et al., 1997). PTH is a vital, parathyroid-derived peptide that plays a key role in many metabolic functions. In particular, it regulates cell proliferation and differentiation of keratinocytes in the skin.
Beim Fehlen von oder Mangel an hPTH treten gravierende Erkrankungen der Haut wie Entzündung, Verdickung und Schuppung auf, da PTH die Zellproliferation der unreifen Keratinozyten hemmt und gleichzeitig deren Umwandlung in reife Hornzellen, die die oberste Schicht der Haut bilden, fördert (Whitfield et al. 2004). Daher können solche entzündlich-schuppenden (erythematom-squamöse) Hauterkrankungen durch die topische Zuführung von PTH therapiert werden. Die Behandlung dieser Erkrankungen, die sowohl die Bereitstellung eines hochreinen Wirkstoffes als auch die Applikation auf die Haut umfasst, wird im folgenden beschrieben und ist Gegenstand der Erfindung.In the absence or lack of hPTH, serious skin diseases such as inflammation, thickening and scaling occur as PTH inhibits the cell proliferation of the immature keratinocytes and at the same time promotes their conversion into mature horny cells which form the uppermost layer of the skin (Whitfield et al. 2004). Therefore, such inflammatory-scaly (erythematic-squamous) skin diseases can be treated by the topical delivery of PTH. The treatment of these diseases, which comprises both the provision of a high-purity active substance and the application to the skin, is described below and is the subject of the invention.
Hemoparatide ist, wie aus der Wirksamkeit und Konzentration dieses Peptides im Blutplasma hervorgeht, die bedeutendste bioaktive Form des PTH im menschlichen Körper. Die oben beschriebenen Wirkungen von hPTH auf die Haut entstehen im Organismus daher nicht durch das vollständige Molekül, sondern durch die prozessierte Form hPTH-1-37.Hemoparatide is the most important bioactive form of PTH in the human body, as evidenced by the efficacy and concentration of this peptide in blood plasma. The effects of hPTH on the Skin does not develop in the organism therefore not by the complete molecule, but by the processed form hPTH-1-37.
US-B-6066618 offenbart ein Verfahren zur Inhibierung der Proliferation von Säugetierhautzellen, wobei hPTH 1-34 als antiproliferatives Peptid eingesetzt wird. Ferner sind in DE-A-19508672 zyklische Parathormonfragmente von z. B. hPTH(l-34) offenbart, welche u. a. zur Behandlung von Psoriasis eingesetzt werden können. Beide Offenbarungen haben körperfremde Derivate von humanem Parathormon zum Inhalt, bei denen daher verstärkt unerwünschte Wirkungen und eine schlechtere Bioverfügbarkeit zu erwarten sind.US-B-6066618 discloses a method for inhibiting the proliferation of mammalian skin cells, wherein hPTH 1-34 is used as the antiproliferative peptide. Furthermore, in DE-A-19508672 cyclic parathyroid hormone fragments of z. B. hPTH (I-34) discloses which u. a. can be used to treat psoriasis. Both disclosures have foreign derivatives of human parathyroid hormone in the content, which are therefore expected to be more adverse effects and a poorer bioavailability.
Darüber hinaus sind in WO-A-2004/024758 Parathormon-Peptide aus Fischen offenbart. Diese eignen sich u. a. zur Behandlung von Psoriasis.In addition, WO-A-2004/024758 discloses parathyroid hormone peptides from fish. These are u. a. for the treatment of psoriasis.
Auch US-A-2007/0117157 beschreibt die Behandlung von Psoriasis mit Parathormon-Peptiden aus Fischen. Die dort beschriebenen PTH-Derivate besitzen jedoch eine Aminosäure-Homologie von nur 53% gegenüber humanem PTH und weisen eine deutlich veränderte biologische Aktivität auf.Also US-A-2007/0117157 describes the treatment of psoriasis with parathyroid hormone peptides from fish. However, the PTH derivatives described therein have an amino acid homology of only 53% to human PTH and have a markedly altered biological activity.
Die WO 02/28420 betrifft Verfahren zur Regulation der Zelldifferentiation und Proliferation, z.B. zur Behandlung von Psoriasis durch Gabe von Nukleinsäuremolekülen, die für PTH kodieren.WO 02/28420 relates to methods of regulating cell differentiation and proliferation, e.g. for the treatment of psoriasis by administration of nucleic acid molecules encoding PTH.
Die WO-A-89/03873 betrifft die Regulation der Zellproliferation durch Verwendung von Peptiden wie PTH (1-34). Diese Anmeldung betrifft aber keine PTH-Peptide 1-37.WO-A-89/03873 relates to the regulation of cell proliferation by using peptides such as PTH (1-34). However, this application does not relate to PTH peptides 1-37.
Die WO 2005-A-007184 betrifft zyklische Analoga des hPTH.WO 2005-A-007184 relates to cyclic analogs of hPTH.
