Nothing Special   »   [go: up one dir, main page]

CN1735433A - Mammalian EPO mimetic CH1 deleted mimetibodies, compositions, methods and uses - Google Patents

Mammalian EPO mimetic CH1 deleted mimetibodies, compositions, methods and uses Download PDF

Info

Publication number
CN1735433A
CN1735433A CNA038201100A CN03820110A CN1735433A CN 1735433 A CN1735433 A CN 1735433A CN A038201100 A CNA038201100 A CN A038201100A CN 03820110 A CN03820110 A CN 03820110A CN 1735433 A CN1735433 A CN 1735433A
Authority
CN
China
Prior art keywords
epo mimetic
polypeptide
deleted mimetibodies
mimetibodies
deleted
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.)
Pending
Application number
CNA038201100A
Other languages
Chinese (zh)
Inventor
G·A·希纳
D·M·奈特
J·格拉耶
B·J·斯卡伦
T·C·内斯珀尔
K·A·库托洛斯基
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Janssen Biotech Inc
Original Assignee
Centocor Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Centocor Inc filed Critical Centocor Inc
Publication of CN1735433A publication Critical patent/CN1735433A/en
Pending legal-status Critical Current

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • 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
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/505—Medicinal preparations containing antigens or antibodies comprising antibodies
    • 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
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/50—Immunoglobulins specific features characterized by immunoglobulin fragments
    • C07K2317/52—Constant or Fc region; Isotype
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2318/00—Antibody mimetics or scaffolds
    • C07K2318/10—Immunoglobulin or domain(s) thereof as scaffolds for inserted non-Ig peptide sequences, e.g. for vaccination purposes
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2319/00—Fusion polypeptide
    • C07K2319/30—Non-immunoglobulin-derived peptide or protein having an immunoglobulin constant or Fc region, or a fragment thereof, attached thereto

Landscapes

  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Engineering & Computer Science (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Biophysics (AREA)
  • Molecular Biology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Biochemistry (AREA)
  • Immunology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Genetics & Genomics (AREA)
  • Animal Behavior & Ethology (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Peptides Or Proteins (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)

Abstract

The present invention relates to at least one novel EPO human CH1-deleted mimetibody or specified portion or variant, including isolated nucleic acids that encode at least one CH1-deleted mimetibody or specified portion or variant, CH1-deleted mimetibody or specified portion or variants, vectors, host cells, transgenic animals or plants, and methods of making and using thereof, including therapeutic compositions, methods and devices.

Description

Mammalian EPO mimetic CH 1 deleted mimetibodies, compositions, method and purposes
Background of invention
Invention field
The present invention relates to that biologically activated protein, fragment or part had specific mammalian EPO mimetic CH 1 deleted mimetibodies, specific part or variant, EPO mimetic CH 1 deleted mimetibodies code nucleic acid and complementary nucleic acid, host cell, and preparation and using method, comprise the treatment prescription, use and install.
Correlation technique
Recombiant protein is the emerging therapeutic agent of a class.This recombinant therapy is making progress aspect protein formulations and the chemical modification.This modification can strengthen the treatment effectiveness of therapeutic protein potentially, for example by prolong half-life (for example, being exposed to proteolytic enzyme by hindering it), strengthens biological activity or reduces harmful side effect.A kind of such modification is to utilize to be merged to receptor protein, as the immunoglobulin fragment of enteracept.People also once utilized the Fc district to make up therapeutic protein, attempted the half-life that provides long or integrated such as Fc receptors bind, A protein binding and the bonded function of complement.
A special and important effect of mammal hemopoietic system is to generate erythrocyte, and this erythrocyte can be with oxygen transport to the intravital a plurality of tissues of animal.Generate erythrocytic process (" erythropoiesis ") and occur in the in life whole of animal incessantly, to remedy ruined erythrocyte.Typical erythrocyte has the short relatively life-span, is generally 100-120 days.Erythropoiesis is a physiological mechanisms that quilt is accurately controlled, thereby can generate the erythrocyte of capacity, realizing appropriate tissue oxygenation, but can not hinder circulation by as many as.
At present, known erythropoiesis mainly is subjected to a kind of acidoglycoprotein, i.e. the control of polypeptide erythropoietin (EPO).The generation of erythropoietin is to be arranged in the result that the single copy gene of mammal chromosome is expressed.The aminoacid sequence of recombinant human EPO (" rHuEPO ") basically with the consensus amino acid sequence that obtains from the EPO in Urina Hominis source.But the glycosylation of rHuEPO is different with the glycosylation of urine EPO and human serum EPO.
In healthy mammalian body, when organizing by the abundant oxygenate of circulation erythrocyte of on-hand quantity, the concentration of EPO in blood plasma is very low.The existence of EPO has stimulated new erythrocytic generation, to substitute those erythrocyte that lose in ageing process.In addition, being created under the hypoxia condition of EPO promoted, wherein, although blood is enough to be full of tissue, the oxygen supply of when injected organism tissue still is lower than the normal physiological level.Hypoxia may hemorrhage, radiation induced erythrocyte destruction, various anemia, high height above sea level or long-term unconscious causing.In contrast, when the erythrocyte number in the circulation surpassed normal structure oxygenate requirement, the generation of EPO had just reduced.
