EP1305342A2 - Polypeptides du domaine d5 humain de production de kininees et ses utilisations - Google Patents

Polypeptides du domaine d5 humain de production de kininees et ses utilisations

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Publication number
EP1305342A2
EP1305342A2 EP01954904A EP01954904A EP1305342A2 EP 1305342 A2 EP1305342 A2 EP 1305342A2 EP 01954904 A EP01954904 A EP 01954904A EP 01954904 A EP01954904 A EP 01954904A EP 1305342 A2 EP1305342 A2 EP 1305342A2
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Prior art keywords
polypeptide
cell
cells
tumor
binding
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German (de)
English (en)
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Andrew P. Mazar
Jose C. Juarez
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Tactic Pharma LLC
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Attenuon LLC
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/81—Protease inhibitors
    • C07K14/8107—Endopeptidase (E.C. 3.4.21-99) inhibitors
    • C07K14/8139—Cysteine protease (E.C. 3.4.22) inhibitors, e.g. cystatin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A61P1/04—Drugs for disorders of the alimentary tract or the digestive system for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P11/00—Drugs for disorders of the respiratory system
    • 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
    • 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
    • A61P17/02—Drugs for dermatological disorders for treating wounds, ulcers, burns, scars, keloids, or the like
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A61P19/02—Drugs for skeletal disorders for joint disorders, e.g. arthritis, arthrosis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P27/00—Drugs for disorders of the senses
    • A61P27/02—Ophthalmic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P27/00—Drugs for disorders of the senses
    • A61P27/02—Ophthalmic agents
    • A61P27/06—Antiglaucoma agents or miotics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
    • A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A61P35/02—Antineoplastic agents specific for leukemia
    • 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
    • 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
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A61P9/10—Drugs for disorders of the cardiovascular system for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2319/00—Fusion polypeptide