Die WO-A-2008/150929 betrifft topikal verabreichbare Zusammensetzung mit hPTH 1-37 zur Behandlung von Psoriasis in bestimmten galenischen Zubereitungen. Die chemische Synthese von Hemoparatide haben verschiedene Produzenten durchgeführt und somit das Produkt für präklinische und klinische Untersuchungen und Studien geliefert. Die Synthese des hPTH ist nicht trivial, sodass sogar Produkte mit hohen Verunreinigungen an unvollständig synthetisierten Peptiden vertrieben wurden.WO-A-2008/150929 relates to topically administrable composition comprising hPTH 1-37 for the treatment of psoriasis in certain galenic preparations. The chemical synthesis of Hemoparatide has been carried out by several producers, providing the product for preclinical and clinical investigations and studies. The synthesis of hPTH is not trivial, so even products with high levels of impurities in incompletely synthesized peptides have been marketed.
Eine Aufgabe der vorliegenden Erfindung ist die Bereitstellung einer hochreinen Form vom Hemoparatide zur Verwendung als Therapeutikum.An object of the present invention is to provide a high purity form of hemoparatide for use as a therapeutic.
Neben den Anforderungen an die Reinheit von Hemoparatide müssen auch galenische Formen gefunden werden, die die Verwendung des hochreinen Peptides optimieren, dem jeweiligen Status und Ziel der Behandlung angepasst sind und es erlauben, Hemoparatide lokal bei den entzündlich- schuppenden (erythemato-squamösen) Hauterkrankungen anzuwenden. Solche Formulierungen für Hemoparatide, auch in hochreiner Form, liegen bisher nicht vor. Deshalb muss für heutige Vorschriften einerseits ein ausreichend reiner Wirkstoff verfügbar sein, der andererseits in entsprechenden galenischen Formen bereitgestellt werden kann, darüber hinaus kommerziell herstellbar ist, und somit als hochreine Form mit höchstem Anspruch angesehen werden kann. Eine weitere Aufgabe kann also darin gesehen werden, das zur Verfügung zu stellende, hochreine hPTH in einer angemessenen galenischen Darreichungsform bereitzustellen.In addition to the purity requirements of hemoparatides, galenic forms must be found that optimize the use of the high-purity peptide, are adapted to the status and purpose of the treatment, and allow Hemoparatide to be used locally in inflammatory scaly (erythemato-squamous) skin conditions , Such formulations for Hemoparatide, even in highly pure form, are not yet available. Therefore, for today's regulations on the one hand, a sufficiently pure active substance must be available, which on the other hand can be provided in appropriate galenic forms, moreover can be produced commercially, and thus can be regarded as a highly pure form with the highest demands. A further object can thus be seen to provide the high-purity hPTH to be provided in a suitable pharmaceutical dosage form.
Diese Aufgaben werden durch die Verwendung gemäß Anspruch 1 und das Verfahren gemäß Anspruch 3 gelöst.These objects are achieved by the use according to claim 1 and the method according to claim 3.
Ein Gegenstand der vorliegenden Erfindung ist die Verwendung von hPTH-1-37 mit der Aminosäurensequenz SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVAL (SEQ ID NO. 1) zur Herstellung eines Arzneimittels zur Behandlung von entzündlich-schuppenden (erythemato-squamösen) Erkrankungen, insbesondere von Psoriasis. In einer Ausführungsform weist das hPTH-1-37 ein Molekulargewicht von 4401.13 Da auf.An object of the present invention is the use of hPTH-1-37 with the amino acid sequence SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVAL (SEQ ID NO: 1) for the manufacture of a medicament for the treatment of inflammatory-scaly (erythemato-squamous) diseases, in particular psoriasis. In one embodiment, hPTH-1-37 has a molecular weight of 4401.13 Da.
Ein weiterer Gegenstand der vorliegenden Erfindung sind Derivate von hPTH- 1-37, nämlich ihre natürlichen und pharmakologisch verträglichen Derivate, insbesondere amidierte, acylierte, phosphorylierte und glycosylierte Derivate zur Herstellung eines Arzneimittels zur Behandlung von entzündlichschuppenden (erythemato-squamösen) Erkrankungen, insbesondere von Psoriasis.Another object of the present invention are derivatives of hPTH-1-37, namely their natural and pharmacologically acceptable derivatives, in particular amidated, acylated, phosphorylated and glycosylated derivatives for the manufacture of a medicament for the treatment of inflammatory scaly (erythemato-squamous) diseases, in particular psoriasis ,
Ein weiterer Aspekt der Erfindung ist ein Verfahren zur Herstellung von hPTH- 1-37 oder seiner Derivate, dadurch gekennzeichnet, dass diese über chemische Synthese aus den Teilsequenzen SVSEIQLMHNL (SEQ ID No. 2) und GKHLNSMERVEWLRKKLQDVHNFVAL (SEQ ID NO. 3) oder SVSEIQLMHNL(SEQ ID No. 2), GKHLNSMERVEW (SEQ ID NO. 4) und LRKKLQDVH N FVAL (SEQ ID No. 5) hergestellt und chromatographisch gereinigt werden.Another aspect of the invention is a process for the preparation of hPTH-1-37 or its derivatives, characterized in that they are synthesized by chemical synthesis from the partial sequences SVSEIQLMHNL (SEQ ID No. 2) and GKHLNSMERVEWLRKKLQDVHNFVAL (SEQ ID No. 3) or SVSEIQLMHNL (SEQ ID No. 2), GKHLNSMERVEW (SEQ ID No. 4) and LRKKLQDVH N FVAL (SEQ ID No. 5) are prepared and purified by chromatography.
