WO1998004686A2 - Enzyme de conversion de preproteines - Google Patents

Enzyme de conversion de preproteines Download PDF

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Publication number
WO1998004686A2
WO1998004686A2 PCT/CA1997/000535 CA9700535W WO9804686A2 WO 1998004686 A2 WO1998004686 A2 WO 1998004686A2 CA 9700535 W CA9700535 W CA 9700535W WO 9804686 A2 WO9804686 A2 WO 9804686A2
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Prior art keywords
human
renin
prorenin
protein
cells
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PCT/CA1997/000535
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English (en)
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WO1998004686A3 (fr
Inventor
Robert Day
Nabil G. Seidah
Rémi Martel
Michel Chretien
Tim Reudelhuber
Guy Leclerc
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Robert Day
Seidah Nabil G
Martel Remi
Michel Chretien
Tim Reudelhuber
Guy Leclerc
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Priority claimed from CA002203745A external-priority patent/CA2203745A1/fr
Application filed by Robert Day, Seidah Nabil G, Martel Remi, Michel Chretien, Tim Reudelhuber, Guy Leclerc filed Critical Robert Day
Priority to EP97932685A priority Critical patent/EP0920494A2/fr
Priority to JP50834398A priority patent/JP2001505870A/ja
Priority to US09/214,555 priority patent/US6380171B1/en
Priority to AU36170/97A priority patent/AU3617097A/en
Publication of WO1998004686A2 publication Critical patent/WO1998004686A2/fr
Publication of WO1998004686A3 publication Critical patent/WO1998004686A3/fr

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/48—Hydrolases (3) acting on peptide bonds (3.4)
    • C12N9/50—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25)
    • C12N9/64—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25) derived from animal tissue
    • C12N9/6421—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25) derived from animal tissue from mammals
    • C12N9/6424—Serine endopeptidases (3.4.21)
    • C12N9/6454—Dibasic site splicing serine proteases, e.g. kexin (3.4.21.61); furin (3.4.21.75) and other proprotein convertases
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P7/00—Drugs for disorders of the blood or the extracellular fluid
    • A61P7/02—Antithrombotic agents; Anticoagulants; Platelet aggregation inhibitors
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides

