EP1148888A2 - Treatment of erectile dysfunctions with a c-type natriuretic polypeptide (cnp) as monotherapy or in combination with phosphodiesterase inhibitors - Google Patents
Treatment of erectile dysfunctions with a c-type natriuretic polypeptide (cnp) as monotherapy or in combination with phosphodiesterase inhibitorsInfo
- Publication number
- EP1148888A2 EP1148888A2 EP00909112A EP00909112A EP1148888A2 EP 1148888 A2 EP1148888 A2 EP 1148888A2 EP 00909112 A EP00909112 A EP 00909112A EP 00909112 A EP00909112 A EP 00909112A EP 1148888 A2 EP1148888 A2 EP 1148888A2
- Authority
- EP
- European Patent Office
- Prior art keywords
- cnp
- phosphodiesterase inhibitors
- combination
- treatment
- type natriuretic
- Prior art date
- 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.)
- Withdrawn
Links
Classifications
-
- 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
- A61K38/2242—Atrial natriuretic factor complex: Atriopeptins, atrial natriuretic protein [ANP]; Cardionatrin, Cardiodilatin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P15/00—Drugs for genital or sexual disorders; Contraceptives
- A61P15/10—Drugs for genital or sexual disorders; Contraceptives for impotence
Definitions
- CNP C-type natriuretic polypeptide
- the present invention relates to a medicament for the treatment of erectile dysfunctions and the use of the C-type natriuretic polypeptide (CNP) and its variants individually or in combination with phosphodiesterase inhibitors for the treatment of erectile dysfunctions.
- CNP C-type natriuretic polypeptide
- cAMP cyclic adenosine monophosphate
- Sildenafil is a PDE5 inhibitor currently used under the Viagra is already successfully used as a therapeutic agent for the treatment of erectile dysfunctions (Boolell et al., 1997). However, undesirable side effects of sildenafil, such as eye flicker, headache or circulatory failure, have been described.
- the object of the present invention was to provide an agent which reduces or avoids the side effects mentioned.
- CNP can also be replaced or supplemented by fragments or derivatives with the physiological effect of the CNP obtained. Partial fragments contained in the CNP prohormone sequence are particularly suitable as fragments.
- the amino acid sequences of the CNP, proCNP and preproCNP are shown below:
- GLSKGCFGLKLDRIGSMSGLGC Suitable derivatives are, in particular, those structures derived from the formula given which arise from the conservative exchange of amino acids in this sequence. Derivatives of the CNP containing amidated, phosphorylated, acylated, alkylated and / or linkages of side chains are also suitable.
- Fragments are also obtained by deletion of amino acids which are not involved in the function and can be used as medicaments according to the invention.
- the medicament preferably also contains necessary auxiliaries and / or carriers which enable or support the galenic application.
- the medicament according to the invention is administered intravenously, it is advisable to dissolve the components in question in appropriate preparations suitable for intravenous administration.
- Other forms of administration are oral, intracavernous, intraurethral, locally topical, transcutaneous.
- Particularly suitable phosphodlesterase inhibitors are those of PDE5 phosphodiesterase. These include, in particular, Zaprinast, Dipyridamol, Milrinon and / or Sildenafil.
- CNP or fragments or derivatives of the CNP with preserved physiological effect of the CNP for the treatment of erectile dysfunctions is claimed.
- CNP or fragments or derivatives of CNP with a physiological value obtained Effect of CNP used in combination with phosphodiesterase inhibitors.
- an increase in the intracellular cAMP level is achieved by stimulating the formation (not the inhibition of hydrolysis) of the PDE3A inhibitor cGMP.
- the figure shows the effect of CNP and ANP / CDD99-126 on smooth muscle preparations of human corpus cavernosum penis in the form of the effect of CNP and ANP / CDD99- 1 26 vorkontrahierter on the relaxation of smooth muscle preparations.
- CNP already shows a significant relaxing effect at a concentration of 1 nM, which finally leads to a relaxation corresponding to 60% of the tone existing after norepinephrine administration when increased to 1 ⁇ M.
- guanylate cyclases In the enzymes that catalyze the synthesis of cGMP, the guanylate cyclases, a distinction must be made between two different subgroups, namely the soluble cytosolic and the membrane-bound guanylate cyclases (for an overview, see Wedel and Garbers, 1997). So far it was assumed that only the soluble cytosolic guanylate cyclases play a role in triggering the erection.
- RNA was isolated from biopsy samples from human corpus cavernosum and the subsequent transcription of the messenger RNA (mRNA) contained in 5 ⁇ g total RNA into copy DNA first strand (cDNA first strand) as already described (Magert et al. , 1995; Magert et al., 1998). Each 1/300 of the first-strand cDNA batches were then used in standard RT-PCR reactions (Saiki et al., 1988) with the primer pairs derived from cDNA database entries and specific for the different guanylate cyclases. The reaction conditions were as follows:
- the products obtained were separated electrophoretically in an agarose gel and, after staining with ethidium bromide, visualized using ultraviolet transmitted light.
- guanylate cyclase B (sense primer GCB-1S: GGGTGCTGTGGCCTCTGG I I I I I CGG, antisense primer GCB-2AS: CCCTGCATC I I I CCACAATTCGAAG) a homogeneous band could be obtained.
- GC-B guanylate cyclase B
- GC-B is the receptor for the peptide hormone C-type natriuretic polypeptide (CNP) formed in the brain, it was checked as follows whether human CNP has relaxing effects on the smooth muscles of the corpus cavernosum or an increase in intracellular concentration of cyclic nucleoside monophosphates induced:
- Strip preparations of human cavernous muscles were in the with a modified KREBS-RINGER solution (NaCI 120 mM, NaHC0 3 25.6 mM, KCI 4.7 mM, CaCI 2 2.5 mM, NaH 2 P0 4 1.2 mM, MgCI 2 1.2 mM, glucose 22 mM, 2Na + (Ca 2+ ) EDTA 0.1 mM, (pH 7.2-7.4) filled bath chambers of an organ bath system (Isolated Organ Apparatus IOA 5306, Fschreib Medical Instruments GmbH, Seeheim, Germany) The bath solution was heated to 37 ° C. and was continuously gassed with a mixture of 95% O2 and 5% CO 2.