In einer Ausgestaltung der Erfindung werden das Hemoparatide (hPTH-1-37) oder dessen Derivate zur Herstellung eines Arzneimittels in verschiedenen galenischen Applikationsformen, insbesondere als Lyophilisat, verwendet. In einer Ausführungsform dieses Arzneimittels wird eine hydrophile Salbe insbesondere nach Deutschem Arzneibuch als galenische Applikationsgrundlage verwendet. Es wurde überraschend festgestellt, dass der Wirkstoff bis auf methionin-oxidierte Metabolite in dieser Formulierung sehr stabil ist. Diese oxidierten Metaboliten kommen auch als natürliche PTH- Formen im Blutplasma vor, können aber durch Arbeiten in Stickstoffatmosphäre vermieden werden und sind als natürlich vorkommende körpereigene Derivate nebenwirkungsfrei.In one embodiment of the invention, the hemoparatides (hPTH-1-37) or its derivatives are used for the preparation of a medicament in various pharmaceutical application forms, in particular as a lyophilisate. In one embodiment of this drug, a hydrophilic ointment is used in particular according to the German Pharmacopoeia as a galenic basis of application. It has surprisingly been found that the active substance is very stable except for methionine-oxidized metabolites in this formulation. These oxidized metabolites also occur as natural PTH forms in the blood plasma, but can be avoided by working in a nitrogen atmosphere and are free from side effects as naturally occurring endogenous derivatives.
Einen weitereren Aspekt der Erfindung stellt ein Arzneimittel dar, enthaltend 300 Mikrogramm bis 30 Milligramm von hPTH-1-37 pro Gramm Zubereitung (SEQ ID No. 1) und/oder 300 Mikrogramm bis 30 Milligramm seiner Derivate. Die Formulierung enthält in einer Ausführungsform typische Salbengrundlagen, in die das Peptid hPTH-1-37 eingearbeitet wird. Eine typische Salbe enthält:A further aspect of the invention is a pharmaceutical composition containing from 300 micrograms to 30 milligrams of hPTH-1-37 per gram of preparation (SEQ ID No. 1) and / or from 300 micrograms to 30 milligrams of its derivatives. The formulation in one embodiment contains typical ointment bases into which the peptide hPTH-1-37 is incorporated. A typical ointment contains:
Emulgierender Cetylstearylalkohol Typ A 15 - 25 g, insbesondere 21 gEmulsifying cetylstearyl alcohol type A 15-25 g, in particular 21 g
Dünnflüssiges Wachs 5 - 15 g, insbesondere 10 gLow-viscosity wax 5 - 15 g, especially 10 g
Glycerol 85% 3 - 8 g, insbesondere 5 gGlycerol 85% 3 - 8 g, especially 5 g
Kaliumsorbat 0,08 - 1,20 g, insbesondere 0,14 gPotassium sorbate 0.08-1.20 g, especially 0.14 g
Wasserfreie Citronensäure 0,01 - 1,5 g, insbesondere 0,07 gAnhydrous citric acid 0.01 - 1.5 g, especially 0.07 g
Benzalkoniumchlorid 50 - 200 mg, insbesondere 100 mgBenzalkonium chloride 50-200 mg, especially 100 mg
EDTA 50 - 200 mg, insbesondere 100 mgEDTA 50-200 mg, especially 100 mg
Gereinigtes Wasser ad 100 mgPurified water ad 100 mg
Die Menge von hPTH 1- 37 kann 0,01 bis 1,0 Gew.%, bezogen auf die Salbengrundlage betragen.The amount of hPTH 1-37 may be 0.01 to 1.0% by weight based on the ointment base.
Beschreibung der FigurenDescription of the figures
Figur 1 offenbart ein HPLC-Chromatogramm des hochreinen HemoparatideFigure 1 discloses an HPLC chromatogram of the high purity hemoparatide
In Figur 2 ist ein MALDI (Matrix assisted laser desorption/ionisation)-MS- Spektrum von hPTH-(l-37)-Endprodukt dargestellt.FIG. 2 shows a MALDI (matrix assisted laser desorption / ionization) MS spectrum of hPTH (l-37) end product.