Definitions

  • This invention relates to protein processing enzymes or pro-hormone convertases (PCs), specifically to PC5 , more specifically to the human PC5.
  • PCs protein processing enzymes or pro-hormone convertases
  • PCs Pro-hormone convertases
  • Renin is an aspartyl protease which makes an important contribution to cardiovascular physiology and pathophysiology through its key role in the synthesis of the vasoactive octapeptide angiotensin II (All) . While the kidney is the primary source of circulating active renin, several additional tissues, including the pituitary and adrenal glands, placenta, uterus, ovary, testes, heart, vasculature and brain express the renin gene (reviewed in 1 ⁇ 4 ) .
  • RAS renin- angiotensin system
  • ACE angiotensin converting enzyme
  • tRAS locally active tissue renin-angiotensin system
  • prorenin which is converted to active renin by the proteolytic removal of a 43 amino acid amino-terminal prosegment.
  • the activity of the RAS within any given tissue would, therefore, be dependent on the existence of proteolytic enzymes capable of converting prorenin to active renin and on the expression of such prorenin processing enzymes (PPEs) in the same cells that express prorenin.
  • PPEs prorenin processing enzymes
  • the identity of the enzyme (s) responsible for the proteolytic activating human prorenin in vivo is still uncertain. Furthermore, it is possible that multiple PPEs exist in humans and these may differ among renin-producing tissues.
  • prohor one convertase PCI has also been shown to cleave human prorenin with the correct site-and organelle specificity in transfected cells 10 and to co-localize with renin in the adrenal medulla and derived tumors 11 , but not in JG cells. 12
  • Mouse PC5 is capable of partially cleaving human prorenin.
  • PCs platelet-derived-growth factors A and B
  • EGF epidermal -growth-factor
  • IGF's insulin-like growth factors I and II
  • TGF's transforming growth factors ⁇ and ⁇
  • PDGF's platelet-derived-growth factors A and B
  • EGF epidermal -growth-factor
  • IGF's insulin-like growth factors I and II
  • TGF's transforming growth factors ⁇ and ⁇
  • Full biological potency is conferred to these growth factors only after cleavage at these sites, by one or more of the PC enzyme family. There is therefore a possibility that manipulating the expression of the PCs would affect cell proliferation via deficient growth factor activation.
  • PTCA vascular endothelial growth factor
  • the present invention relates to the human PC5 (hPC5) .
  • hPC5 human PC5
  • PC5 is a prorenin-processing enzyme (PPE) .
  • PPE prorenin-processing enzyme
  • Silencing the expression of PC5 would find a specific application in inhibiting the production of renin, and a method of inhibiting the production of renin is an object of the invention. Since the production of renin is one of targets of the RAS involved in hypertension.
  • hPC5 is overexpressed in atherosclerotic coronary arteries.
  • Antisense oligonucleotides have been designed, amongst which one has been shown to successfully silence the expression of hPC5 in smooth muscle cells in culture. This antisense inhibited carotid stenosis in a in vivo rabbit carotid injury model. These results indicate that a method of silencing the expression of PC5 would find a specific application in preventing restenosis.
  • PC5 is known to be expressed in CD4 + T cells, along with furin and PC7.
  • the three enzymes are capable of converting HIV gpl60 into its fusiogenic form. Therefore, antisense constructs, particularly the oligonucleotide that successfully inhibited restenosis, will find a use in inhibiting expression of the activity of PC5 towards HIV gpl60.
  • the complete amino acid and nucleotide sequence of hPC5 is described hereinbelow and are another object of this invention.
  • Recombinant vectors and hosts comprising as a new insert, whole or part of hPC5 are also an object of the invention.
  • Oligopeptides derived from the proteic sequence of hPC5 are also an object of the invention.
  • Antibodies directed against the whole protein hPC5 or a part thereof are also an object of the invention.
  • Diagnostic methods and kits comprising oligonucleotides or antibodies binding PC5 nucleic acids or protein or peptides are also an object of the invention.
  • Figure 1 Schematic diagram of the isolated cDNAs encoding hPC5. Restriction enzyme sites used in sub-cloning are denoted. Solid lines represent clones isolated from a phage library. Hatched lines denote the portion of the cDNA isolated by RT-PCR of human adrenal mRNA. The double line represents the portion of the mouse PC5 cDNA (corresponding to the amino terminus of the signal peptide) which was used to complete the cDNA for expression.
  • Figure 2 Nucleotide and derived protein sequence of hPC5 (SEQ ID NOS: 1 and 2, respectively). Proposed signal peptide (solid arrow) and prosegment (open arrow) cleavage sites are denoted based on data from mouse PC5. 15 The underlined sequence represents the portion of the signal peptide from mouse PC5 which was used in the expression vector construction.
  • Figure 3 Distribution on PC5 RNA in various human tissues.
  • Each lane contains 2 ⁇ g poly-A RNA. Filters were hybridized with a radiolabeled probe for hPC5 as described in Materials and Methods. Shown at left is the migration of single strand size standards in kilobases (Kb) . Note that the absolute signal cannot be compared between the two filters as they were of different ages and hybridized at different times.
  • Figure 4 hPC5 cleaves human prorenj n with site and cell specificity.
  • Panel A GH ⁇ cell were co-transfected with expression vectors for the indicated proteins. Supernatants were collected 30 hrs . after transfection and assayed for % active renin [(active renin/total renin) X 100].
  • Panel B Resulting secretion of active renin after co-transfection of CHO cells with an expression vector for prorenin and either a control plasmid (pUC) or hPC5. Bars represent the mean ⁇ S.E.M of 3 independent transfections .
  • Figure 5 Active renin generation in secretory granules of co-transfected GH.C1 cells. Parallel wells of GH 4 C1 cells co- transfected with expression vectors for human prorenin and human PC5 were incubated for 20 min.