But, some morbid state relates to the abnormal erythrocyte generation.Many countries have utilized recombinant human EPO (rHuEPO) in treatment.In the U.S., U.S. food and FAD (FDA) approved the application of rHuEPO aspect the treatment anemia relevant with latter stage nephropathy.Typically suffer from serious anemia for treating the patient that this disease accepts hemodialysis, this be erythrocyte fragmentation of causing of dialysis treatment and ripe before death cause.EPO also is used in the treatment of other type anemia.For example, anemia, the anemia relevant, the anemia of being correlated with, the relevant anemia of AIDS and the anemia relevant that can adopt EPO therapeutical chemistry therapy to bring out with precocity with multiple congenital imbalance with myelodysplasia.In addition, EPO also can play a role in other field, recovers the normal plasma cell specific volume as promoting quickly in bone marrow transplantation patient, the patient who prepares autotransfusion and the ferrum overload imbalance patient body.
Erythropoietin (EPO) is by 165 aminoacid and 4 glycoprotein hormoneses that sugar chain is formed, and combines by the special receptor with erythrocyte precursor surface, erythropoiesis brought into play the effect of main regulatory factors.This combination shows that they will breed and be divided into mature erythrocyte.Erythropoietin receptor is that erythropoietin is had high-affinity, and contains 484 amino acid whose glycoproteins.For this erythropoietin receptor, the equal dimerization that part brings out may be one of activatory critical events of control.
Erythropoietin has the short relatively half-life.The clearance rate of the erythropoietin that intravenous is used meets first order kinetics, and it is about 3-4 hour at the intravital circulating half-life of CRF patient.In the therapeutic dose scope, but the detection level of blood plasma erythropoietin is maintained to few 24 hours.Behind the subcutaneous administration erythropoietin, reach the peak value serum levels in 5-24 hour, and after slowly reduce.Cmax after erythropoietin is used and t 1/2Be respectively 1.80 ± 0.7U/mL and 19.0 ± 5.9 hours.
The initial dose scope of erythropoietin is 50-150U/kg, and is inferior on every Wendesdays.The dosage of erythropoietin must be used at Different Individual, so that hematocrit maintains in the above-mentioned suggestion target zone.For surgical patients, the recommended dose of erythropoietin is subcutaneous injection 10 days, the operation same day and an operation back subcutaneous injection 4 days 300U/kg/ days before operation, or optionally weekly subcutaneous injection 600U/kg (perform the operation preceding 21,14 and 7 days), add and used this dosage the 4th time the same day of performing the operation.
Several studies group is by filtering out the peptide that erythropoietin receptor is had affinity from any phage display peptide library, thereby identifies the little peptide mimics of erythropoietin.Its sequence and erythropoietin do not have homology.In functional trial, there are some kinds of above-mentioned peptides to show activity, but only are active 1/100,000 of recombinant erythropoietin.Although people have carried out some trials by the covalent dimer or the polymer of preparation peptide mimics for improving tiring of above-mentioned peptide, the activity of above-claimed cpd is still low by 1 than erythropoietin by mole, 000-10,000 times.
The peptide sequence in erythropoietin source also by claim for being synergic.Also there is report to show that the activity of the dimerization sequence that comprises a plurality of or all natural erythropoietin sequence has been enhanced.These chemical compounds have very low or do not have the oral bioavailability rate, and its activity also makes it in the present commercial feasibility that do not have.
Therefore, the improvement of EPO therapeutic protein need be provided and/or modify version, overcoming an above-mentioned one or more difficult problem, and other difficult problem known in the art.
Summary of the invention
The method that place like this is described and/or put into practice, and in conjunction with method known in the art, the invention provides isolating human EPO mimetic CH 1 deleted mimetibodies, the immunoglobulin, pyrolysis product and other specific part and the variant thereof that comprise modification, EPO mimetic CH 1 deleted mimetibodies compositions, coding or complementary nucleic acid, carrier, host cell, compositions, prescription, device, transgenic animal, transgenic plant also are provided, and methods for making and using same.
The present invention also provides place like this to describe and/or at least a separation EPO mimetic CH 1 deleted mimetibodies known in the art or specific part or variant.This EPO mimetic CH 1 deleted mimetibodies can randomly comprise at least one CH3 district, this CH3 district directly links to each other with at least one CH2 district, this CH2 district directly links to each other with at least one hinge region or its fragment, this hinge region or its fragment directly link to each other with at least one part of V district, this part of V district directly links to each other with any joint sequence, this joint sequence directly links to each other with at least one therapeutic peptide, and this therapeutic peptide then randomly further directly links to each other with at least one part of at least one variable antibody sequence.In a kind of preferred embodiment of a pair of CH3-CH2-hinge-part J sequence-joint-therapeutic peptide with optional N-terminal antibody sequence, be randomly by associating or covalent bond, for example, but be not limited only to the Cys-Cys disulfide bond connect into right.In a kind of embodiment, the EPO mimetic CH 1 deleted mimetibodies comprises molecular formula (I): (V1 (n)-Pep (n)-Flex (n)-V2 (n)-pHinge (n)-CH2 (n)-CH3 (n)) (m), wherein V1 is at least one part of immune globulin variable region N-terminal, Pep is the EPO mimic peptide of at least one biologically active, Flex is by making analogue body have optional orientation and binding characteristic, thereby provide the polypeptide of structural flexibility, V2 is at least one part of immune globulin variable region C-terminal, pHtinge is at least one part of immunoglobulin variable hinge region, CH2 is at least one part of immunoglobulin CH2 constant region, CH3 is at least one part of immunoglobulin CH 3 constant regions, n and m can be the integer among the 1-10, simulate dissimilar immunoglobulin molecules, for example, but be not limited only to IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, IgM, IgD, IgE etc., or its combination.