Figur 3 offenbart ein Chromatogramm der Kapillarzonenelektrophorese (CZE) von hPTH-(l-37)-Endprodukt.Figure 3 discloses a chromatogram of capillary zone electrophoresis (CZE) of hPTH (l-37) end product.
Die Figuren 4A und 4B offenbaren HPLC-Chromatogramme von hPTH-(l-37).Figures 4A and 4B disclose HPLC chromatograms of hPTH- (1-37).
Figur 4A zeigt ein Chromatogramm von frisch hergestelltem hPTH-(l-37).Figure 4A shows a chromatogram of freshly prepared hPTH- (1-37).
Figur 4B ist ein Chromatogramm von neun Monaten gelagertem hPTH-(l-37).Figure 4B is a chromatogram of nine months of stored hPTH- (1-37).
Es sind nur sehr geringe Mengen an methionin-oxidierten Metaboliten nachweisbar (siehe Satelliten-Peaks). Ferner offenbart Figur 5 den Vergleich zwischen den HPLC-Chromatogrammem von frisch hergestelltem hPTH-1-37Only very small amounts of methionine-oxidized metabolites are detectable (see satellite peaks). Further, Figure 5 discloses the comparison between the HPLC chromatograms of freshly prepared hPTH-1-37
(A) und einem kommerziell erhältlichen Präparat. Die überraschend gute Qualität lässt sich durch das Fehlen von Sateliten-Peaks in 5A gegenüber 5B ersehen.(A) and a commercially available preparation. The surprisingly good Quality can be seen from the lack of satellite peaks in 5A versus 5B.
Die Herstellung des Wirkstoffes und dessen Formulierung ist im Folgenden beschrieben :The preparation of the active ingredient and its formulation is described below:
Chemische Synthese von hPTH-1-37Chemical synthesis of hPTH-1-37
Die erfindungsgemäßen Peptide werden durch chemische Synthese in Lösung oder am festen Träger hergestellt. Hierzu können verschiedeneThe peptides of the invention are prepared by chemical synthesis in solution or on the solid support. This can be different
Schutzgruppenkombinationen, z.B. die Fmoc- oder Boc-geschützteProtecting group combinations, e.g. the Fmoc or Boc-protected
Aminosäuren eingesetzt werden. Neben der schrittweisenAmino acids are used. In addition to the gradual
Festphasenpeptidsynthese nach dem klassischen Merrifield-Prinzip können die erfindungsgemäßen Peptide aus dem geschütztem Peptidfragment durch konvergente Synthese gewonnen werden.Solid phase peptide synthesis according to the classical Merrifield principle, the peptides according to the invention can be obtained from the protected peptide fragment by convergent synthesis.
hPTH-1-37 mit der AminosäuresequenzhPTH-1-37 with the amino acid sequence
SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVAL
wurde unter Verwendung Fmoc(fluorenylmethoxycarbonyl)-geschützter Aminosäuren durch schrittweise Festphasensynthese hergestellt. Die Synthese wurde auf einem mit Fmoc-Leucin beladenen Wang-Harz (0,69 mmol/g, 100 - 200 mesh) als festem Träger durchgeführt. Die Aktivierung der Fmoc- Aminosäuren, die in 10-fachem molaren Überschuss eingesetzt wurden, wurde mit [(2-(lH-Benzotriazol-l-yl)-l,l,3,3-tetramethyluronium-hexa-fluoro-phos- phat] (HBTU) unter Zusatz von 1-Hydroxybenzotriazol (HOBt) und Diisopropylethylamin (DIEA) in N-Methyl-2-pyrrolidinon (NMP) bei Raumtemperatur durchgeführt. Acylierungsreaktionen wurden typischerweise für 45 Minuten durchgeführt. Folgende Aminosäurederivate wurden zur Synthese eingesetzt: Fmoc-L-Ala, Fmoc-L-Arg(Pbf), Fmoc-L-Asn(Trt), Fmoc-L- Asp(OtBu), Fmoc-L-Glu(OtBu), Fmoc-L-Gln(Trt), Fmoc-Gly, Fmoc-L-His(Trt), Fmoc-L-Ile, Fmoc-L-Leu, Fmoc-L-Lys(Boc), Fmoc-L-Met, Fmoc-L-Phe, Fmoc-L- Ser(tBu), Fmoc-L-Trp(Boc) und Fmoc-L-Val. Die temporären Fmoc- Schutzgruppen wurden mit 20% Piperidin in N-Methyl-2-pyrrolidinon innerhalb von 2-10 Minuten abgespalten. Nach Entfernung der Fmoc-Schutzgruppe wurde das Peptidylharz sorgfältig und mehrfach mit NMP und anschließend Dichlormethan gewaschen und getrocknet. Das trockene Harz wurde anschließend zur Abspaltung des Peptides in einer frisch hergestellten Mischung aus Trifluoressigsäure, Ethandithiol und Wasser (94 : 3 : 3, vol, 20 ml pro 1 g Peptidylharz) suspendiert und für drei Stunden geschüttelt. Die Mischung wurde filtriert, der Rückstand mit weiterer Abspaltmischung gewaschen und die vereinigten Filtrate werden unter Kühlung langsam zum 10-fachen Volumen kaltem tert-Butylmethylether gegeben. Das ausgefallene entschützte peptidische Material wurde über Nacht bei +4°C gelagert und anschließend durch Filtration oder Zentrifugation isoliert und im Vakuum getrocknet.was prepared by using Fmoc (fluorenylmethoxycarbonyl) -protected amino acids