Definitions

  • angiostatin the C-terminal hemopexin domain of matrix metalloprotease 2 (MMP-2) (Brooks PC et al, Cell 92:391-400, 1998)
  • MMP-2 matrix metalloprotease 2
  • the N- terminal 16 kD fragment of prolactin and a 29 kDa fragment of fibronectin O'Reilly MS et al, Cell 7P:315-328, 1994; Folkman, J., Sci. Amer. 250:150-154, 1996.
  • the two-chain molecule, HK a After kallikrein-mediated cleavage, the two-chain molecule, HK a , retains the trinodular structure, though the three globular regions rearrange in a pattern resembling vertices of a triangle, h this structure, the anionic surface binding and prekallikrein binding regions are more closely apposed.
  • HK a Compared to the antiproliferative effects, the anti-adhesive effects of HK a appear to be of less importance since EC adhesion was only modestly inhibited at HK a concentrations > 100 nM, whereas anti-proliferative effects were observed at concentrations as low as ⁇ 1 nM.
  • HKNKGKKNGKHNGWKT SEQ ID NO:6
  • HKNKGKKN SEQ ID NO:7
  • Another The polypeptide or peptide fragment as above has between about 4 and about 16 amino acids and includes one or more repeats of a sequence HKXK (SEQ ID NO: 8) where X is a neutral or aromatic amino acid.
  • the D5 fusion polypeptide may comprise any of the above polypeptides, peptide fragments, homologues or other functional derivatives, fused to a second polypeptide.
  • the binding partner molecule is a protein or peptide that increases the expression, stability or biologic or pharmacologic activity of the fusion polypeptide when compared to the D5 polypeptide, fragment homologue or derivative alone.
  • a preferred binding partner molecule is thioredoxin, calmodulin binding protein, maltose-binding protein or glutathione-S-transferase.
  • the linker in the above fusion is preferably cleavable by an enzyme that is present and active in the vicinity of, or in cells of, a tumor, such that the first fusion partner is released from the fusion polypeptide when the enzyme acts on the fusion polypeptide.
  • Preferred enzymes are a matrix metalloprotease, urokinase, a cathepsin, plasmin or thrombin, which act to release D5 in vivo (or in situ ) in the tumor milieu.
  • a preferred linker is a peptide having the sequence VPRGSD (SEQ ID NO:9) or DDKDWH ⁇ SEQ ID NO:10).
  • nucleic acid sequence that is linked in frame to the first nucleic acid sequence or to the linker nucleic acid sequence and that encodes a second polypeptide. Also included is an isolated nucleic acid molecule that hybridizes with any of the above nucleic acid molecules under stringent conditions.
  • the above polypeptide or a biologically active fragment, homologue or other functional derivative may be produced by recombinant expression of the above nucleic acid molecules.
  • an expression vector comprising a nucleic acid encoding any of the above polypeptides or functional derivatives, operatively linked to
  • Preferred expression vectors are plasmids or viral vectors.
  • the present invention is directed to an antibody that is specific for an epitope of a human kininogen D5 domain polypeptide, preferably a linear or conformational epitope of the polypeptide having SEQ ID NO:2.
  • the antibody may be specific for an epitope present in a peptide selected from the group consisting of
  • an angiogenic endothelial cell-targeting-targeting therapeutic composition comprises (a) an effective amount of the above polypeptide, fusion polypeptide, fragment, homologue or functional derivative of any of claims bound directly or indirectly to a therapeutically active moiety, such as a radionuclide; and (b) a therapeutically acceptable carrier.
  • the above diagnostic composition is used in a method for detecting the presence of angiogenic endothelial cells (i) in a tissue, (ii) in an organ or (iii) in a biological sample, which tissue, organ or sample is suspected of having angiogenically-activated endothelial cells.
  • the method comprises (a) contacting the tissue, organ or sample with the diagnostic composition; and (b) detecting the presence of the label associated with the tissue, organ or sample.
  • the contacting is in vivo. Both the contacting and the detecting may be in vivo.
  • the invention is also directed to an affinity ligand useful for binding to angiogenic endothelial cells or to a D5 domain binding site.
  • These ligands comprise the above polypeptide, fusion polypeptide, fragment, homologue or functional derivative immobilized to a solid support or carrier.
  • the ligand is used in a method for isolating a D5 domain binding molecule from a complex mixture, the method comprising: (a) contacting the mixture with the affinity ligand; (b) allowing any of the binding molecules to bind to the ligand; (c) removing unbound material from the ligand; and (d) eluting the bound D5 domain binding material.