  • FIG 8 Western blot analysis of PCS protein in rabbit smooth muscle cells treated with either antisense. sense or mismatch PC5 oligonucleotides.
  • the specific PC5 band is identified (see arrow) by comparison with proteins extracted from rat supraoptic nucleus (SON) and SK-N-MCIXC cells (human neuroepithelioma) .
  • SON rat supraoptic nucleus
  • SK-N-MCIXC cells human neuroepithelioma
  • Figure 9 Rabbit i n vivo test of the PC5 antisense ODN as compared to the control sense and random ODNs . It shows a decreased stenosis due to the presence of a PC5 antisense when compared to the sense and random controls.
  • Figure 10 Proprotein convertase immunoreactivity in human atherecto y specimens. It shows the presence of the enzymes of the pro-hormone convertase family which are present in the specimens .
  • Figure 11 Illustrates differences between cDNA sequences of PC6 (Miranda et al . SEQ ID No. 4) and PC5 (present invention) .
  • Figure 12 Illustrates differences between protein sequences of PC6 (Miranda et al . SEQ ID No. 5) and PC5 (present invention) .
  • cDNA library construction and screening A cDNA library derived from total human adrenal RNA was constructed by Stratagene (La Jolla, CA) in the phage vector Uni-Zap XR. Six hundred thousand phage plaques were screened initially using radioactive probes and standard methodologies. 14 The initial hybridization probe was a 320 base pair DNA fragment derived from reverse-transcriptase PCR of human brain RNA using information derived from an unidentified human cDNA sequence tag in Genbank (Accession # M85522) with a high degree of similarity to the previously cloned mouse PC5.
  • a 1070 base pair fragment (excluding the poly A tail) was excised from hPC5A, labeled and used to re- screen an additional 600,000 phage from the cDNA library.
  • a second phage clone (hPC5B) was isolated and found to contain an 1807 base pair cDNA insert overlapping hPC5A and extending toward the 5' end of the cDNA ( Figure 1) .
  • Reverse- transcriptase PCR One microgram of poly A+ RNA from total human adrenal (Clontech Laboratories, Palo Alto, CA) was subjected to reverse transcriptase polymerase chain reaction (RT-PCR) using a published procedure 16 and the following oligonucleotides: Forward oligonucleotide; derived from a region corresponding to the signal peptide of mouse PC5. 15 An artificial Hindlll restriction enzyme cleavage site added to the 5 ' -end of the amplified fragment for the purpose of cloning is underlined: 5 ' -CCAAGCTTGGCTGCTGTGCGTGCTGGC-3 '
  • Reverse oligonucleotide derived from the 5 ' -end of the phage hPC5B.
  • An internal Bglll restriction enzyme site is underlined: 5 ' -CTGCCTCAGATCTGTAGTG- •
  • the entire RT-PCR reaction was repeated 4 times and 4 independently derived clones of the amplified fragment were sequenced and the sequences were compared.
  • Genbank accesion #U49114 represents the consensus sequence, defined as any nucleotide appearing in 3/4 clones.
  • GH 4 C1 cells were plated in 6 -well culture dishes at a density of 5X10 5 cells per well. Twenty four hours later, medium was changed and the cells were transfected by the DEAE-dextran method using a commercial kit (CellPhect Transfection kit, Pharmacia Biotech, Baie D'urfe, Quebec, Canada) according to manufacturer's instructions.
  • GH 4 C1 transfected with the human prorenin and hPC5 expression vectors were stimulated to release secretory granules by depolarization using a previously published technique. 21 Forty hrs. after co- transfection, the culture medium in parallel wells of transfected cells was replaced with pre-warmed medium supplemented to a final concentration of 50 mmol/L with either NaCl (control) or KCl (secretagogue) . The media were collected after 20 min. and assayed for renin/prorenin. A potassium-dependent increase in the percent active renin contained in cell supernatants was taken as an indication of active renin release from the secretory granules of the transfected cells. Results shown in Figure 5 represent the mean of three independent transfection experiments .
  • PC5.MAP antibody or a 1:200 dilution of a polyclonal rabbit antiserum against recombinant human prorenin.
  • immune complexes were revealed by incubation with protein A-colloidal gold (15 nm particles) synthesized from tetra-chloroauric acid (BDH) according to the method of Ghitescu and Bendayan. 22 Gold particles were enhanced for viewing in the light microscope by incubation with silver (IntenSETMM Silver Enhancement Kit, Amersham Life Science, Oakville, Ontario, Canada) and sections were counter-stained with hematoxylin and methyl green.
  • Immune complexes on human adrenal sections were detected with a 1:200 dilution of biotin- labeled donkey anti -rabbit IgG and a 1:300 dilution of streptavidin-horseradish peroxidase complex (Amersham Life Science, Oakville, Ontario, Canada) and were incubated with diaminobenzidine and hydrogen peroxide (Sigma Chemicals, St. Louis, MO) as chro ogen. All positive staining patterns were subsequently verified for specificity by omission of the first antibody.
  • PC5 RNA Northern analysis of poly A RNA from a variety of human tissues reveals a major band of approximately 6.6 Kb and a minor band at approximately 3.8 Kb (Figure 3).
  • PC5 RNA is detected in the brain, heart, placenta, lung, thyroid gland and testes and at lower levels in the skeletal muscle, kidney and pancreas, small intestine and stomach. In the adrenal gland, PC5 is particularly enriched in the cortex ( Figure 3) . Because PC5 RNA appears to be expressed in a number of tissues previously reported to contain active renin, we have tested the ability of hPC5 to cleave human prorenin in a cell co-transfection assay ( Figure 4A) .
  • RNA encoding angiotensinogen and renin have been detected in preparations from the human adrenal zona glomerulosa, fasciculata and medulla 24,25 , confirming that both renin and its substrate are synthesized within the human adrenal gland.
  • ACE inhibition or blockade of angiotensin receptors inhibits aldosterone release from human adrenal tissue explants 26 , suggesting that the local RAS plays an active role in the regulation of aldosterone secretion from the adrenal gland.
  • renin would be in the appropriate intracellular compartment to be activated by PC5 in the adrenal cortex.