by stepwise solid-phase synthesis. The synthesis was performed on a Fmoc-leucine loaded Wang resin (0.69 mmol / g, 100-200 mesh) as a solid support. The activation of the Fmoc amino acids, which were used in a 10-fold molar excess, was treated with [(2- (1H-benzotriazol-1-yl) -1,3,3-tetramethyluronium hexa-fluorophosphate ] (HBTU) with the addition of 1-hydroxybenzotriazole (HOBt) and diisopropylethylamine (DIEA) in N-methyl-2-pyrrolidinone (NMP) at room temperature.Acylation reactions were typically carried out for 45 minutes.The following amino acid derivatives were used for synthesis: Fmoc- L-Ala, Fmoc-L-Arg (Pbf), Fmoc-L-Asn (Trt), Fmoc-L-Asp (OtBu), Fmoc-L-Glu (OtBu), Fmoc-L-Gln (Trt), Fmoc -Gly, Fmoc-L-His (Trt), Fmoc-L-Ile, Fmoc-L-Leu, Fmoc-L-Lys (Boc), Fmoc-L-Met, Fmoc-L-Phe, Fmoc-L- Ser (tBu), Fmoc-L-Trp (Boc) and Fmoc-L-Val. The temporary Fmoc protecting groups were cleaved with 20% piperidine in N-methyl-2-pyrrolidinone within 2-10 minutes. After removal of the Fmoc protecting group, the peptidyl resin was washed thoroughly and repeatedly with NMP followed by dichloromethane and dried. The dry resin was then suspended to cleave the peptide in a freshly prepared mixture of trifluoroacetic acid, ethanedithiol and water (94: 3: 3, vol, 20 ml per 1 g peptidyl resin) and shaken for three hours. The mixture was filtered, the residue was washed with further elimination mixture and the combined filtrates are added slowly with cooling to 10 times the volume of cold tert-butyl methyl ether. The precipitated deprotected peptidic material was stored overnight at + 4 ° C and then isolated by filtration or centrifugation and dried in vacuo.
Das Rohpeptid wurde in 10% Essigsäure gelöst und chromatographisch gereinigt (Waters Prep-Pak C18, 47 x 300 mm; Eluens A: 0,7% Trifluoressigsäure (TFA) in Wasser; Eluens B: 0,7% TFA in Acetonitril / Wasser 4: 1 (vol); Gradient: 35 - 55% Eluens B in 40 Minuten; Detektion : UV bei 215 nm; Flussrate: 40 ml/min). Fraktionen, die das Produkt in ausreichender Reinheit enthalten (bestimmt durch analytische HPLC) wurden vereinigt und gefriergetrocknet. Zum Austausch des Trifluoracetatgegenions gegen Acetat wurde das trockene Produkt in 10% Essigsäure aufgenommen und in Gegenwart von Essigsäure/Acetat chromatographiert (Waters Prep-Pak C18, 47 x 300 mm, equilibriert mit 0,1 M Amoniumacetatlösung; Eluens A: 10% Essigsäure in Wasser; Eluens B: 2% Essigsäure in Acetonitril / Wasser 4 : 1 (vol); Gradient: 10-60% Eluens B in 40 Minuten; Detektion : UV bei 215 nm; Flussrate: 40ml/min). Fraktionen genügender Reinheit wurden vereinigt und gefriergetrocknet. Trockenes Endprodukt wurde anschließend durch RP-HPLC (Abbildung 1), MALDI-MS (Abbildung 2) und Kapillarzonenelektrophorese (Abbildung 3) analysiert, und es wurde überraschend ein hochreines Produkt gefunden, dessen Spezifikation sich von früheren Synthesen durch deutlich bessere Qualität unterscheidet (siehe Abbildung 4A und 4B). Die überraschend gute Qualität ist auch in einer Tabelle (Tabelle 1) gegenüber kommerziell erhältlichem Produkt (Abbildung 5A gegenüber 5B) zu erkennen.The crude peptide was dissolved in 10% acetic acid and purified by chromatography (Waters Prep-Pak C18, 47 x 300 mm, eluent A: 0.7% trifluoroacetic acid (TFA) in water, eluant B: 0.7% TFA in acetonitrile / water 4 : 1 (vol), gradient: 35-55% Eluent B in 40 minutes, detection: UV at 215 nm, flow rate: 40 ml / min). Fractions containing the product in sufficient purity (determined by analytical HPLC) were combined and freeze-dried. To replace the trifluoroacetate counterion with acetate, the dry product was taken up in 10% acetic acid and chromatographed in the presence of acetic acid / acetate (Waters Prep-Pak C18, 47 x 300 mm, equilibrated with 0.1 M ammonium acetate solution, eluent A: 10% acetic acid in Water: eluent B: 2% acetic acid in acetonitrile / water 4: 1 (vol), gradient: 10-60% eluent B in 40 minutes, detection: UV at 215 nm, flow rate: 40 ml / min). Fractions of sufficient purity were combined and freeze-dried. Dry final product was then analyzed by RP-HPLC (Figure 1), MALDI-MS (Figure 2), and capillary zone electrophoresis (Figure 3), and surprisingly, a high purity product was found, the specification of which has been significantly improved by previous syntheses better quality (see Figures 4A and 4B). The surprisingly good quality can also be seen in a table (Table 1) compared to commercially available product (Figure 5A vs. 5B).