Therefore, EPO mimetic CH 1 deleted mimetibodies of the present invention is under the situation that keeps inherent character and function, simulated at least one part of antibody or immunoglobulin structure or function, also provide in a kind of therapeutic peptide and external, the body thereof simultaneously or original position institute is intrinsic or the characteristic or the activity of acquisition.Adopt method described herein, and, can change a plurality of parts of antibody of the present invention and the therapeutic peptide part of at least a EPO mimetic CH 1 deleted mimetibodies in conjunction with methods known in the art.
One aspect of the present invention provides the isolated nucleic acid molecule that comprises specific polynucleotide, with the complementary isolated nucleic acid molecule of these specific polynucleotide, with these specific polynucleotide have remarkable homogeneity isolated nucleic acid molecule or with the isolated nucleic acid molecule of its hybridization, the wherein specific analogue body of this specific polynucleotide encoding or its specific part or variant, and comprise its at least one particular sequence, zone, part or variant.The present invention further provides the recombinant vector that comprises at least a above-mentioned separation EPO mimetic CH 1 deleted mimetibodies nucleic acid molecules, the host cell that contains above-mentioned nucleic acid and/or recombinant vector, and the preparation and/or the application process of above-mentioned EPO mimetic CH 1 deleted mimetibodies nucleic acid, carrier and/or host cell.
At least a EPO mimetic CH 1 deleted mimetibodies of the present invention or specific part or variant have been simulated combining of this analogue body Pep part and at least one part, perhaps have at least a biologic activity of at least a protein, subunit, fragment, part or its combination in any.
The present invention also provides at least a separation EPO mimetic CH 1 deleted mimetibodies described herein and/or known in the art or specific part or variant, wherein EPO mimetic CH 1 deleted mimetibodies or specific part or variant have at least a activity, for example, but to be not limited only to molecular formula be the peptide of the corresponding at least a biologically active of the Pep part of I or the known organism activity of polypeptide.Therefore, can filter out according to known method and to have corresponding active EPO mimetic CH 1 deleted mimetibodies, this corresponding activity is such as active at a kind of protein or its segmental at least a neutralization.
The present invention also provides at least a compositions, and said composition has comprised at least a separation EPO mimetic CH 1 deleted mimetibodies or the specific part or the variant of (a) coding nucleic acid described herein and/or EPO mimetic CH 1 deleted mimetibodies; (b) a kind of suitable carriers or diluent.According to known method, this carrier or diluent can randomly can be accepted carrier or diluent for pharmacopedics.Said composition can randomly further comprise at least a other chemical compound, protein or compositions.
The present invention also provides at least a method that is used at least a EPO mimetic CH 1 deleted mimetibodies of host cell inner expression or specific part or variant, comprise and cultivate host cell described herein and/or known in the art under given conditions, under this condition, at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant are can detect and/or callable amount is expressed.
The present invention further provides at least a EPO mimetic CH 1 deleted mimetibodies in ad hoc approach or the compositions, specific part or variant, when using this EPO mimetic CH 1 deleted mimetibodies with the treatment effective dose, when specific part or variant, can be according to multiple different situations, for example, but be not limited only to before relevant disease known in the art or treatment condition, afterwards or during needs be used for regulating, treatment or alleviate following at least a symptom, comprise the imbalance of bone and joint, cardiovascular disorder, tooth or mouthful imbalance, cutaneous disorder, ear, nose or throat imbalance, endocrine or Metabolic disorder, gastrointestinal disorder, gynecological's imbalance, liver or gallbladder imbalance, obstetrics' imbalance, the blood imbalance, immunity or allergia imbalance, infectious disease, the imbalance of flesh skeleton, the tumor imbalance, nervous disorder, nutritional disorder, the ophthalmology imbalance, the department of pediatrics imbalance, poison and lack of proper care, the psychiatry imbalance, the kidney imbalance, lung imbalance or other any known imbalance (are seen for example by the complete TheMerck Manual that is incorporated herein by reference, 17 ThEd., Merck Research Laboratories, MerckandCo., Whitehouse Station, NJ (1999)).
The present invention further provides at least a EPO mimetic CH 1 deleted mimetibodies in ad hoc approach or the compositions, specific part or variant, when using this EPO mimetic CH 1 deleted mimetibodies with the treatment effective dose, when specific part or variant, can be used for regulating, treatment or alleviation cell, tissue, organ, at least a immunity among animal or the patient, cardiovascular, infectiousness, pernicious and/or sacred disease symptom, and/or according to multiple different situations, for example, but be not limited only to before relevant disease known in the art and/or described herein or treatment condition, afterwards or during needs and use.
The present invention also provides at least a treatment or the EPO mimetic CH 1 deleted mimetibodies at least a of the present invention of prevention effective dose or compositions, device and/or the method for specific part or variant of passing that be used to send.
The present invention further provides at least a anti-idiotype antibody at least a EPO mimetic CH 1 deleted mimetibodies of the present invention.This anti-idiotype antibody comprises arbitrary protein matter or the peptide that contains specific molecular, this specific molecular comprises at least one part of immunoglobulin molecules, for example, but be not limited only at least one complementary determining region (CDR) of heavy or light chain or its ligand binding moiety, heavy chain or variable region of light chain, heavy chain or constant region of light chain, framework region, or its arbitrary portion, and this immunoglobulin molecules is competitively in conjunction with the EPO receptor binding domain of at least a EPO mimetic CH 1 deleted mimetibodies of the present invention.Idiotype antibody of the present invention can comprise or derive from any mammal, for example, but is not limited only to the idiotype antibodies of the mankind, mice, rabbit, rat, rodent, primate etc.