  • a method for isolating or enriching cells expressing D5 domain binding sites from a cell mixture comprises (a) contacting the cell mixture with the affinity ligand of claim 57;
  • Figure 1 Primary sequence and genetic structure of high molecular weight kininogen. Numbers 1-626 are amino acid (aa) locations with leader sequence -18 to -1. Letters A-J are the locations of intron/exon junctions. Domain 1 (aa 1-113) is coded by exons 1, 2 and 3. Domain 2 (aa 114-234) is coded by exons 4, 5 and 6. Domain 3 (aa 235-357) is coded by exons 7, 8 and 9.
  • Domain 4 (aa 358-383) is coded by exon IO BK - Domain 5 (aa 384-502) is coded by the 5' portion of exon IO HK - Domain 6 (aa-503-626) is coded by the 3' portion of exon IO HK - Curved arrows indicate kallikrein cleavage sites. Boxed “O” and “N” are the locations of O- and N- linked carbohydrate chains, respectively.
  • XI is the putative factor XI binding sequence.
  • PK is the putative prekallikrein binding sequence (from DeLa Cadena 47 ).
  • NMR and molecular dynamic analysis where one examines regions that are defined by the active site of D5.
  • the capacity for motion in these structures is considered along with the impact of restraining the motion at particular sites on rigidity and biological activity of the molecule.
  • Conventional approaches of protein engineering are applied.
  • stability is increased by introducing one or more Cys residues into strategic positions , where the formation of disulfide bonds between two Cys residues increases stability.
  • Another approach is based on introduction of residues that form ⁇ helices at sites that do not impede the polypeptide' s biological activity, for example at the N- and C- termini. These helices have a charged face and a hydrophobic face, and because of the highly charged nature of the polypeptide, hydrophobic residues in the helices will enter into from helix-helix interactions that further stabilize the polypeptide.
  • a peptidomimetic agent may be an unnatural peptide or a non-peptide agent which recreates the stereospatial properties of the binding elements of D5 such that it has the binding activity or biological activity of D5. Similar to the linear peptides corresponding to D5, a peptidomimetic will have a binding face (which interacts with the D5BS) and a non-binding face. Again, similar to linear peptides of D5, the non-binding face of a peptidomimetic will contain functional groups which can be modified by various therapeutic moieties without modifying the binding face of the peptidomimetic. A preferred embodiment of a peptidomimetic would contain an aniline on the non-binding face of the molecule.
  • the CBP-D5 fusion is predicted to have a pl of 9.0 and can thus be purified using cation exchange such as SP (sulfopropyl)-Sepharose developed at pH 8.5.
  • SP sulfopropyl
  • Very few proteins have as basic of a pi as CBP- D5 and thus, only CBP-D5 would be positively charged at pH and capable of sticking to the column.
  • This protocol has been used for the one-step purification of the CBP-D5 fusion to homogeneity.
  • An active D5 fusion polypeptide can be expressed and purified in this manner regardless of fusion partner.
  • polypeptides as well as their variants and chemical derivatives, including peptidomimetics, must bind to endothelial cells or a fraction of these cells that contains the D5BS, preferably with an IC 50 ⁇ lO ⁇ M, more preferably ⁇ l ⁇ M. This activity is characterized in greater detail below.
  • Inducible non-fusion expression vectors include pTrc (Amann et al, (1988) Gene 69: 301-315) and pET lid (Studier et al, Gene Expression Technology: Methods in Enzymology 185, Academic Press, San Diego, Calif. (1990) 60-89). While target gene expression relies on host RNA polymerase transcription from the hybrid trp-lac fusion promoter in pTrc, expression of target genes inserted into pET 1 Id relies on transcription from the T7 gnlO-lacO fusion promoter mediated by coexpressed viral RNA polymerase (T7gnl). Th is viral polymerase is supplied by host strains BL21(DE3) or HMS174(DE3) from a resident ⁇ prophage harboring a T7gnl under the transcriptional control of the lacUV 5 promoter.
  • the enhancer is preferably placed upstream from the promoter with which it interacts to stimulate gene expression.
  • the endogenous viral LTR may be rendered enhancer-less and substituted with other desired enhancer sequences which confer tissue specificity or other desirable properties such as transcriptional efficiency on the D5-encoding DNA molecule of the present invention.
  • Retrovirus-mediated gene delivery generally requires target cell proliferation for gene transfer (Miller, D.G. et al, Mol. Cell. Biol. 10:4239 (1990). This condition is met by certain of the preferred target cells into which the present DNA molecules are to be introduced, i. e. , actively growing tumor cells.
  • Gene therapy of cystic fibrosis using transfection by plasmids using any of a number of methods and by retroviral vectors has been described by Collins et al, U.S. Patent 5,240,846.