Typische Charakteristik für hPTH-1-37 und hochreines Hemoparatide PräparatTypical characteristics of hPTH-1-37 and high purity hemoparatide preparation
Tab. 1 : Die wesentlichen Spezifikationen, die für die Form des Peptides als hochreines Produkt herangezogen werden.Tab. 1: The essential specifications that are used for the shape of the peptide as a high-purity product.
Die hohe Stabilität des Präparates von hochreinem Hemoparatide in lyophilisierter Form bei einer Temperatur von +4°C ist durch entsprechende Analytik belegt, wobei frisches Material (Abbildung 4A) mit neun Monaten gelagertem Material (Abbildung 4B) verglichen ist. Es erscheinen nach dieser Lagerzeit nur geringe Methionin-oxidierte Metabolite.The high stability of the preparation of high purity hemoparatide in lyophilised form at a temperature of + 4 ° C is proven by appropriate analysis, with fresh material (Figure 4A) at nine months stored material (Figure 4B). After this storage time, only small methionine-oxidized metabolites appear.
Herstellung einer neuen Formulierungen von HemoparatidePreparation of a new formulation of Hemoparatide
Für die galenische Anwendung eignen sich vorzugsweise Cremes auf Öl-inWasser-Basis, in deren wässrige Phase das wasserlösliche Hemoparatide eingearbeitet ist.For the galenic application are preferably creams on an oil-in-water basis, in the aqueous phase, the water-soluble hemoparatide is incorporated.
Eine solche Cremegrundlage stellt die hydrophile Salbe nach Deutschem Arzneibuch (Unguentum emulsificans aquosum DAB) dar.One such cream base is the hydrophilic ointment according to the German Pharmacopoeia (Unguentum emulsificans aquosum DAB).
Emulgierender Cetylstearylalkohol Typ A 21 gEmulsifying cetylstearyl alcohol type A 21 g
Dünnflüssiges Wachs 10 gLow-viscosity wax 10 g
Glycerol 85% 5 gGlycerol 85% 5 g
Kaliumsorbat 0,14 gPotassium sorbate 0.14 g
Wasserfreie Citronensäure 0,07 gAnhydrous citric acid 0.07 g
Benzalkoniumchlorid 100 mgBenzalkonium chloride 100 mg
EDTA 100 mgEDTA 100 mg
Gereinigtes Wasser ad 100 gPurified water ad 100 g
Tab. 2 : Alle eingesetzten Rohstoffe sind Pharma-konform.Tab. 2: All raw materials used are pharmaceutical compliant.
Hemoparatide wurde in Konzentrationen von 0,03 Gew.-%, 0,1 Gew.-% und 0,3 Gew.-% in diese Grundlage eingebracht.Hemoparatide was incorporated into this base in concentrations of 0.03 wt%, 0.1 wt% and 0.3 wt%.
Es wurde überraschend festgestellt, dass der Wirkstoff bis auf methionin- oxidierte Metabolite in dieser Formulierung sehr stabil ist. Diese oxidierten Metaboliten kommen auch als natürliche PTH-Formen im Blutplasma vor, können aber durch Arbeiten in Stickstoffatmosphäre vermieden werden und sind als natürlich vorkommende körpereigene Derivate nebenwirkungsfrei. Zur Benutzung der Creme-Formulierung wird diese wie üblich in galenischen Einheiten vorzugsweise zweimal täglich dünn auf die Haut aufgetragen.It has surprisingly been found that the active substance is very stable except for methionine-oxidized metabolites in this formulation. These oxidized metabolites also occur as natural PTH forms in the blood plasma, but can be avoided by working in a nitrogen atmosphere and are free from side effects as naturally occurring endogenous derivatives. To use the cream formulation, it is applied as usual in galenic units preferably twice a day thinly on the skin.