The present invention also provide at least a isolated nucleic acid molecule that comprises specific polynucleotide, with the complementary isolated nucleic acid molecule of these specific polynucleotide, or with the isolated nucleic acid molecule of this specific multi-nucleotide hybrid, at least a EPO mimetic CH 1 deleted mimetibodies of this specific polynucleotide encoding anti-idiotype antibody wherein, and comprise its at least one particular sequence, zone, part or variant.The present invention further provides the recombinant vector that comprises above-mentioned EPO mimetic CH 1 deleted mimetibodies anti-idiotype antibody coding nucleic acid molecule, the host cell that contains above-mentioned nucleic acid and/or recombinant vector, and the preparation and/or the application process of above-mentioned anti-idiotype antibody nucleic acid, carrier and/or host cell.
The present invention also provides at least a method that is used at least a EPO mimetic CH 1 deleted mimetibodies of host cell inner expression or EPO mimetic CH 1 deleted mimetibodies anti-idiotype antibody, this method comprises cultivates host cell described herein under given conditions, wherein under this condition, at least a EPO mimetic CH 1 deleted mimetibodies or anti-idiotype antibody are can detect and/or callable amount is expressed.
The present invention also provides at least a compositions, and said composition comprises (a) separation EPO described herein mimetic CH 1 deleted mimetibodies code nucleic acid and/or EPO mimetic CH 1 deleted mimetibodies; (b) a kind of suitable carriers or diluent.According to known carrier or diluent, this carrier or diluent may optionally be pharmacopedics can accept carrier or diluent.Said composition can randomly further comprise at least a other chemical compound, protein or compositions.
At least a EPO mimetic CH 1 deleted mimetibodies method or compositions have been the present invention further provides, but the administering therapeutic effective dose with regulate or treatment cell, tissue, organ, animal or patient at least a protein conditions associated, and/or be applied to known in the art and/or described herein conditions associated before, afterwards or during.
The present invention also provides at least a compositions, device and/or the method that is used to send the EPO mimetic CH 1 deleted mimetibodies at least a of the present invention of passing treatment or prevention effective dose.
At least a EPO mimetic CH 1 deleted mimetibodies method or compositions have been the present invention further provides, at least a EPO that can be used among diagnosis cell, tissue, organ, animal or the patient is conditions associated, and/or be applied to known in the art and/or described herein conditions associated before, afterwards or during.
The present invention also provides at least a compositions, device and/or the method for at least a EPO mimetic CH 1 deleted mimetibodies of the present invention to diagnose of passing that be used to send.
On the one hand, the invention provides at least a isolating mammalian EPO mimetic CH 1 deleted mimetibodies, this analogue body comprises at least one Pep (n) district, this zone comprises at least one part of at least one sequence among the SEQ ID NO:1-42 as shown in table 1 below, perhaps randomly have described herein or known in the artly one or morely substitute, disappearance or insertion portion.On the other hand, the invention provides at least a isolating mammalian EPO mimetic CH 1 deleted mimetibodies, wherein this EPO mimetic CH 1 deleted mimetibodies can combine with at least one specific epitope specificity, or randomly have described herein or known in the artly one or morely substitute, disappearance or insertion portion, wherein above-mentioned epi-position comprises individual at least a part of 1-3 or land at least, and this part can combine with at least one part of at least one sequence among the NO:1-42 of SEQ ID shown in the following table 1.
Above-mentioned at least a EPO mimetic CH 1 deleted mimetibodies can randomly further comprise at least a conjugated protein, and this is conjugated protein to have and be selected from least 10 -9M, at least 10 -10M, at least 10 -11M or at least 10 -12The affinity of one of M; At least a activity of at least a protein or its part of having neutralized basically.The present invention also provides the isolating nucleic acid of at least a separation mammalian EPO mimetic CH 1 deleted mimetibodies of encoding; Comprise the isolating nucleic acid carrier of this isolating nucleic acid, and/or comprise the protokaryon or the eukaryotic host cell of this isolating nucleic acid.This host cell can randomly be at least and be selected from COS-1, COS-7, HEK293, BHK21, CHO, BSC-1, Hep G2,653, SP2/0,293, HeLa, myeloma or lymphoma cell, or the cell of one of the deriving arbitrarily of above-mentioned cell, immortalization or transformant.The present invention also provides the method that is used to generate at least a EPO mimetic CH 1 deleted mimetibodies, that this method is included in is external, translation EPO mimetic CH 1 deleted mimetibodies code nucleic acid in the body or under the original position condition, makes this EPO mimetic CH 1 deleted mimetibodies can detect or callable amount is expressed.
The present invention also provides and has comprised the compositions that at least a separation mammalian EPO mimetic CH 1 deleted mimetibodies and at least a pharmacopedics can be accepted carrier or diluent.Said composition can randomly further comprise at least a specific compound or the protein of effective dose, and this specific compound or protein are selected from least a detectable label or reporter molecule, anti-infectives, cardiovascular (CV) system medicine, central nervous system (CNS) medicine, autonomic nervous system (ANS) medicine, respiratory drugs, gastrointestinal (GI) tract drug, hormonal medicaments, the medicine that is used for body fluid or electrolyte balance, blood substance, antitumor drug, immunoregulation medicament, eye, ear or nose medication, local application, nutritional drugs, the TNF antagonistic, antirheumatic, muscle relaxant, anesthetis, on-steroidal anti-inflammatory medicaments (NSAID), analgesic, anesthetics, tranquilizer, local anesthetic, neuromuscular blocking agents, antibacterial, antipsoriatic, corticosteroid, anabolic steroid, erythropoietin, immunity, immunoglobulin, immunosuppressant, growth hormone, hormone replaces medicine, radiopharmaceutical, antidepressants, psychosis, analeptic, the asthma medicine, the β synergist, suck steroid, epinephrine or analog, cytokine or cytokine antagonist.