  • rats transgenic for mouse Ren-2 renin [TGR (mRen-2) 27] display fulminant hypertension 29 which correlates best with the expression of the mouse prorenin in the adrenal gland.
  • the principal source of circulating active renin in humans is the JG cells of the kidney.
  • low levels of hPC5 RNA can be detected by Northern blot analysis in a sample of total kidney poly-A RNA (Fig. 3) , we were unable to localize PC5 immunostaining in kidney sections (Fig. 6) raising the possibility that PC5 is expressed at low levels in diffuse cell types in the kidney.
  • PC5 is expressed at low levels in diffuse cell types in the kidney.
  • hPC5 In the mouse, two forms of hPC5 have been predicted based on cloned cDNAs; the first would be analogous to the hPC5 cDNA described in this study and to that cloned from rat tissues 15,23 while the second, called PC6B, is extended at its 3 '-end due to a differential RNA splicing event. 35 Although the hPC5 cDNA we have cloned is only roughly 3 Kb in length, the major RNA band seen in human tissues is of approximately 6.6 Kb. The identity of the longer band hybridizing to the hPC5 probe is currently unknown.
  • PC5 RNA species exist in mammals that are extended at their 5' -ends.
  • human tissues may be particularly enriched in a homologue to PC6B which was not picked up in our screenings.
  • Recent data suggest that the alternate C-terminal tail present on PC6B may serve to retain the enzyme in the Golgi network, while the "short" form of mouse PC5 is targeted to dense core secretory granules (N.G. Seidah, unpublished) .
  • These data and the results of our co- transfection assays would suggest that the "short" form of hPC5 described here is the form which would be active in renin processing in secretory granules .
  • PC5 enzymes isolated from humans and mice show a remarkably high degree of conservation at the nucleotide and protein sequence levels. This degree of similarity is higher than that seen for the other mammalian PC enzymes which seem to diverge in the C-terminal half of the enzyme. 36,37 This high degree of sequence conservation may reflect an essential function of PC5 (and the C-terminus of PC5) in mammals.
  • PC5 is linked to smooth muscle proliferation
  • PC could be a potential target of smooth muscle cell proliferation
  • Changes in PC levels in the process of restenosis is a distinct possibility since in previous studies using animal models or cell lines, we have shown that PC levels can be regulated or even be induced.
  • human restenosed coronary tissues from patients. These tissues were screened for each of the PC mRNAs using in vivo hybridization histochemistry in order to obtain information within an anatomical context. Coronaries with partial or total occlusions demonstrated dramatically increased PC5 mRNA levels within smooth muscle tissues, whereas coronary tissue without occlusions were PC5 negative.
  • PC5 is either strongly up-regulated or induced in the human coronary arteries during the active process of stenosis (fig. 7) . To our knowledge this is the first indication that a specific PC is directly linked to smooth muscle proliferation.
  • a specific antisense oligonucleotide (ODN) was shown to drastically inhibit smooth muscle proliferation using an in vi tro model of rabbit smooth muscle in culture.
  • ODN antisense oligonucleotide
  • Table 1 Incubating rabbit smooth muscle cells with a PC5 antisense 17-mer oligonucleotide shown in Table 1 caused a dose-dependent inhibition smooth muscle proliferation with a maximal inhibitory effect of 81.6% + 1.6% at 10 mM (mean of three experiments done in quadruplicates) .
  • PC5 is decreased in the affected cells (fig. 8) .
  • Phosphorothioate antisense ODNs were synthesized on a DNA/RNA synthesizer following standard procedure (Applied Biosystems) . Conjugation of oligomers with cholesterol was achieved with 3 ' -cholesterol-VN CPG (Clontech) , a virtual nucleotide (VN) glass reagent that introduces a cholesterol label to the 3' terminus of an oligonucleotide via solid- phase synthesis.
  • VN virtual nucleotide
  • New Zealand rabbits male or female (2 Kg) were intramuscularly sedated with xylazine (2 mg/Kg) and anesthetized with ketamine (100 mg/Kg) prior to surgical exposure of left carotid artery. Segments of 10 mm of carotids were transiently isolated by temporary ligatures and rinsed with 0.9% sodium chloride via a cannula until there was no more visible evidence of blood components.
  • Carotid arteries were transfected with 80 ⁇ mol/L of antisense ODNs in a 1 cm portion either alone or conjugated to cholesterol for a period of 30 minutes. The volume infused was 100 ⁇ l, and no visible loss of volume was noted throughout the incubation period.
  • a total of 36 New Zealand white rabbit carotid arteries were injured with a 2.5 mm balloon catheter serially inflated for 1 minute to 4 , 6, 8 and 10 atm with gentle traction, allowing 45 seconds between inflations.
  • a second injury was imposed at the same arterial site which was then transfected in a 1 cm portion with 80 ⁇ mol/L (100 ⁇ l of volume injected) of therapeutic molecules or with 100 ⁇ L of NaCl 0.9% as control.
  • Intimal/medial areas were evaluated by computer analysis on histological sections derived from transfected arteries two weeks following the second injury and transfection procedure.
  • antisense oligonucleotides may be added to optimize this method of preventing restenosis, such as those silencing the expression of other convertases, namely PC2, which are also observed in atherectomy specimens (see Figure 10) .
  • PC2 convertases
  • the development of drugs based on the inhibition or the inactivation of the convertases is of great interest because the drugs can easily be delivered directly at the affected site during the intervention by the cardiologist.
  • the sequences of the oligonucleotides used are: Antisense GCAACTTGCCAGAGCAT SEQ ID NO: 3 Sense ATGCTCTGGCAAGTTGC Random AATCCGTGAGACCAGTC .
  • PC5 is involved in the cleavage of HIV gpl ⁇ O into gpl20 and gp41.
  • PC5, PC7 and furin are known to be present in CD4 + T lymphocytes. All three enzymes cleave HIV gpl60 to gpl20 and gp41 as well as a synthetic peptide covering the junction wherein cleavage occurs in gpl60. Since the 17-mer antisense ODN defined in Seq ID. No.