Nachweis der mikrobiellen Stabilität im KonservierunqsbelastunqstestDemonstration of microbial stability in the preservation challenge test
Im Konservierungsbelastungstest wurde die mikrobielle Produktstabilität der erfindungsgemäßen Rezeptur nach Pharma-Richtlinien (Plattengussverfahren) nachgewiesen. Die Ergebnisse zeigen, dass alle Anforderungen, die für eine Hautmedikation notwendig sein werden, erfüllt werden.In the preservation stress test, the microbial product stability of the formulation according to the invention was demonstrated according to pharmaceutical guidelines (slab casting method). The results show that all the requirements that will be necessary for skin medication are met.
Verträglichkeit und Sicherheit der Hemoparatide-CremeformulierunqTolerability and Safety of Hemoparatide Cream Formulation
Beim Menschen ist die gute Verträglichkeit des hochreinen Produktes bei subcutaner Injektion in wässriger Lösung durch eine Studie Phase I belegt. Erfindungsgemäß entwickelte Formulierungen für die topische Anwendung des hochreinen Wirkstoffes Hemoparatide können also in hohem Maß als sicher und verträglich gelten (siehe Figur 6). In humans, the good tolerability of the high purity product in subcutaneous injection in aqueous solution is evidenced by a Phase I study. Formulations developed according to the invention for the topical application of the high-purity active substance hemoparatide can therefore be considered to be safe and compatible to a high degree (see FIG. 6).
Nachweis des Behandlungserfolges durch Anwendung von Hemoparatide-Creme bei Psoriasis:Proof of treatment success by using Hemoparatide cream for psoriasis:
Die hPTH-1-37 Creme wurde täglich zwei Mal dünn auf dem seitlichen Wadenareal rechts dorsal zu den Pfeilen aufgetragen :The hPTH-1-37 cream was applied twice a day thinly on the lateral calf area to the right, dorsal to the arrows:
1. Das behandelte Areal hat eine Fläche von ca. 4 mal 10 cm.1. The treated area has an area of approx. 4 x 10 cm.
2. Es wurden jeweils morgens und abends ca. 250 mg Salbe gleichmäßig aufgetragen, indem die Salbe mit dem Zeigefinger verteilt wurde. 3. Die Salbe pro Applikation enthält insgesamt > 20 μg hPTH-1-37 also 40 μg pro Tag.2. In the morning and in the evening, about 250 mg of ointment were evenly applied by spreading the ointment with the index finger. 3. The ointment per application contains a total of> 20 μg hPTH-1-37, ie 40 μg per day.
4. linkes Bild zeigt den rechten Unterschenkel vor Behandlung, rechtes Bild nach 14 Tagen der Behandlung.4. Left picture shows the right lower leg before treatment, right picture after 14 days of treatment.
Deutlich ist die Auflockerung der Effloreszenz und Rückgang der entzündlichen Reaktion nach 14 Tagen zu erkennen. Der Juckreiz lässt auch in den angrenzenden Arealen nach, und die Wirkung ist bei längerer Behandlung und Absetzen der Medikation nach über 4 Wochen nachhaltig.Clearly the loosening of the efflorescence and decrease of the inflammatory reaction can be recognized after 14 days. The itching also subsides in the adjoining areas, and the effect is lasting with longer treatment and discontinuation of the medication after more than 4 weeks.
Literaturliterature
Hock D, Mägerlein M, Heine G, Ochlich PP, Forssmann WG (1997) Isolation and characterization of the bioactive circulating human parathyroid hormone, hPTH-1-37. FEBS Lett. 400: 221-225.Hock D, Mägerlein M, Heine G, Ochlich PP, Forssmann WG (1997) Isolation and characterization of the bioactive circulating human parathyroid hormone, hPTH-1-37. FEBS Lett. 400: 221-225.
Whitfield JF (2004) Taming psoriatic keratinocytes - PTHs' uses go up another notch. JCB 93: 251-256. Whitfield JF (2004) Taming psoriatic keratinocytes - PTHs' uses go up another notch. JCB 93: 251-256.

Claims

Patentansprüche claims
1. Verwendung von hPTH-1-37 mit der Aminosäurensequenz SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVAL (SEQ ID NO. 1) oder seiner natürlichen und pharmakologisch verträglichen Derivate, insbesondere amidierte, acylierte, phosphorylierte und glycosylierte Derivate zur Herstellung eines Arzneimittels zur Behandlung von entzündlich-schuppenden (erythemato-squamösen) Erkrankungen, insbesondere von Psoriasis, wobei hPTH-1-37 (SEQ ID NO. 1) in einer Menge von 300 μg bis mg pro Gramm Arzneimittel vorliegt.1. Use of hPTH-1-37 with the amino acid sequence SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVAL (SEQ ID NO: 1) or its natural and pharmacologically acceptable derivatives, in particular amidated, acylated, phosphorylated and glycosylated derivatives for the manufacture of a medicament for the treatment of inflammatory-scaly (erythematous) squamous) diseases, particularly psoriasis, wherein hPTH-1-37 (SEQ ID NO: 1) is present in an amount of 300 μg to mg per gram of drug.