The present invention further provides can with bonded anti-idiotype antibody of at least a EPO mimetic CH 1 deleted mimetibodies of the present invention specificity or fragment.
The present invention also provides the method for the disease symptoms that is used for diagnosing or treat cell, tissue, organ or animal, comprises
(a) make cell, tissue, organ or animal contact or it is used the compositions of at least a separation mammalian EPO mimetic CH 1 deleted mimetibodies of the present invention who comprises effective dose.This method can randomly further comprise the effective dose that adopts per kilogram cell, tissue, organ or animal weight to use 0.001-50mg.This method can randomly further comprise by at least a parenteral that is selected from, subcutaneous, intramuscular, intravenous, intraarticular, in the bronchus, in the abdomen, in the capsule, in the cartilage, intracavity, in the body cavity, in the cerebellum, Intraventricular, colonic, in the cervix uteri, gastric, in the liver, in the cardiac muscle, in the bone, in the pelvis, in the pericardium, intraperitoneal, in the pleura, in the prostate, in the lung, internal rectum, in the kidney, in the retina, in the spinal column, in the synovial membrane, intrathoracic, intrauterine, intravesical, medicine group, vagina, rectum, cheek, the Sublingual, the mode of intranasal or percutaneous contacts or uses.This method can randomly further be included in before (a) contact or use, simultaneously or use at least a compositions afterwards, at least a specialization that said composition contains effective dose contains thing or protein, and this chemical compound or protein are selected from least a detectable label or reporter molecule, anti-infectives, cardiovascular (CV) system medicine, central nervous system (CNS) medicine, autonomic nervous system (ANS) medicine, respiratory drugs, gastrointestinal (GI) tract drug, hormonal medicaments, the medicine that is used for body fluid or electrolyte balance, blood substance, antitumor, immunoregulation medicament, eye, ear or nose medication, local application, nutritional drugs, the TNF antagonistic, antirheumatic, muscle relaxant, anesthetis, on-steroidal anti-inflammatory medicaments (NSAID), analgesic, anesthetics, tranquilizer, local anesthetic, neuromuscular blocking agents, antibacterial, antipsoriatic, corticosteroid, anabolic steroid, erythropoietin, immunity, immunoglobulin, immunosuppressant, growth hormone, hormone replaces medicine, radiopharmaceutical, antidepressants, psychosis, analeptic, the asthma medicine, the β synergist, suck steroid, epinephrine or analog, cytokine or cytokine antagonist.
The present invention also provides the medical apparatus that comprises at least a isolating mammalian EPO mimetic CH 1 deleted mimetibodies of the present invention, and wherein this device is fit to by at least a parenteral that is selected from, subcutaneous, intramuscular, intravenous, intraarticular, in the bronchus, in the abdomen, in the capsule, in the cartilage, intracavity, in the body cavity, in the cerebellum, Intraventricular, colonic, in the cervix uteri, gastric, in the liver, in the cardiac muscle, in the bone, in the pelvis, in the pericardium, intraperitoneal, in the pleura, in the prostate, in the lung, internal rectum, in the kidney, in the retina, in the spinal column, in the synovial membrane, intrathoracic, intrauterine, intravesical, medicine group, vagina, rectum, cheek, the Sublingual, the mode of intranasal or percutaneous contacts or uses at least a EPO mimetic CH 1 deleted mimetibodies.
The present invention also provides the goods that are used for human medicine or diagnostic uses, comprises at least a container that separates mammalian EPO mimetic CH 1 deleted mimetibodies of packaging material and the present invention who contains solution or lyophilized form.These goods can randomly comprise container as parenteral, subcutaneous, intramuscular, intravenous, intraarticular, in the bronchus, in the abdomen, in the capsule, in the cartilage, intracavity, in the body cavity, in the cerebellum, Intraventricular, colonic, in the cervix uteri, gastric, in the liver, in the cardiac muscle, in the bone, in the pelvis, in the pericardium, intraperitoneal, in the pleura, in the prostate, in the lung, internal rectum, in the kidney, in the retina, in the spinal column, in the synovial membrane, intrathoracic, intrauterine, intravesical, medicine group, vagina, rectum, cheek, the Sublingual, an ingredient of intranasal or percutaneous delivery device or system.
The present invention also provides the method that is used to prepare at least a separation mammalian EPO mimetic CH 1 deleted mimetibodies of the present invention, and this method comprises host cell or transgenic animal or transgenic plant or the plant cell that the EPO mimetic CH 1 deleted mimetibodies that can express recyclable amount is provided.The present invention further provides at least a EPO mimetic CH 1 deleted mimetibodies by method for preparing.
The present invention also provides at least a method that is used for expressing at host cell at least a EPO mimetic CH 1 deleted mimetibodies or anti-idiotype antibody, this method comprises cultivates host cell described herein under given conditions, under this condition, at least a EPO mimetic CH 1 deleted mimetibodies is can detect and/or callable amount is expressed.
The present invention further provides any invention described herein.
Detailed Description Of The Invention
The invention provides isolating, reorganization and/or synthetic analogue body or specific part or variant, and the compositions and the coding nucleic acid molecule that comprise at least a specific polynucleotide, at least a EPO mimetic CH 1 deleted mimetibodies of this specific polynucleotide encoding.The invention described above analogue body or specific part or variant have comprised specific EPO mimetic CH 1 deleted mimetibodies sequence, zone, fragment and specific variants thereof, and the present invention also provides the methods for making and using same of above-mentioned nucleic acid and analogue body or specific part or variant, comprises therapeutic composition, method and apparatus.