- a modified KREBS-RINGER solution NaCI 120 mM, NaHC0 3 25.6 mM, KCI 4.7 mM, CaCI 2 2.5 mM, NaH 2 P0 4 1.2 mM, MgCI 2 1.2 mM, glucose 22 mM, 2N
- tissue concentrations of cyclic nucleotides To determine the time and dose-dependent effects of ANP / CDD99-126 and CNP on the intracellular concentration of cAMP and cGMP, strips of cavernous musculature were incubated with KREBS-RINGER solution in 2 ml reaction vessels analogous to the organ bath experiments, continuously gassed with carbogen and with 6 ⁇ M NOR stimulated. Subsequently, the incubation with ANP / CDD9 9 was - 12 6 and CNP (0.1, 10 and 1000 nM) for 10 min.
- silicafil an orally active type 5 cyclic GMP-specific phosphodiesterase inhibitor for the treatment of penile erectile dysfunction. Int. J. Impot. Res. 8: 47-52.
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Pharmacology & Pharmacy (AREA)
- Engineering & Computer Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Endocrinology (AREA)
- Veterinary Medicine (AREA)
- Chemical & Material Sciences (AREA)
- Public Health (AREA)
- General Health & Medical Sciences (AREA)
- Medicinal Chemistry (AREA)
- Animal Behavior & Ethology (AREA)
- Gastroenterology & Hepatology (AREA)
- Molecular Biology (AREA)
- Epidemiology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Immunology (AREA)
- Zoology (AREA)
- Cardiology (AREA)
- Gynecology & Obstetrics (AREA)
- Reproductive Health (AREA)
- Organic Chemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Enzymes And Modification Thereof (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
- Peptides Or Proteins (AREA)
Abstract
The invention relates to medicaments containing a combination of CNP or fragments or derivatives with obtained physiological action of the CNP and phosphodiesterase inhibitors.
Description
Behandlung von erektilen Dysfunktionen mit C-Typ Natriureti- schem Polypeptid (CNP) als Monotherapie oder in Kombination mit PhosphodiesterasehemmernTreatment of erectile dysfunctions with C-type natriuretic polypeptide (CNP) as monotherapy or in combination with phosphodiesterase inhibitors
Gegenstand der vorliegenden Erfindung ist ein Arzneimitttel zur Behandlung von erektilen Dysfunktionen sowie die Verwendung des C-Typ Natriuretischen Polypeptids (CNP) und seiner Varianten einzeln oder in Kombination mit Phosphodiesterase-Hemmstoffen zur Behandlung erek- tiler Dysfunktionen.The present invention relates to a medicament for the treatment of erectile dysfunctions and the use of the C-type natriuretic polypeptide (CNP) and its variants individually or in combination with phosphodiesterase inhibitors for the treatment of erectile dysfunctions.
Die zur penilen Erektion führende Relaxation der glatten Muskulatur des Corpus cavernosum penis wird durch ein komplexes Zusammenspiel verschiedener Faktoren ausgelöst, wobei der letztendlich gebildete intrazelluläre sekundäre Botenstoff zyklisches Adenosinmonophosphat (cAMP) eine essentielle Rolle spielt (Stief et al., 1997). Dessen Konzentration wird einerseits durch die Aktivität cAMP-Synthese katalysierender Enzyme, nämlich der Adenylatzyklasen, andererseits durch cAMP abbauende (hydrolysierende) Enzyme, die Phosphodiesterasen (PDEs), bestimmt. Es sind bisher mehr als zehn verschiedene Phosphodiestera- se-Isoenzyme bekannt, unter denen manche cAMP, andere jedoch auch den verwandten sekundären Botenstoff zyklisches Guanosinmo- nophosphat (cGMP) hydrolysieren (Beavo und Reifsnyder, 1990; Burns et al., 1996; Taher et al., 1997). Da eine cAMP-hydrolysierende Phosphodiesterase, die PDE3A, durch cGMP gehemmt wird, ist die cGMP-hydrolysierende Phosphodiesterase 5 (PDE5) in den letzten Jahren selbst in den Mittelpunkt der Erforschung erektiler Dysfunktionen geraten. Sildenafil ist ein PDE5-Hemmstoff, der zur Zeit unter der Be-
Zeichnung Viagra bereits erfolgreich als Therapeutikum zur Behandlung erektiler Dysfunktionen in Anwendung ist (Boolell et al., 1997). Unerwünschte Nebenwirkungen von Sildenafil, wie beispielsweise Augen- flimmern, Kopfschmerzen oder Kreislaufversagen, wurden allerdings beschrieben.The relaxation of the smooth muscles of the corpus cavernosum penis, which leads to penile erection, is triggered by a complex interplay of various factors, whereby the ultimately formed intracellular secondary messenger cyclic adenosine monophosphate (cAMP) plays an essential role (Stief et al., 1997). Its concentration is determined on the one hand by the activity of enzymes catalyzing cAMP, namely the adenylate cyclases, and on the other hand by cAMP-degrading (hydrolyzing) enzymes, the phosphodiesterases (PDEs). To date, more than ten different phosphodiester isoenzymes are known, some of which hydrolyze cAMP, but others also the related secondary messenger cyclic guanosine monophosphate (cGMP) (Beavo and Reifsnyder, 1990; Burns et al., 1996; Taher et al., 1997). Since a cAMP-hydrolyzing phosphodiesterase, PDE3A, is inhibited by cGMP, cGMP-hydrolyzing phosphodiesterase 5 (PDE5) has itself become the focus of research into erectile dysfunctions in recent years. Sildenafil is a PDE5 inhibitor currently used under the Viagra is already successfully used as a therapeutic agent for the treatment of erectile dysfunctions (Boolell et al., 1997). However, undesirable side effects of sildenafil, such as eye flicker, headache or circulatory failure, have been described.
Aufgabe der vorliegenden Erfindung war es, ein Mittel bereitzustellen, das die genannten Nebenwirkungen vermindert oder vermeidet.The object of the present invention was to provide an agent which reduces or avoids the side effects mentioned.