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Abstract

Un clone d'ADNc codant pour la prohormone convertase humaine PC5 a été isolé de l'ARN messager des surrénales humaines. La séquence protéique qui en a été déduite devrait coder pour une préprotéine PC5 de 915 acides aminés, ayant un degré très élevé d'homologie avec ses homologues du rat et de la souris précédemment codées. L'ARNm de la PC5 est détecté dans de multiples tissus humains, notamment le cerveau, les surrénales, la thyroïde, le coeur, le placenta, les poumons et les testicules. Il n'était pas détectable dans le foie et il est présent à des taux inférieurs dans les muscles squelettiques, les reins, le pancréas, l'intestin grêle et l'estomac. La co-transfection des vecteurs d'expression de la PC5 humaine et de la prorénine humaine dans des cellules GH4C1 cultivées provoque la sécrétion de rénine active. L'activation de la prorénine humaine par la PC5, qui dépend d'une paire d'acides aminés basiques aux positions 42 et 43 du prosegment de la prorénine, n'a lieu que dans des cellules contenant des granules sécrétoires à coeur dense. La PC5 humaine a été colocalisée par immunochimie avec la rénine dans la zone glomérulée des surrénales, ce qui suggère qu'elle pourrait participer à l'activation d'un système rénine-angiotensine local dans le cortex surrénal humain. La PC5 est surexprimée dans les coronaires athéroscléreuses. L'atténuation de l'expression de la PC5 par un oligonucléotide antisens spécifique a inhibé efficacement la prolifération des cellules des muscles lisses en culture. En outre, l'antisens a inhibé la sténose de la carotide dans un modèle de lésion de la carotide. Ces résultats indiquent que l'atténuation de la PC5 s'applique à la prévention de la resténose. Les PC pourraient être des cibles de choix pour le traitement de toutes les maladies prolifératives mettant en jeu leur action sur un facteur de croissance donné. Enfin, l'oligonucléotide antisens de la PC5 est à utiliser pour atténuer l'activité de cette enzyme envers la gp160 du VIH, puisqu'ils coexistent dans les lymphocytes T CD4+ et que la glycoprotéine virale peut être clivée par la PC5.
PCT/CA1997/000535 1996-07-26 1997-07-25 Enzyme de conversion de preproteines WO1998004686A2 (fr)