The present invention also provides at least a isolating EPO mimetic CH 1 deleted mimetibodies described herein and/or known in the art or specific part or variant.This EPO mimetic CH 1 deleted mimetibodies can randomly comprise at least one CH3 district, this CH3 district directly links to each other with at least one CH2 district, this CH2 district directly links to each other with at least one hinge region or its fragment, this hinge region or its fragment directly link to each other with at least one part of V district, this part of V district directly links to each other with any joint sequence, this any joint sequence directly links to each other with at least one therapeutic peptide, and this therapeutic peptide then randomly further directly links to each other with at least one part of at least one variable antibody sequence.
In a kind of preferred embodiment, the EPO mimetic CH 1 deleted mimetibodies comprises molecular formula (I): (V1 (n)-Pep (n)-Flex (n)-V2 (n)-pHinge (n)-CH2 (n)-CH3 (n)) (m) wherein V1 is at least one part of immune globulin variable region N-terminal, Pep is the peptide of at least one biologically active, Flex is by making analogue body have optional orientation and binding characteristic, thereby provide the polypeptide of structural flexibility, V2 is at least one part of immune globulin variable region C-terminal, pHinge is at least one part of immunoglobulin variable hinge region, CH2 is at least one part of immunoglobulin CH2 constant region, CH3 is at least one part of immunoglobulin CH3 constant region, n and m can be the integer among the 1-10, simulate dissimilar immunoglobulin molecules, for example, but be not limited only to IgG1, IgG2, IgG3, IgG4, IgA, IgM, IgD, IgE etc., or its combination.Monomer during m=1 can for example, link to each other with other monomer but be not limited only to the Cys-Cys disulfide bond by associating or covalent bond.Therefore, EPO mimetic CH 1 deleted mimetibodies of the present invention has been simulated the antibody structure with antibody inherent character and function, a kind of therapeutic peptide is provided simultaneously and in external, body or original position intrinsic or the characteristic or the activity that obtain.Adopt method described herein, and, can change a plurality of parts of antibody of the present invention and the therapeutic peptide part of at least a EPO mimetic CH 1 deleted mimetibodies in conjunction with methods known in the art.
" EPO mimetic CH 1 deleted mimetibodies " used herein, " EPO mimetic CH 1 deleted mimetibodies part " or " EPO mimetic CH 1 deleted mimetibodies fragment " and/or " EPO mimetic CH 1 deleted mimetibodies variant " and similar analogies, all have or stimulate at least a protein, for example, but be not limited only among the SEQ ID NO:1-1110 at least a proteinic at least a part in conjunction with or at least a biological activity, such as external, original position and/or preferred intravital part in conjunction with or active.For example, suitable EPO mimetic CH 1 deleted mimetibodies specific part of the present invention or variant can be in conjunction with at least a protein ligands, and comprise at least a protein ligands, receptor, soluble recepter etc.But suitable EPO mimetic CH 1 deleted mimetibodies, specific part or the variant also signalling of scalable, raising, change, at least a protein acceptor of activation or other can be measured or detection of active.
Useful analogue body is characterised in that the suitable affinity that has with protein ligands or receptors bind in the inventive method and compositions, and optional and preferably have a hypotoxicity.Particularly, EPO mimetic CH 1 deleted mimetibodies with following situation is all useful in the present invention, i.e. independent component wherein, such as variable region, constant region (no CH1 part) and frame part, or its arbitrary portion (for example variable heavy or J, the D of light chain or the part in V district; Hinge region, constant heavy chain or light chain etc.) independent and/or common optional and preferably have a reduced immunogenicity.Can be applicable to that analogue body of the present invention is optional to be characterised in that but it has the long-term treatment patient, and can be from well to relief of symptoms capitally, and the low ability of toxicity.Reduced immunogenicity and/or high-affinity, and other not bright characteristic all may work to its attainable therapeutic effect." reduced immunogenicity " is defined in herein and is less than about 75%, or preferably be less than and significant HAMA, HACA occur among about 50,45,40,35,30,25,20,15,10,9,8,7,6,5,4,3,2 and/or 1% the patient or NAHA replys through treatment, and/or (the two antigen enzyme immunoassays of employing are determined as and are less than about 300 to occur low titre in through the patient of treatment, preferably be less than about 100) (seeing for example Elliott et al., Lancet344:1125-1127 (1994)).
Effectiveness
Isolating nucleic acid of the present invention can be used to generate at least a EPO mimetic CH 1 deleted mimetibodies, fragment or its specific variants, this EPO mimetic CH 1 deleted mimetibodies, fragment or its specific variants can be used at cell, tissue, organ or animal were worked in (comprising the mammal and the mankind), to regulate, treatment, it is conditions associated to alleviate at least a protein, help its morbidity of prevention, or reduce its symptom, this protein is conditions associated to be selected from, but be not limited only at least a immune disorder or disease, cardiovascular disorder or disease, infectious disease, malignant diseases and/or nervous disorder or disease, anemia; Immunity/autoimmune; And/or cancer/infectious disease, and other is known or particular proteins is conditions associated.
This method can comprise to the above-mentioned adjusting of needs, treatment, alleviation, prevention or reduce particular composition or the pharmaceutical composition that symptom, influence or machine-processed cell, tissue, organ, animal or patient use effective dose that said composition comprises at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant.This effective dose can comprise that single or multiple uses the amount of about 0.0001-500mg/kg, or single or multiple is used the amount that makes serum-concentration reach 0.0001-5000ug/ml serum, or any effective range or numerical value wherein, it is used and is described herein with method for measurement of concentration or the known method of association area.