Diese Aufgabe wird gelöst durch ein Arzneimittel enthaltend eine Kombination von CNP und Phosphodiesteraseinhibitoren. CNP kann auch durch Fragmente oder Derivate mit erhaltener physiologischer Wirkung des CNP ersetzt oder ergänzt sein. Als Fragmente kommen insbesondere in der CNP-Prohormonsequenz enthaltene Teilfragmente in Frage. Die Aminosäuresequenzen des CNP sowie des proCNP und des preproCNP sind nachstehend eingeblendet:This object is achieved by a medicament containing a combination of CNP and phosphodiesterase inhibitors. CNP can also be replaced or supplemented by fragments or derivatives with the physiological effect of the CNP obtained. Partial fragments contained in the CNP prohormone sequence are particularly suitable as fragments. The amino acid sequences of the CNP, proCNP and preproCNP are shown below:
Human preproCNP (Prepropeptid)Human preproCNP (prepropeptide)
MHLSQLLACALLLTLLS RPSEAKPGAPPKVPRTPPAEELAEPQAAGGGMHLSQLLACALLLTLLS RPSEAKPGAPPKVPRTPPAEELAEPQAAGGG
QKKGDKAPGGGGANLKGDRSRLLRDLRVDTKSRAAWARLLQEHPNARQKKGDKAPGGGGANLKGDRSRLLRDLRVDTKSRAAWARLLQEHPNAR
KYKGANKKGLSKGCFGLKLDRIGSMSGLGCKYKGANKKGLSKGCFGLKLDRIGSMSGLGC
Human proCNP (Propeptid)Human proCNP (propeptide)
KPGAPPKVPRTPPAEELAEPQAAGGGQKKGDKAPGGGGANLKGDRSR LLRDLRVDTKSRAAWARLLQEHPNARKYKGANKKGLSKGCFGLKLDRI GSMSGLGCKPGAPPKVPRTPPAEELAEPQAAGGGQKKGDKAPGGGGANLKGDRSR LLRDLRVDTKSRAAWARLLQEHPNARKYKGANKKGLSKGCFGLKLDRI GSMSGLGC
Human CNP (reifes Peptid)Human CNP (mature peptide)
GLSKGCFGLKLDRIGSMSGLGC
Als Derivate kommen insbesondere solche von der wiedergegebenen Formel abgeleitete Strukturen in Betracht, die durch konservativen Austausch von Aminosäuren in dieser Sequenz entstehen. Des weiteren kommen amidierte, phosphorylierte, acylierte, alkylierte und/oder Verknüpfungen von Seitenketten enthaltende Derivate des CNP in Frage.GLSKGCFGLKLDRIGSMSGLGC Suitable derivatives are, in particular, those structures derived from the formula given which arise from the conservative exchange of amino acids in this sequence. Derivatives of the CNP containing amidated, phosphorylated, acylated, alkylated and / or linkages of side chains are also suitable.
Auch durch Deletion an der Funktion nicht beteiligter Aminosäuren werden Fragmente erhalten, die erfindungsgemäß als Arzneimittel eingesetzt werden können.Fragments are also obtained by deletion of amino acids which are not involved in the function and can be used as medicaments according to the invention.
Vorzugsweise enthält das Arzneimittel weiterhin notwendige Hilfsund/oder Trägerstoffe, die die galenische Applikation ermöglichen oder unterstützen. Bei intravenaler Applikation des erfindungsgemäßen Arzneimittels empfiehlt sich die Lösung der betreffenden Komponenten in entsprechenden zur intravenösen Verabreichung geeigneten Zubereitungen. Weitere Applikationsformen sind oral, intracavernös, intraurethral, lokal topisch, transcutan.The medicament preferably also contains necessary auxiliaries and / or carriers which enable or support the galenic application. When the medicament according to the invention is administered intravenously, it is advisable to dissolve the components in question in appropriate preparations suitable for intravenous administration. Other forms of administration are oral, intracavernous, intraurethral, locally topical, transcutaneous.
Als Phosphodlesteraseinhibitoren kommen insbesondere diejenigen der PDE5-Phosphodiesterase in Betracht. Dazu gehören insbesondere Zapri- nast, Dipyridamol, Milrinon und/oder Sildenafil.Particularly suitable phosphodlesterase inhibitors are those of PDE5 phosphodiesterase. These include, in particular, Zaprinast, Dipyridamol, Milrinon and / or Sildenafil.
Erfindungsgemäß wird die Verwendung von CNP oder Fragmenten oder Derivaten des CNP mit erhaltener physiologischer Wirkung des CNP zur Behandlung von erektilen Dysfunktionen beansprucht. In einer vorteilhaften Ausgestaltung der erfindungsgemäßen Verwendung werden CNP oder Fragmente oder Derivate des CNP mit erhaltener physiologischer
Wirkung des CNP in Kombination mit Phosphodiesteraseinhibitoren eingesetzt.According to the invention, the use of CNP or fragments or derivatives of the CNP with preserved physiological effect of the CNP for the treatment of erectile dysfunctions is claimed. In an advantageous embodiment of the use according to the invention, CNP or fragments or derivatives of CNP with a physiological value obtained Effect of CNP used in combination with phosphodiesterase inhibitors.
Erfindungsgemäß wird eine Erhöhung des intrazellulären cAMP-Spiegels durch die Stimulation der Bildung (nicht der Hemmung der Hydrolyse) des PDE3A-Hemmstoffes cGMP erreicht.According to the invention, an increase in the intracellular cAMP level is achieved by stimulating the formation (not the inhibition of hydrolysis) of the PDE3A inhibitor cGMP.
Bei Tests mit männlichen Patienten führte sowohl die intravenale, als auch die intracavernale Applikation von CNP zu einer sofortigen Erektion, die ca. eine Stunde lang anhielt. Erstaunlicherweise ergab sich bei der kombinierten Anwendung mit Phosphodiesterasehemmem wie Sil- denafil, daß deren für die Erzeugung einer Erektion notwendigen Dosen durch die gleichzeitige Applikation von CNP deutlich gesenkt werden konnten. Dabei wurden bei einigen Probanden auch die bei ausschließlicher Sildenafil-Behandlung auftretenden Nebenwirkungen wie Augen- flimmem oder Kopfschmerzen teilweise bis zum vollständigen Verschwinden verringert.In tests on male patients, both intravenous and intracavernal application of CNP resulted in an immediate erection that lasted for about an hour. Surprisingly, when used in combination with phosphodiesterase inhibitors such as sildenafil, it was found that the doses required to produce an erection could be significantly reduced by the simultaneous application of CNP. In some of the subjects, the side effects such as flare-ups or headaches that occurred with exclusive sildenafil treatment were partially reduced until they completely disappeared.