  • the DNA molecules encoding the D5 sequences may be packaged into retrovirus vectors using packaging cell lines that produce replication-defective retroviruses, as is well-known in the art (see, for example, Cone, R.D. et al, Proc. Natl. Acad. Sci. USA 81 :6349-6353 (1984); Mann, R.F. et al, Cell 33:153-159 (1983); Miller, A.D. et al, Molec. Cell. Biol 5:431-437 (1985),; Sorge, J., et al, Molec. Cell. Biol. 4:1730-1737 (1984); Hock, R.A. et al, Nature 320:257 (1986); Miller, A.D. et al, Molec. Cell. Biol. 6:2895-2902 (1986). Newer packaging cell lines which are efficient an safe for gene transfer have also been described (Bank et al, U.S. 5,278,056.
  • adenovirus vectors for human gene therapy include the fact that recombination is rare, no human malignancies are known to be associated with such viruses, the adenovirus genome is double stranded DNA which can be manipulated to accept foreign genes of up to 7.5 kb in size, and live adenovirus is a safe human vaccine organisms.
  • Adeno- associated virus is also useful for human therapy (Samulski, R.J. et al, EMBO J. 10:3941 (1991) according to the present invention.
  • transduced D5 molecules may not require stable expression. Rather, transient expression of the polypeptide may be sufficient for transduced cells to perform their biological/pharmacological function.
  • EC growth factors such as VEGF
  • Cells are then be cultured for an additional 48 hours at 37° C, at which time the relative cell numbers in each well is determined using the Cell Titer ® Aq ueous cell proliferation assay (Promega). Briefly, 20 ⁇ l of a 19:1 (V/V) mixture of (3-(4,5-dimethylthiazol-2-yl)-5-(3- carboxymethylphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) and phenazine methosulfate (PMS) are be added to each well, and after an additional hour of incubation, A 9 o is measured.
  • the present inventor has quantitated ECs following incubation with HK a using both this method and manual cell counting, and observed a direct linear correlation.
  • this tumor produces metastases, preferentially in the lungs.
  • the primary tumor exerts anti-metastatic effects and must first be excised before study of the metastatic phase (see also U.S. 5,639,725).
  • Single-cell suspensions are prepared from solid tumors by treating minced tumor tissue with a solution of 0.3 %> trypsin. Cells are washed 3 times with PBS (pH 7.4) and suspended in PBS. Viability of the 3LL cells prepared in this way is generally about 95-99% (by trypan blue dye exclusion). Viable tumor cells (e.g., 3 x 10 4 - 5 x 10 6 ) suspended in 0.05 ml PBS are injected subcutaneously, either in the dorsal region or into one hind foot pad of C57BL/6 mice. Visible tumors appear after 3-4 days after dorsal sc injection of 10 cells. The day of tumor appearance and the diameters of established tumors are measured by caliper every two days.
  • the D5 polypeptides can also be made detectable by coupling them to a phosphorescent or a chemiluminescent compound. The presence of the chemiluminescent-tagged peptide is then determined by detecting the presence of luminescence that arises during the course of a chemical reaction. Examples of particularly useful chemiluminescers are luminol, isoluminol, theromatic acridinium ester, imidazole, acridinium salt and oxalate ester. Likewise, a bioluminescent compound may be used to label the peptides. Bioluminescence is a type of chemiluminescence found in biological systems in which a catalytic protein increases the efficiency of the chemiluminescent reaction.
  • auxiliary agents e.g., preservatives, stabilizers, wetting agents, buffers, or salts for influencing osmotic pressure and the like.
  • Preferred vehicles for non-sprayable topical preparations include ointment bases, e.g., polyethylene glycol-1000 (PEG- 1000); conventional creams such as HEB cream; gels; as well as petroleum jelly and the like.
  • ointment bases e.g., polyethylene glycol-1000 (PEG- 1000); conventional creams such as HEB cream; gels; as well as petroleum jelly and the like.
  • sprayable aerosol preparations wherein the active compound, preferably in combination with a solid or liquid inert carrier material, is packaged in a squeeze bottle or in admixture with a pressurized volatile, normally gaseous propellant.
  • the aerosol preparations can contain solvents, buffers, surfactants, perfumes, and/or antioxidants in addition to the compounds of the invention.
  • compositions of the invention may comprise, in addition to the D5 polypeptide, one or more additional anti-tumor agents, such as a mitotic inhibitor, e.g., vinblastine; alkylating agents, e.g., cyclophosphamide; a folate inhibitor, e.g., methotrexate, piritrexim or trimetrexate; an antimetabolite, e.g., 5-fluorouracil and cytosine arabinoside; an intercalating antibiotic, e.g., adriamycin and bleomycin; an enzyme or enzyme inhibitors, e.g., asparaginase, topoisomerase inhibitors such as etoposide; or a cytokine or "biological response modifier," e.g., an interferon or an interleukin.