Quote as proof
Whole publications cited herein or patent be owing to embodied this area present situation simultaneously with the present invention, and/or description of the invention and implementation method are provided, thereby be incorporated herein by reference by complete.Publication refers to any science or patent publications, or other any data of being provided of media form arbitrarily, comprises all records, electronics or layout.Following list of references all is incorporated herein by reference by complete: Ausubel, et al., ed., Current Protocolsin Molecular Biology, John Wiley ﹠amp; Sons, Inc., NY, NY (1987-2002); Sambrook, et al., Molecular Cloning:ALaboratory Mannual, 2 NdEdition, Cold Spring Harbor, NY (1989); Harlow and Lane, Antibodies, a Laboratory Manual, Cold SpringHarbor, NY (1989); Colligan, et al., eds., Current Protocolsin Immunology, John Wiley ﹠amp; Sons, Inc., NY (1994-2002); Colligan et al., Current Protocols in Protein Science, JohnWiley ﹠amp; Sons, NY, NY, (1997-2002).
Analogue body of the present invention
The EPO mimetic CH 1 deleted mimetibodies can comprise at least one CH3 district, this CH3 district directly links to each other with at least one CH2 district, this CH2 district directly links to each other with at least one hinge region or its fragment, this hinge region or its fragment directly link to each other with at least one part of V district, this part y district directly links to each other with an optional joint sequence, should directly link to each other with at least one therapeutic peptide by optional joint sequence, this therapeutic peptide then randomly further directly links to each other with at least one part of at least one variable antibody sequence.In a kind of preferred embodiment, a pair of CH3-CH2-hinge-part J sequence-joint-therapeutic peptide with optional N-terminal antibody sequence is by associating or covalent bond, for example, connecting paired but be not limited only to the Cys-Cys disulfide bond.Therefore, EPO mimetic CH 1 deleted mimetibodies of the present invention has been simulated the antibody structure with antibody inherent character and function, a kind of therapeutic peptide is provided simultaneously and in external, body or original position intrinsic or the characteristic or the activity that obtain.Adopt method described herein, and, can change a plurality of parts of antibody of the present invention and the therapeutic peptide part of at least a EP0 mimetic CH 1 deleted mimetibodies in conjunction with methods known in the art.
Therefore, analogue body of the present invention provides at least a proper characteristics that can compare with known protein matter, for example, but be not limited only at least a in the persistent period of the clearance rate of affinity, raising or the reduction of the half-life of prolongation, the activity of raising, more special activity, raising, elite or more suitable active hypotype, less immunogenicity, quality that at least a expection therapeutic effect is enhanced or prolongation, the less characteristics such as side effect.
The fragment of the analogue body of formula (I) can be passed through enzymatic lysis known in the art and/or described herein, synthetic or recombinant technique preparation.Utilize the specific antibodies gene also can prepare the analogue body of multiple clipped form, the one or more termination codoies in this specific antibodies gene have been imported into the upstream of natural termination site.A plurality of parts of analogue body can be linked together by chemistry by routine techniques, perhaps by utilizing gene engineering to be prepared as in abutting connection with albumen.For example, can express the nucleic acid of at least one constant region in the coding human antibodies chain, with generation can be applicable in the analogue body of the present invention in abutting connection with albumen.For example see Ladner et al., U.S.Patent No.4,946,778 and Bird, R.E.et al., Science, 242:423-426 (1988), the part of relevant single-chain antibody.
Term used herein " human analogue body " refers to a kind of antibody, wherein this proteinic each part (for example EPO simulating peptide, framework, C L, C HDistrict (C for example H2, C H3), hinge, (V L, V H)) all be required basic non-immunogenicity in the mankind basically, and only have less sequence variation or variation.For the human antibodies or analogue body of the present invention of unmodified, this variation or variation are optional and preferably keep in the mankind or reduced immunogenicity.Therefore, human antibodies and corresponding EPO mimetic CH 1 deleted mimetibodies of the present invention and chimeric or humanized antibody is completely different.Be to be noted that human antibodies and EPO mimetic CH 1 deleted mimetibodies can be generated by inhuman animal or cell, this inhuman animal or cell can be expressed human immunoglobulin (for example heavy chain) gene of rearrangement on function.
At suitable part, for example isolating and/or epo protein matter receptor or part, or its part (comprising synthetic molecules, for example synthetic peptide) can be designed the human analogue body that is specific at least a protein ligands or its receptor.The ligand binding domain of at least a protein or its part or the known technology of sequence can be identified and characterize to utilization, can prepare above-mentioned analogue body.
In a kind of preferred embodiment, the EPO mimetic CH 1 deleted mimetibodies comprises molecular formula (I):
(V1 (n)-Pep (n)-Flex (n)-V2 (n)-pHinge (n)-CH2 (n)-CH3 (n)) (m), wherein V1 is at least one part of immune globulin variable region N-terminal, Pep is the EPO simulating peptide of at least one biologically active, Flex is by making analogue body have optional orientation and binding characteristic, thereby provide the polypeptide of structural flexibility, V2 is at least one part of immune globulin variable region C-terminal, pHinge is at least one part of immunoglobulin variable hinge region, CH2 is at least one part of immunoglobulin CH2 constant region, CH3 is at least one part of immunoglobulin CH3 constant region, n and m can be the integer among the 1-10, simulated dissimilar immunoglobulin molecules, for example, but be not limited only to IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, IgM, IgD, IgE etc., or its combination.Monomer during m=1 can for example, link to each other with other monomer but be not limited only to the Cys-Cys disulfide bond by associating or covalent bond.