Die Figur zeigt die Wirkung von CNP und ANP/CDD99-126 auf Präparationen glatter Muskulatur des humanen Corpus cavernosum penis in Form der Wirkung von CNP und ANP/CDD99-126 auf die Relaxation vorkontrahierter Präparate glatter Muskulatur. Im Gegensatz zu ANP/CDD99-126 zeigt CNP bereits bei einer Konzentration von 1 nM einen signifikanten relaxierenden Effekt, der bei Erhöhung auf 1 μM schließlich zu einer Relaxation entsprechend 60% des nach Norepinephringabe bestehenden Tonus führt.
Beispiel 1The figure shows the effect of CNP and ANP / CDD99-126 on smooth muscle preparations of human corpus cavernosum penis in the form of the effect of CNP and ANP / CDD99- 1 26 vorkontrahierter on the relaxation of smooth muscle preparations. In contrast to ANP / CDD99-126, CNP already shows a significant relaxing effect at a concentration of 1 nM, which finally leads to a relaxation corresponding to 60% of the tone existing after norepinephrine administration when increased to 1 μM. example 1
Bei den die Synthese von cGMP katalysierenden Enzymen, den Guanylatzyklasen, ist zwischen zwei verschiedenenen Untergruppen, nämlich den löslichen zytosolischen und den membranständigen Guanylatzyklasen zu unterscheiden (zur Übersicht siehe Wedel und Garbers, 1997). Bisher wurde davon ausgegangen, daß bei der Auslösung der Erektion ausschließlich die löslichen zytosolischen Guanylatzyklasen eine Rolle spielen. Um eine eventuelle Beteiligung membranständiger Guanylatzyklasen zu überprüfen, wurde in einem ersten Schritt die Expression der Gene für die membranständigen Guanylatzyklasen GC-A, GC-B und GC-C sowie für zwei retinale membranständige Guanylatzyklasen (Datenbank-Identifier: HSRETGC und HSRETCG) in Corpus cavernosum - Gewebe wie folgt mit Hilfe der RT-PCR-Technik überprüft:In the enzymes that catalyze the synthesis of cGMP, the guanylate cyclases, a distinction must be made between two different subgroups, namely the soluble cytosolic and the membrane-bound guanylate cyclases (for an overview, see Wedel and Garbers, 1997). So far it was assumed that only the soluble cytosolic guanylate cyclases play a role in triggering the erection. In order to check the possible involvement of membrane-bound guanylate cyclases, the expression of the genes for the membrane-bound guanylate cyclases GC-A, GC-B and GC-C as well as for two retinal membrane-bound guanylate cyclases (database identifier: HSRETGC and HSRETCG) was carried out in the corpus cavernosum - tissue checked as follows using the RT-PCR technique:
Aus Biopsieproben von humanem Corpus cavernosum erfolgte die Isolierung von Gesamt-RNA und die anschließende Umschreibung der in 5 μg Gesamt-RNA enthaltenen Boten-RNA (mRNA) in Copy-DNA- Erststrang (cDNA-Erststrang) wie bereits beschrieben (Magert et al., 1995; Magert et al., 1998). Je 1/300 der cDNA-Erststrangansätze wurden anschließend in Standard-RT-PCR-Reaktionen (Saiki et al., 1988) mit den aus cDNA-Datenbankeintragungen abgeleiteten, für die verschiedenen Guanylatzyklasen spezifischen Primerpaaren eingesetzt. Die Reaktionsbedingungen waren dabei wie folgt:Total RNA was isolated from biopsy samples from human corpus cavernosum and the subsequent transcription of the messenger RNA (mRNA) contained in 5 μg total RNA into copy DNA first strand (cDNA first strand) as already described (Magert et al. , 1995; Magert et al., 1998). Each 1/300 of the first-strand cDNA batches were then used in standard RT-PCR reactions (Saiki et al., 1988) with the primer pairs derived from cDNA database entries and specific for the different guanylate cyclases. The reaction conditions were as follows:
1 X Denaturierung 94°C 5 min
danach je 35 X Denaturierung 98°C 1 sec1 x denaturation 94 ° C 5 min then 35 x denaturation 98 ° C for 1 sec
Primer-Hybridisierung 50°C 30 secPrimer hybridization 50 ° C 30 sec
Elongation 72°C 1 minElongation 72 ° C 1 min
abschließend 1 X Elongation 72°C 7 minfinally 1 X elongation 72 ° C 7 min
Die erhaltenen Produkte wurden elektrophoretisch in einem Agarosegel getrennt und nach Ethidiumbromidfärbung mit Hilfe von Ultraviolettem Durchlicht sichtbar gemacht.The products obtained were separated electrophoretically in an agarose gel and, after staining with ethidium bromide, visualized using ultraviolet transmitted light.
Tatsächlich konnte mit den für die Guanylatzyklase B (GC-B) spezifischen Primern (Sense-Primer GCB-1S: GGGTGCTGTGGCCTCTGG I I I I I CGG, Antisense-Primer GCB-2AS: CCCTGCATC I I I I CCACAATTCGAAG) eine homogene Bande im entsprechenden Größenbereich erhalten werden. Die Isolierung des enspre- chenden DNA-Fragmentes aus dem Agarosegel, die anschließende Sequenzierung mit einem DNA-Fluoeszenzsequenzer unter Verwendung der zur Amplifikation eingesetzten Primer und der Vergleich der erhaltenen Sequenz mit den Eintragungen der EMBL/GenBank-Datenbanken bestätigten es endgültig als GC-B spezifisch.In fact, with the primers specific for guanylate cyclase B (GC-B) (sense primer GCB-1S: GGGTGCTGTGGCCTCTGG I I I I I CGG, antisense primer GCB-2AS: CCCTGCATC I I I CCACAATTCGAAG) a homogeneous band could be obtained. The isolation of the corresponding DNA fragment from the agarose gel, the subsequent sequencing with a DNA fluorescence sequencer using the primers used for amplification and the comparison of the sequence obtained with the entries in the EMBL / GenBank databases finally confirmed it as GC-B specific.