  • a mitotic inhibitor e.g., vinblastine
  • alkylating agents e.g., cyclophosphamide
  • Preferred doses of the radionuclide conjugates are a function of the specific radioactivity to be delivered to the target site which varies with tumor type, tumor location and vascularization, kinetics and biodistribution of the D5 polypeptide carrier, energy of radioactive emission by the nuclide, etc.
  • Those skilled in the art of radiotherapy can readily adjust the dose of the D5 polypeptide in conjunction with the dose of the particular nuclide to effect the desired therapeutic benefit without undue experimentation.
  • an effective dose of I-D5 is between about 1 and 1000 ⁇ Ci per gram of tumor for an extracranial tumor.
  • the therapeutic dosage administered is an amount which is therapeutically effective, as is known to or readily ascertainable by those skilled in the art.
  • the dose is also dependent upon the age, health, and weight of the recipient, kind of concurrent treatment(s), if any, the frequency of treatment, and the nature of the effect desired, such as, for example, anti-inflammatory effects or anti-bacterial effect.
  • Diphtheria toxin and Pseudomonas exotoxin A are also single chain proteins, and their binding and toxicity functions reside in separate domains of the same protein chain with full toxin activity requiring proteolytic cleavage between the two domains.
  • Pseudomonas exotoxin A has the same catalytic activity as diphtheria toxin.
  • Ricin has been used therapeutically by binding its toxic ⁇ -chain, to targeting molecules such as antibodies to enable site-specific delivery of the toxic effect.
  • Bacterial toxins have also been used as anti-tumor conjugates.
  • the present pharmaceutical compositions are intended for the treatment of any of diseases or conditions that involve ocular neovascularization, chief among them, prohferative diabetic retinopathy, neovascular age-related macular degeneration,, retinopathy of prematurity, sickle cell retinopathy and retinal vein occlusion.
  • Fab, F(ab') 2 , Fv and scFv fragments or forms of the antibodies useful in the present invention may be used for the detection and quantitation of the D5 polypeptides in the same manner as an intact antibody.
  • Conventional fragments are typically produced by proteolytic cleavage, using enzymes such as papain (for Fab fragments) or pepsin (for F(ab') 2 fragments).
  • Fv fragments are described in (Hochman, J. et al. (1973) Biochemistry 12:1130- 1135; Sharon, J, et ⁇ /.(1976) Biochemistry 15:1591-1594). ).
  • scFv polypeptides include the hypervariable regions from the Ig of interest and recreate the antigen binding site of the native Ig while being a fraction of the size of the intact Ig (Skerra, A. et al. (1988) Science, 240: 1038- 1041; Pluckthun, A. et al (1989) Methods Enzymol 178: 497-515; Winter, G. et al. (1991) Nature, 349: 293-299); Bird et al, (1988) Science 242:423; Huston et al (1988) Proc. Natl. Acad. Sci. USA 85:5879; U.S. Patents No. 4,704,692, 4,853,871, 4,94,6778, 5,260,203, 5,455,030.
  • Chimeric antibodies are Ig molecules wherein different parts of the molecule are derived from different animal species.
  • An example is an Ig having a variable region derived from a murine mAb and a human Ig constant region.
  • Chimeric antibodies and methods for their production are known in the art ( Cabilly et al, Proc. Natl. Acad. Sci. USA 57:3273-3277 (1984); Morrison et al, Proc. Natl. Acad. Sci.
  • the antibodies, or fragments of antibodies, useful in the present invention may be used to quantitatively or qualitatively detect the presence of aD5 polypeptide. For example, it may be desired to monitor the level of a D5 polypeptide in the circulation or in the tissues of a subject receiving therapeutic doses of the polypeptide. Thus, the antibodies (of fragments thereof) useful in the present invention may be employed histologically to detect the presence of D5
  • Antibodies to D5 epitopes are also used to treat tumors. They may inhibit interactions between endothelial cells and ECM, so that their binding to D5 results in a similar outcome to treatment with D5 peptides and fusion proteins.
  • the expression vector was transformed into BL21(DE3) Gold cells (Stratagene) and subclones were grown and induced with 1 mM IPTG at 37°C for 4 hours. SDS-PAGE analysis revealed that the majority of expressed CBP-D5 was in inclusion bodies.
  • a 500 ml preparation of cells was grown, harvested and the pellet washed once with 50 mM Tris/ 1 mM EDTA, pH 8.0 (TE buffer).
  • the cell suspension was lysed with 0.25 mg/ml lysozyme in TE buffer.
  • the lysate was homogenized with 4% Tergitol and centrifuged at 10,000 x g for 45 minutes.
  • the inclusion body pellet was washed several times with TE buffer.