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Application Number Priority Date Filing Date Title
EP97932685A EP0920494A2 (fr) 1996-07-26 1997-07-25 Enzyme de conversion de preproteines
JP50834398A JP2001505870A (ja) 1996-07-26 1997-07-25 プロタンパク質変換酵素
US09/214,555 US6380171B1 (en) 1996-07-26 1997-07-25 Pro-protein converting enzyme
AU36170/97A AU3617097A (en) 1996-07-26 1997-07-25 Pro-protein converting enzyme

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US2100896P 1996-07-26 1996-07-26
US60/021,008 1996-07-26
CA002203745A CA2203745A1 (fr) 1996-07-26 1997-04-25 Enzyme de conversion pro-proteinique
CA2,203,745 1997-04-25

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Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1999034823A2 (fr) * 1998-01-09 1999-07-15 University Of Utah Procedes de prevention et de traitement des maladies fibreuses provoquees par l'accumulation excessive d'une matrice extracellulaire induite par le tgf beta a l'aide d'inhibiteurs de la renine
WO2003011328A1 (fr) * 2001-07-31 2003-02-13 Prince Henry's Institute Of Medical Research Activite enzymatique liee a la grossesse
WO2003060109A2 (fr) * 2002-01-15 2003-07-24 Bayer Healthcare Ag Regulation d'une subtilase humaine

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DECROLY E ET AL: "THE CONVERTASES FURIN AND PC1 CAN BOTH CLEAVE THE HUMAN IMMUNODEFICIENCY VIRUS (HIV)-1 ENVELOPE GLYCOPROTEIN GP160 INTO GP120 (HIV-I SU) AND GP41 (HIV-I TM)" JOURNAL OF BIOLOGICAL CHEMISTRY, vol. 269, no. 16, 22 April 1994, pages 12240-12247, XP002023318 *
HŸLÈNE C ET AL: "SPECIFIC REGULATION OF GENE EXPRESSION BY ANTISENSE, SENSE AND ANTIGENE NUCLEIC ACIDS" BIOCHIMICA ET BIOPHYSICA ACTA, vol. 1049, 1 January 1990, pages 99-125, XP000570355 *
MERCURE C. ET AL.: "Prohormone convertase PC5 is a candidate processing enzyme for prorenin in the human adrenal cortex" HYPERTENSION, vol. 28, no. 4, October 1996, DALLAS, US, pages 840-846, XP002046402 *
MIRANDA L. ET AL.: "Isolation of the human PC6 gene encoding the putative host protease for HIV-1 gp160 processing in CD4+ T lymphocytes" PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF USA, vol. 93, July 1996, WASHINGTON US, pages 7695-7700, XP002046403 cited in the application *

Cited By (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1999034823A2 (fr) * 1998-01-09 1999-07-15 University Of Utah Procedes de prevention et de traitement des maladies fibreuses provoquees par l'accumulation excessive d'une matrice extracellulaire induite par le tgf beta a l'aide d'inhibiteurs de la renine
WO1999034823A3 (fr) * 1998-01-09 1999-09-16 Univ Utah Procedes de prevention et de traitement des maladies fibreuses provoquees par l'accumulation excessive d'une matrice extracellulaire induite par le tgf beta a l'aide d'inhibiteurs de la renine
WO2003011328A1 (fr) * 2001-07-31 2003-02-13 Prince Henry's Institute Of Medical Research Activite enzymatique liee a la grossesse
AU2002355612B2 (en) * 2001-07-31 2007-11-01 Prince Henry's Institute Of Medical Research Pregnancy-related enzyme activity
EP2316474A1 (fr) 2001-07-31 2011-05-04 Prince Henry's Institute of Medical Research Antagonistes de l'enzyme PC 5/6 liée à la grossesse
WO2003060109A2 (fr) * 2002-01-15 2003-07-24 Bayer Healthcare Ag Regulation d'une subtilase humaine
WO2003060109A3 (fr) * 2002-01-15 2004-02-19 Bayer Healthcare Ag Regulation d'une subtilase humaine

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AU3617097A (en) 1998-02-20
WO1998004686A3 (fr) 1998-04-23
EP0920494A2 (fr) 1999-06-09

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