Beispiel 2Example 2
Da bekannt ist, daß die GC-B den Rezeptor für das im Gehirn gebildete Peptidhormon C-Typ Natriuretisches Polypeptid (CNP) darstellt, wurde wie folgt überprüft, ob humanes CNP relaxierende Effekte auf die glatte Muskulatur des Corpus cavernosum ausübt bzw. eine Erhöhung der
intrazellulären Konzentration zyklischer Nucleosidmonophosphate indu- zuiert:Since it is known that GC-B is the receptor for the peptide hormone C-type natriuretic polypeptide (CNP) formed in the brain, it was checked as follows whether human CNP has relaxing effects on the smooth muscles of the corpus cavernosum or an increase in intracellular concentration of cyclic nucleoside monophosphates induced:
Funktionelle Organbad-Studien:Functional organ bath studies:
Streifenpräparate humaner cavernöser Muskulatur wurden in den mit einer modifizierten KREBS-RINGER-Lösung (NaCI 120 mM, NaHC03 25,6 mM, KCI 4,7 mM, CaCI2 2,5 mM, NaH2P04 1,2 mM, MgCI2 1,2 mM, Glu- cose 22 mM, 2Na+ (Ca2+) EDTA 0,1 mM, (pH 7,2 - 7,4) gefüllten Badkammern einer Organbad-Anlage (Isolated Organ Apparatus IOA 5306, Föhr Medical Instruments GmbH, Seeheim, Deutschland) fixiert. Die Badlösung war auf 37°C temperiert und wurde kontinuierlich mit einem Gemisch aus 95% O2 und 5% CO2 begast. Dem Anlegen einer passiven Vorspannung von 5 mN (0,5 g) folgte eine Äquillibrierungsphase von 60 min. Anschließend wurden die Muskelstreifen mit 6 μM Norepinephrin (NOR) stimuliert. Nach dem Erreichen stabiler Kontraktionsplateaus wurden die in unserem Hause chemisch synthetisierten Peptide human Atrial Natriuretic Peptid / Cardiodilatin (ANP/CDD99-126) (zur Übersicht siehe Forssmann et al., 1998) und human C-Type Natriuretic Peptide (CNP) (Ogawa et al., 1992) in kumulativer Dosierung (0,01 - 1000 nM) zugegeben. Mechanische Spannungs-änderungen der Muskulatur wurden mit einem analog-digitalen Meßdaten-erfassungssystem (MacLab®, AD Instruments, Castle Hill, Australien) registriert.Strip preparations of human cavernous muscles were in the with a modified KREBS-RINGER solution (NaCI 120 mM, NaHC0 3 25.6 mM, KCI 4.7 mM, CaCI 2 2.5 mM, NaH 2 P0 4 1.2 mM, MgCI 2 1.2 mM, glucose 22 mM, 2Na + (Ca 2+ ) EDTA 0.1 mM, (pH 7.2-7.4) filled bath chambers of an organ bath system (Isolated Organ Apparatus IOA 5306, Föhr Medical Instruments GmbH, Seeheim, Germany) The bath solution was heated to 37 ° C. and was continuously gassed with a mixture of 95% O2 and 5% CO 2. A passive prestress of 5 mN (0.5 g) followed The muscle strips were then stimulated with 6 μM norepinephrine (NOR) and after reaching stable contraction plateaus, the peptides human Atrial Natriuretic Peptide / Cardiodilatin (ANP / CDD99-126), which had been chemically synthesized in our company, were (see Forssmann et al., 1998) and human C-type natriuretic peptides (CNP) (Ogawa et al., 1992) in cumulative r Dosage (0.01 - 1000 nM) added. Mechanical changes in muscle tension were recorded with an analog-digital measurement data acquisition system (MacLab®, AD Instruments, Castle Hill, Australia).
Bestimmung der Gewebekonzentrationen zyklischer Nukleotide:
Zur Bestimmung zeit- und dosisabhängiger Effekte von ANP/CDD99-126 und CNP auf die intrazelluläre Konzentration von cAMP und cGMP wurden Streifen cavernöser Muskulatur analog zu den Organbad- Experimenten in 2 ml Reaktionsgefäßen mit KREBS-RINGER Lösung inkubiert, kontinuierlich mit Carbogen begast und mit 6 μM NOR stimuliert. Anschließend erfolgte die Inkubation mit ANP/CDD99-126 und CNP (0,1, 10 und 1000 nM) für 10 min. Diese erfolgte in Gegenwart von 1 mM Isobutylmethylxanthin (IBMX) um die Degradierung zyklischer Nukleotide durch die Aktivität von Phosphodiesterasen zu verhindern. Nach dem Ende der Inkubationszeit wurden die Gewebestreifen in vorbereitete Reaktionsgefäße transferiert, in flüssigem Stickstoff schockgefroren und anschließend pulverfein homogenisiert. Die Extraktion zyklischer Nukleotide erfolgte durch Resuspendierung der Gewebeho- mogenate in 1 ml 70% Ethanols, Sonifizierung der Suspensionen für 7 min und anschließender Zentrifugation (10 min bei 1.700 g). Die alkoholischen Überstande wurden abgezogen und gefriergetrocknet, die Lyophillisate in 1 ml RIA-Puffer (50 mM NaAc pH 6.0) aufgenommen. Jeweils 200 μl der resuspendierten Proben wurden in Reaktionsgefäßen vorgelegt und mit jeweils 6.7 μl eines Gemisches aus Triethyiamin/ Essigsäure (Verhältnis 2: 1) acetyliert. Die Quantifizierung von cAMP und cGMP erfolgte mit radioimmunchemischen Methoden unter Verwendung von Kaninchen-Antikörpern gegen acetyliertes cAMP und cGMP.Determination of tissue concentrations of cyclic nucleotides: To determine the time and dose-dependent effects of ANP / CDD99-126 and CNP on the intracellular concentration of cAMP and cGMP, strips of cavernous musculature were incubated with KREBS-RINGER solution in 2 ml reaction vessels analogous to the organ bath experiments, continuously gassed with carbogen and with 6 μM NOR stimulated. Subsequently, the incubation with ANP / CDD9 9 was - 12 6 and CNP (0.1, 10 and 1000 nM) for 10 min. This was done in the presence of 1 mM isobutylmethylxanthine (IBMX) to prevent the degradation of cyclic nucleotides by the activity of phosphodiesterases. After the end of the incubation period, the tissue strips were transferred to prepared reaction vessels, snap-frozen in liquid nitrogen and then homogenized to a fine powder. Cyclic nucleotides were extracted by resuspending the tissue homogenates in 1 ml of 70% ethanol, sonifying the suspensions for 7 min and then centrifuging (10 min at 1,700 g). The alcoholic supernatants were drawn off and freeze-dried, and the lyophilisates were taken up in 1 ml of RIA buffer (50 mM NaAc pH 6.0). In each case 200 μl of the resuspended samples were placed in reaction vessels and acetylated with 6.7 μl of a mixture of triethyiamine / acetic acid (ratio 2: 1). The quantification of cAMP and cGMP was carried out by radioimmunochemical methods using rabbit antibodies against acetylated cAMP and cGMP.