2. Verwendung gemäß Anspruch 1, wobei hPTH-1-37 (SEQ ID NO. 1) in einer Menge von 0,6 bis 3 Gew.% im Arzneimittel vorliegt. 3. Verfahren zur Herstellung von hPTH-1-37 oder seiner Derivate, dadurch gekennzeichnet, dass diese über chemische Synthese aus den Teilsequenzen SVSEIQLMHNL (SEQ ID No. 2) und GKHLNSMERVEWLRKKLQDVHNFVAL (SEQ ID NO . 2. Use according to claim 1, wherein hPTH-1-37 (SEQ ID NO: 1) is present in an amount of 0.6 to 3% by weight in the drug. 3. A process for the preparation of hPTH-1-37 or its derivatives, characterized in that these are prepared by chemical synthesis from the partial sequences SVSEIQLMHNL (SEQ ID No. 2) and GKHLNSMERVEWLRKKLQDVHNFVAL (SEQ ID NO.
3) oder SVSEIQLM H N L(SEQ ID NO . 2), GKHLNSMERVEW (SEQ ID No. 4) und LRKKLQDVHNFVAL (SEQ ID NO. 5) hergestellt und chromatographisch gereinigt werden.3) or SVSEIQLM H N L (SEQ ID NO: 2), GKHLNSMERVEW (SEQ ID NO 4), and LRKKLQDVHNFVAL (SEQ ID NO: 5) are prepared and purified by chromatography.
4. Salbe enthaltend 300 μg bis 30 mg pro Gramm Salbe hPTH-1-37 SEQ ID No. 1 oder seiner natürlichen und pharmakologisch verträglichen Derivate, insbesondere amidierte, acylierte, phosphorylierte und glycosylierte Derivate, insbesondere 6 mg bis 30 mg hPTH-1-37 pro Gramm Salbe.4. Ointment containing 300 μg to 30 mg per gram ointment hPTH-1-37 SEQ ID NO. 1 or its natural and pharmacologically acceptable derivatives, in particular amidated, acylated, phosphorylated and glycosylated derivatives, in particular 6 mg to 30 mg hPTH-1-37 per gram ointment.
5. Salbe nach Anspruch 4, wobei die Salbengrundlage ist:5. ointment according to claim 4, wherein the ointment base is:
Emulgierender Cetylstearylalkohol Typ A 15 - 25 g, insbesondere 21 gEmulsifying cetylstearyl alcohol type A 15-25 g, in particular 21 g
Dünnflüssiges Wachs 5 - 15 g, insbesondere 10 gLow-viscosity wax 5 - 15 g, especially 10 g
Glycerol 85% 3 - 8 g, insbesondere 5 gGlycerol 85% 3 - 8 g, especially 5 g
Kaliumsorbat 0,08 - 1,20 g, insbesondere 0,14 gPotassium sorbate 0.08-1.20 g, especially 0.14 g
Wasserfreie Citronensäure 0,01 - 1,5 g, insbesondere 0,07 gAnhydrous citric acid 0.01 - 1.5 g, especially 0.07 g
Benzalkoniumchlorid 50 - 200 mg, insbesondere 100 mgBenzalkonium chloride 50-200 mg, especially 100 mg
EDTA 50 - 200 mg, insbesondere 100 mgEDTA 50-200 mg, especially 100 mg
Gereinigtes Wasser ad 100 mg Purified water ad 100 mg
EP09783807A 2008-10-07 2009-10-07 PRODUCTION AND USE OF HIGH-PURITY HEMOPARATIDE (hPTH-1-37) FOR THE TREATMENT OF INFLAMMATORY SCALING SKIN DISEASES Withdrawn EP2344182A1 (en)

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PCT/EP2009/063017 WO2010040769A1 (en) 2008-10-07 2009-10-07 PRODUCTION AND USE OF HIGH-PURITY HEMOPARATIDE (hPTH-1-37) FOR THE TREATMENT OF INFLAMMATORY SCALING SKIN DISEASES

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EP0415924B1 (en) * 1987-10-20 1996-05-29 HOLICK, Michael F Regulation of cell proliferation and differentiation using peptides
DE19957918A1 (en) * 1999-11-25 2001-06-13 Ulrich Doht Disinfectant cleaner for cleaning and care and process for its manufacture
JP2004500404A (en) * 2000-03-27 2004-01-08 カール　−　ツァイス　−　シュティフツング Novel cosmetic, body care, cleansing and nutritional supplement compositions containing bioactive glass and methods of manufacture and use thereof
AU2003239869A1 (en) * 2002-05-23 2003-12-12 Michael Holick Use of a parathyroid hormone peptide analogs for the treatment of vaginal atrophy
WO2008150929A1 (en) * 2007-05-29 2008-12-11 Manhattan Pharmaceuticals, Inc. Topical compositions comprising a macromolecule and methods of using same

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