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Abstract

Cette invention a trait à des peptides issus du domaine D5 humain de production de kinine ainsi qu'aux peptides hybrides de celui-ci, lesquels ont une action inhibitrice de l'angiogenèse. On utilise ces peptides pour le diagnostic et le traitement d'états pathologiques associés à une migration et à une prolifération des cellules endothéliales, notamment pour le traitement du cancer. L'invention concerne également des molécules d'acide nucléique codant ces peptides, des anticorps contre ces peptides, des cellules exprimant ces peptides ainsi que des procédés d'isolation de ceux-ci.
EP01954904A 2000-07-24 2001-07-24 Polypeptides du domaine d5 humain de production de kininees et ses utilisations Withdrawn EP1305342A2 (fr)

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AU2012201537B2 (en) * 2002-07-15 2014-06-05 Board Of Regents, The University Of Texas System Peptides binding to phosphatidylethanolamine and their use in treating viral infections and cancer
US7378386B2 (en) 2002-07-15 2008-05-27 Board Of Regents, The University Of Texas System Anti-viral treatment methods using phosphatidylethanolamine-binding peptide derivatives
US6987270B2 (en) 2003-05-07 2006-01-17 General Electric Company Method to account for event losses due to positron range in positron emission tomography and assay of positron-emitting isotopes
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WO2014194259A1 (fr) * 2013-05-30 2014-12-04 Arizona Board Of Regents, A Body Corporate Of The State Of Arizona, Acting For And On Behalf Of Arizona State University Procédés et compositions pour traiter des maladies cérébrales
WO2016004281A1 (fr) * 2014-07-03 2016-01-07 Chunlei Liu Polypeptides comprenant un canal ionique thermosensible lié au domaine 5 d'un kininogène, des acides nucléiques, et leurs utilisations pour la modulation cellulaire et des traitements

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CA2345382A1 (fr) * 1998-10-16 2000-04-27 Immunex Corporation Inhibiteurs de l'activation et du recrutement plaquettaires
JP2002529474A (ja) * 1998-11-10 2002-09-10 テンプル・ユニバーシティ−オブ・ザ・コモンウェルス・システム・オブ・ハイアー・エデュケイション 高分子量キニノゲン及びそのペプチド類似体による血管形成の抑制

Non-Patent Citations (5)

* Cited by examiner, † Cited by third party
Title
FUKUOKA Y ET AL., IMMUNOLOGY LETTERS, vol. 38, 1993, pages 153 - 158 *
HITOMI Y ET AL., VIRAL IMMUNOLOGY, vol. 8, 1995, pages 109 - 119 *
KAPUST RB ET AL., PROTEIN SCIENCE, vol. 8, 1999, pages 1668 - 1674 *
SHEFFIELD P ET AL., PROTEIN EXPRESSION AND PURIFICATION, vol. 15, 1999, pages 34 - 39 *
SPIEGEL H ET AL., PLANT SCIENCE, vol. 149, 1999, pages 63 - 71 *

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WO2002014369A2 (fr) 2002-02-21
WO2002014369A9 (fr) 2003-04-03

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