In den Testreihen mit verschiedenen Peptidkonzentrationen konnte schon ab 1 nM CNP eine signifikante Relaxation erreicht werden, die bei Erhöhung auf 1 μM sogar 60% der Ausgangstension entsprach. Mit ANP/CDD99-126 (dem Liganden der Guanylatzyklase A) zeigte sich in
entsprechenden Konzentrationen keine signifikante Relaxation, was die Bedeutung von CNP und seines Rezeptors, der Guanylatzyklase B, für die Tonusregulation der glatten Muskulatur des Penis bestätigt (siehe Fig.).In the test series with different peptide concentrations, a significant relaxation was achieved from 1 nM CNP, which even increased to 60% of the initial tension when increased to 1 μM. With ANP / CDD99- 12 6 (the ligand of guanylate cyclase A) showed in corresponding concentrations, no significant relaxation, which confirms the importance of CNP and its receptor, guanylate cyclase B, for the tone regulation of the smooth muscles of the penis (see FIG.).
Referenzen:Credentials:
Beavo, J.A. und Reifsnyder, D.H. (1990). Primary sequence of cyclic nucleotide phosphodiesterase isoenzymes and the design of selective inhibitors. Trends Pharmac. Sei. 11: 150-155.Beavo, J.A. and Reifsnyder, D.H. (1990). Primary sequence of cyclic nucleotide phosphodiesterase isoenzymes and the design of selective inhibitors. Trends Pharmac. Be. 11: 150-155.
Boolell, M., Allen, M.J., Ballard, S.A., Gepi-Attee, S., Muirhead, G.J., Naylor, A.M., Osterloh, I.H. und Gingell, C. (1997). Sildenafil: an orally active type 5 cyclic GMP-specific phosphodiesterase inhibitor for the treatment of penile erectile dysfunetion. Int. J. Impot. Res. 8: 47-52.Boolell, M., Allen, M.J., Ballard, S.A., Gepi-Attee, S., Muirhead, G.J., Naylor, A.M., Osterloh, I.H. and Gingell, C. (1997). Sildenafil: an orally active type 5 cyclic GMP-specific phosphodiesterase inhibitor for the treatment of penile erectile dysfunction. Int. J. Impot. Res. 8: 47-52.
Burns, F., Zhao, A.Z. und Beavo, J.A. (1996). Cyclic nucleotide phosphodiesterases: gene complexity, regulation by phosphorylation, and physiological implications. Adv. Pharmacol. 36: 29-48.Burns, F., Zhao, A.Z. and Beavo, J.A. (1996). Cyclic nucleotide phosphodiesterases: gene complexity, regulation by phosphorylation, and physiological implications. Adv. Pharmacol. 36: 29-48.
Forssmann, W.G., Richter, R. und Meyer, M. (1998). The endoerine heart and natriuretic peptides: histochemistry, cell biology, and funetio- nal aspects of the renal urodilatin System. Histochem. Cell. Biol. 110: 335-357.Forssmann, W.G., Richter, R. and Meyer, M. (1998). The endoerine heart and natriuretic peptides: histochemistry, cell biology, and functional aspects of the renal urodilatin system. Histochem. Cell. Biol. 110: 335-357.
Magert, HJ., Reinecke, M., David, I., Raab, H.R., Adermann, K., Zucht, H.D., Hill, O., Hess, R. und Forssmann, W.G. (1998). Uroguanylin : Gene structure, expression, processing as a peptide hormone, and co-storage with somatostatin in gastrointestinal D-cells. Regul. Pept. 73: 165-176.
Magert, HJ., Hadrys, T., Cieslak, A., Gröger, A., Feller, S. und Forssmann, W.G. (1995). cDNA sequence and expression pattern of the pu- tative pheromone carrier aphrodisin. Proc. Natl. Acad. Sei. USA 92: 2091-2095.Magert, HJ., Reinecke, M., David, I., Raab, HR, Adermann, K., Zucht, HD, Hill, O., Hess, R. and Forssmann, WG (1998). Uroguanylin: Gene structure, expression, processing as a peptide hormone, and co-storage with somatostatin in gastrointestinal D-cells. Regul. Pept. 73: 165-176. Magert, HJ., Hadrys, T., Cieslak, A., Gröger, A., Feller, S. and Forssmann, WG (1995). cDNA sequence and expression pattern of the putative pheromone carrier aphrodisin. Proc. Natl. Acad. Be. USA 92: 2091-2095.
Ogawa, Y., Nakao, K., Nakagawa, O., Komatsu, Y., Hosoda, K., Suga, S., Arai, H., Nagata, K., Yoshida, N. und Imura, H. (1992). Human C- type natriuretic peptide. Characterization of the gene and peptide. Hy- pertension 19: 809-813.Ogawa, Y., Nakao, K., Nakagawa, O., Komatsu, Y., Hosoda, K., Suga, S., Arai, H., Nagata, K., Yoshida, N. and Imura, H. ( 1992). Human C-type natriuretic peptides. Characterization of the gene and peptide. Hypertension 19: 809-813.
Saiki, R.K., Gelfand, D.H., Stoffel, S., Scharf, S.J., Higuchi, R., Hörn, G.T., Mullis, K.B. und Ehrlich, H.A. (1998). Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science 239: 487-491Saiki, R.K., Gelfand, D.H., Stoffel, S., Scharf, S.J., Higuchi, R., Hörn, G.T., Mullis, K.B. and Ehrlich, H.A. (1998). Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science 239: 487-491
Stief, CG., Noack, T. und Anderson, K.E. (1997). Signal transduetion in cavernous smooth muscle. World J. Urol. 15: 27-31.Stief, CG., Noack, T. and Anderson, K.E. (1997). Signal transduction in cavernous smooth muscle. World J. Urol. 15: 27-31.
Taher, A., Meyer, M., Stief, CG., Jonas, U. und Forssmann, W.G. (1997). Cyclic nucleotide phosphodiesterase in human cavernous smooth muscle. World J. Urol. 15: 32-35.Taher, A., Meyer, M., Stief, CG., Jonas, U. and Forssmann, W.G. (1997). Cyclic nucleotide phosphodiesterase in human cavernous smooth muscle. World J. Urol. 15: 32-35.
Wedel, B.J. und Garbers, D.L. (1997). New insights on the functions of the guanylyl cyclase reeeptors. FEBS Lett. 410: 29-33.
Wedel, B.J. and Garbers, D.L. (1997). New insights on the functions of the guanylyl cyclase reeeptors. FEBS Lett. 410: 29-33.
Claims
1. Arzneimittel enthaltend eine Kombination von CNP oder Fragmenten oder Derivaten mit erhaltener physiologischer Wirkung des CNP und Phosphodiesteraseinhibitoren.1. Medicament containing a combination of CNP or fragments or derivatives with preserved physiological effect of the CNP and phosphodiesterase inhibitors.
2. Arzneimittel nach Anspruch 1 enthaltend weiterhin Hilfs- und/oder Trägerstoffe.2. Medicament according to claim 1 further comprising auxiliaries and / or carriers.
3. Arzneimittel nach Anspruch 2, wobei mittels der Hilfs- und/oder Trägerstoffe das Arzneimittel intravenös, oral, intracavernös, intraurethral, lokal topisch, transcutan verabreichbar ist.3. Medicament according to claim 2, wherein by means of the excipients and / or carriers, the medicament can be administered intravenously, orally, intracavernously, intraurethrally, locally topically, transcutaneously.
4. Arzneimittel nach einem der Ansprüche 1 bis 3, wobei die Phosphodiesteraseinhibitoren Zaprinast, Dipyridamol, Milrinon und/oder Silde- nafil sind.4. Medicament according to one of claims 1 to 3, wherein the phosphodiesterase inhibitors are zaprinast, dipyridamole, milrinone and / or sildenafil.
5. Verwendung von CNP oder Fragmenten oder Derivaten des CNP mit erhaltener physiologischer Wirkung des CNP zur Behandlung von erektilen Dysfunktionen5. Use of CNP or fragments or derivatives of the CNP with preserved physiological effect of the CNP for the treatment of erectile dysfunctions
6. Verwendung nach Anspruch 5 in Kombination mit Phosphodiesteraseinhibitoren. 6. Use according to claim 5 in combination with phosphodiesterase inhibitors.
Applications Claiming Priority (3)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| DE19903087 | 1999-01-27 | ||
| DE19903087A DE19903087A1 (en) | 1999-01-27 | 1999-01-27 | Treatment of erectile dysfunction with C-type natriuretic polypeptide (CNP) as monotherapy or in combination with phosphodiesterase inhibitors |
| PCT/EP2000/000634 WO2000044399A2 (en) | 1999-01-27 | 2000-01-27 | Treatment of erectile dysfunctions with a c-type natriuretic polypeptide (cnp) as monotherapy or in combination with phosphodiesterase inhibitors |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| EP1148888A2 true EP1148888A2 (en) | 2001-10-31 |
Family
ID=7895476
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| EP00909112A Withdrawn EP1148888A2 (en) | 1999-01-27 | 2000-01-27 | Treatment of erectile dysfunctions with a c-type natriuretic polypeptide (cnp) as monotherapy or in combination with phosphodiesterase inhibitors |
Country Status (7)
| Country | Link |
|---|---|
| US (1) | US20020128176A1 (en) |
| EP (1) | EP1148888A2 (en) |
| JP (1) | JP2002536299A (en) |
| AU (1) | AU3150800A (en) |
| CA (1) | CA2361239A1 (en) |
| DE (1) | DE19903087A1 (en) |
| WO (1) | WO2000044399A2 (en) |
Families Citing this family (16)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| TWI265925B (en) | 1999-10-11 | 2006-11-11 | Pfizer | Pyrazolo[4,3-d]pyrimidin-7-ones useful in inhibiting type 5 cyclic guanosine 3',5'-monophosphate phosphodiesterases(cGMP PDE5), process and intermediates for their preparation, their uses and composition comprising them |
| US20030139384A1 (en) * | 2000-08-30 | 2003-07-24 | Dudley Robert E. | Method for treating erectile dysfunction and increasing libido in men |
| NZ524601A (en) * | 2000-08-30 | 2006-04-28 | Unimed Pharmaceuticals Inc | Method for treating erectile dysfunction and increasing libido in men |
| US6503894B1 (en) | 2000-08-30 | 2003-01-07 | Unimed Pharmaceuticals, Inc. | Pharmaceutical composition and method for treating hypogonadism |
| US7041786B2 (en) * | 2001-03-29 | 2006-05-09 | Callisto Pharmaceuticals | Guanylate cyclase receptor agonists for the treatment of tissue inflammation and carcinogenesis |
| ZA200803087B (en) | 2005-10-12 | 2009-11-25 | Unimed Pharmaceuticals Llc | Improved testosterone gel and method of use |
| AU2008261102B2 (en) | 2007-06-04 | 2013-11-28 | Bausch Health Ireland Limited | Agonists of guanylate cyclase useful for the treatment of gastrointestinal disorders, inflammation, cancer and other disorders |
| US8207295B2 (en) * | 2008-06-04 | 2012-06-26 | Synergy Pharmaceuticals, Inc. | Agonists of guanylate cyclase useful for the treatment of gastrointestinal disorders, inflammation, cancer and other disorders |
| US8969514B2 (en) | 2007-06-04 | 2015-03-03 | Synergy Pharmaceuticals, Inc. | Agonists of guanylate cyclase useful for the treatment of hypercholesterolemia, atherosclerosis, coronary heart disease, gallstone, obesity and other cardiovascular diseases |
| KR20100059859A (en) * | 2007-09-11 | 2010-06-04 | 몬도바이오테크 래보래토리즈 아게 | Use of c-type natriuretic peptide, alone or incombination with neuropeptide af, as a therapeutic agent |
| CA2930674A1 (en) * | 2008-06-04 | 2009-12-10 | Synergy Pharmaceuticals Inc. | Agonists of guanylate cyclase useful for the treatment of gastrointestinal disorders, inflammation, cancer and other disorders |
| JP2011528375A (en) * | 2008-07-16 | 2011-11-17 | シナジー ファーマシューティカルズ インコーポレイテッド | Guanylate cyclase agonists useful for the treatment of gastrointestinal disorders, inflammation, cancer, and other disorders |
| JP5964589B2 (en) | 2008-12-03 | 2016-08-03 | シナジー ファーマシューティカルズ インコーポレイテッド | Preparation of guanylate cyclase C agonist and method of use thereof |
| CA2810243C (en) | 2010-09-15 | 2021-04-20 | Synergy Pharmaceuticals Inc. | Formulations of guanylate cyclase c agonists and methods of use |
| US9616097B2 (en) | 2010-09-15 | 2017-04-11 | Synergy Pharmaceuticals, Inc. | Formulations of guanylate cyclase C agonists and methods of use |
| CA2905438A1 (en) | 2013-03-15 | 2014-09-25 | Synergy Pharmaceuticals Inc. | Agonists of guanylate cyclase and their uses |
Family Cites Families (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CA1319099C (en) * | 1987-01-23 | 1993-06-15 | James A. Nathanson | Atriopeptins, guanylate cyclase activators, and phosphodiesterase inhibitors as treatment for glaucoma, hydrocephalus and cerebral edema (cranial fluid volume dysfunction) |
| JP2809533B2 (en) * | 1991-01-31 | 1998-10-08 | 壽之 松尾 | CNP analog peptide |
| JP3418405B2 (en) * | 1996-05-10 | 2003-06-23 | アイコス コーポレイション | Carboline derivative |
-
1999
- 1999-01-27 DE DE19903087A patent/DE19903087A1/en not_active Ceased
-
2000
- 2000-01-27 AU AU31508/00A patent/AU3150800A/en not_active Abandoned
- 2000-01-27 CA CA002361239A patent/CA2361239A1/en not_active Abandoned
- 2000-01-27 EP EP00909112A patent/EP1148888A2/en not_active Withdrawn
- 2000-01-27 WO PCT/EP2000/000634 patent/WO2000044399A2/en not_active Application Discontinuation
- 2000-01-27 JP JP2000595701A patent/JP2002536299A/en active Pending
-
2001
- 2001-07-26 US US09/912,425 patent/US20020128176A1/en not_active Abandoned
Non-Patent Citations (1)
| Title |
|---|
| See references of WO0044399A2 * |
Also Published As
| Publication number | Publication date |
|---|---|
| WO2000044399A3 (en) | 2000-12-14 |
| JP2002536299A (en) | 2002-10-29 |
| CA2361239A1 (en) | 2000-08-03 |
| US20020128176A1 (en) | 2002-09-12 |
| WO2000044399A2 (en) | 2000-08-03 |
| AU3150800A (en) | 2000-08-18 |
| DE19903087A1 (en) | 2000-08-10 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| WO2000044399A2 (en) | Treatment of erectile dysfunctions with a c-type natriuretic polypeptide (cnp) as monotherapy or in combination with phosphodiesterase inhibitors | |
| Corbin et al. | Phosphodiesterase type 5 as a pharmacologic target in erectile dysfunction | |
| De Souza et al. | Corticotropin-releasing factor: Physiology, pharmacology, and role in central nervous system disorders | |
| Rehman et al. | Diminished neurogenic but not pharmacological erections in the 2-to 3-month experimentally diabetic F-344 rat | |
| Klinger et al. | Pulmonary hemodynamic responses to brain natriuretic peptide and sildenafil in patients with pulmonary arterial hypertension | |
| KR20090047546A (en) | Combination treatment for diabetes mellitus | |
| KR20070036079A (en) | Multiple agent therapy for sexual dysfunction | |
| DE10012373A1 (en) | Treating erectile dysfunction using new or known imidazo(1,5-a)pyrido(3,2-e)pyrazinone derivatives, as cGMP-selective phosphodiesterase V inhibitors free of cardiovascular side-effects | |
| Gann et al. | The neuroendocrine response to multiple trauma | |
| DE102005042877A1 (en) | Use of PDE1A polypeptides, or nucleic acid, for identifying their specific inhibitors, which are useful for treatment and prevention of cardiac insufficiency | |
| Takagi et al. | α-Human atrial natriuretic peptide, carperitide, reduces infarct size but not arrhythmias after coronary occlusion/reperfusion in dogs | |
| Eardley | The role of phosphodiesterase inhibitors in impotence | |
| Mondritzki et al. | New pulmonary hypertension model in conscious dogs to investigate pulmonary-selectivity of acute pharmacological interventions | |
| Saussine et al. | Renin stimulating properties of parathyroid hormone-related peptide in the isolated perfused rat kidney | |
| JPH08501296A (en) | Methods for treating renin-related disorders with amylin antagonists | |
| Abd-Elfattah et al. | Myocardial protection in beating heart cardiac surgery: I: pre-or postconditioning with inhibition of es-ENT1 nucleoside transporter and adenosine deaminase attenuates post-MI reperfusion-mediated ventricular fibrillation and regional contractile dysfunction | |
| Monteiro et al. | Adenosine deaminase and adenosine uptake inhibitions facilitate ventilation in rats | |
| US6465465B1 (en) | Process for the treatment of erectile dysfunction and product therefor | |
| Vesely | Cardiac and renal hormones: anticancer effects in vitro and in vivo | |
| DE10002200A1 (en) | Treatment of erectile dysfunction by administration of a stimulator of guanylate cyclase, e.g. C-Type natriuretic polypeptide, optionally with a phosphodiesterase inhibitor | |
| Chiu et al. | Effect of sildenafil on renin secretion in human subjects | |
| Dundore et al. | Sodium nitroprusside potentiates the depressor response to the phosphodiesterase inhibitor zaprinast in rats | |
| Németh et al. | Impairment of neurogenic inflammatory and anti-inflammatory responses in diabetic rats | |
| DE10004289A1 (en) | Treatment of erectile dysfunction without causing undesirable side-effects, using phosphodiesterase IV inhibitor, optionally in combination with other phosphodiesterase inhibitor or adenylate cyclase stimulant | |
| Kellner et al. | C-type natriuretic peptide exerts stimulatory effects on the corticotropin-releasing hormone-induced secretion of hormones in normal man |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PUAI | Public reference made under article 153(3) epc to a published international application that has entered the european phase |
Free format text: ORIGINAL CODE: 0009012 |
|
| 17P | Request for examination filed |
Effective date: 20010721 |
|
| AK | Designated contracting states |
Kind code of ref document: A2 Designated state(s): AT BE CH CY DE DK ES FI FR GB GR IE IT LI LU MC NL PT SE |
|
| AX | Request for extension of the european patent |
Free format text: AL;LT;LV;MK;RO;SI |
|
| 17Q | First examination report despatched |
Effective date: 20020729 |
|
| STAA | Information on the status of an ep patent application or granted ep patent |
Free format text: STATUS: THE APPLICATION IS DEEMED TO BE WITHDRAWN |
|
| 18D | Application deemed to be withdrawn |
Effective date: 20021210 |