In a kind of preferred embodiment, at least a EPO mimetic CH 1 deleted mimetibodies of the present invention or specific part or variant are by at least a cell line, cell mixing system, immortalized cell, or the clonal population of immortalization and/or cultured cell generates.Adopt suitable method just can obtain the protein production cell of immortalization.Preferably, this at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant are prepared by specific nucleic acid or carrier are provided, this nucleic acid or carrier contain the DNA that is derived from least one human immunoglobulin site, perhaps have the sequence similar basically to this at least one human immunoglobulin site, wherein above-mentioned at least one human immunoglobulin site is reset by function or acceptable function is reset, above-mentioned nucleic acid or carrier can further comprise analogue body structure described herein, for example, but be not limited only to molecular formula (I), wherein as known in the art, the terminal variable region of C-and N-part can be used as V1 and V2, hinge region can be used as pHinge, CH2 can be used as CH2, and CH3 can be used as CH3.
Term used herein " function rearrangement " refers to derive from the nucleic acid fragment in specific immunoglobulins site, wherein this immunoglobulin locus has experienced V (D) J reorganization, thereby has generated the immunoglobulin gene of coding immunoglobulin chain (for example heavy chain) or its any part.The immunoglobulin gene that function is reset can obtain directly or indirectly identifying by adopting following appropriate method, suitable method for example nucleotide sequencing, is utilized the hybridization that probe carried out (for example Western blotting, RNA trace) can be annealed to the coding junction between the genetic fragment or is adopted the primer that can be annealed to the coding junction between the genetic fragment to enzymatic amplification (for example polymerase chain reaction) that immunoglobulin gene carried out.Whether cell has generated EPO mimetic CH 1 deleted mimetibodies or its part or variant also can be determined by suitable method, wherein this EPO mimetic CH 1 deleted mimetibodies or its part or variant have comprised specific variable region or have contained the variable region of particular sequence (for example, at least one Pep sequence).
Analogue body of the present invention, specific part and variant also can be by following method preparations, promptly utilize at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant code nucleic acid, to obtain transgenic animal or mammal, for example goat, milch cow, horse, sheep etc. make it generate above-mentioned analogue body or specific part or variant in milk.Adopt with the similar known method of method that is applied to antibody coding sequence and just can obtain above-mentioned animal.See, for example, but be not limited only to U.S. Patent No. 5,827,690; 5,849,992; 4,873,316; 5,849,992; 5,994,616; 5,565,362; 5,304,489 etc., these patents all by complete introducing herein as a reference.
Analogue body of the present invention, specific part and variant also can be in addition by following method preparations, promptly utilize at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant code nucleic acid, with the plant cell that obtains transgenic plant and cultivation (for example, but be not limited only to Nicotiana tabacum L. and Semen Maydis), make it at plant part or therefrom generate above-mentioned analogue body or specific part or variant in institute's cultured cells.A limiting examples, promptly the transgene tobacco leaf of express recombinant protein has been successfully used to the recombiant protein that provides a large amount of, for example, utilizes inducible promoter.See, for example Cramer et al., Curr.Top.Microbol.Immunol.240:95-118 (1999) and the list of references of quoting thereof.Equally, transgenic Semen Maydis or corn also have been used to be equal to by other recombination system generation or by the proteinic mammalian proteins matter of natural origin purification gained with commercial production levels expression biologic activity.See, for example Hood etal., Adv.Exp.Med.Biol.464:127-147 (1999) and the list of references of quoting thereof.Comprise antibody fragment, for example strand analogue body (scFv ' s) is also generated from the transgenic plant seed that comprises tobacco seed and potato tubers in large quantities at interior antibody.See, for example Conrad et al., Plant Mol.Biol.38:101-109 (1998) and the list of references of quoting thereof.Therefore, analogue body of the present invention, specific part and variant also can be according to known methods, utilize transgenic plant and prepared.Also see Fischer etal. for example, Biotechnol.Appl.Biochem.30:99-108 (in October, 1999), Maet al., Trends Biotechnol.13:522-7 (1995); Ma et al., PlantPhysiol.109:341-6 (1995); Whitelam et al., Biochem.Soc.Trans.22:940-944 (1994); And the list of references of being quoted in the above-mentioned literary composition.Above-mentioned list of references all is introduced in this as a reference by complete.
Analogue body of the present invention is affinity (K widely D) in conjunction with human protein's part.In a kind of preferred embodiment, at least a human EPO mimetic CH 1 deleted mimetibodies of the present invention can be randomly with high-affinity in conjunction with at least a protein ligands.For example, at least a EPO mimetic CH 1 deleted mimetibodies of the present invention can be equal to or less than about 10 -7The K of M D, or more preferred, be equal to or less than about 0.1-9.9 (or any range or numerical value wherein) X10 -7, 10 -8, 10 -9, 10 -10, 10 -11, 10 -12Or 10 -13M, or the K of any range or numerical value wherein DIn conjunction with at least a protein ligands.
The EPO mimetic CH 1 deleted mimetibodies can be able to measuring by adopting any appropriate method to the affinity or the affinity of at least a protein ligands, for example is used to measure the method for antibody-antigen binding affinity or affinity.(see, Berzofsky for example, et al., at Fundamental Immunology, Paul, W.E., Ed., Raven Press:NewYork, " the Antibody-Antigen Interaction " among the NY (1984); Kuby, Janis Immunology, W.H.Freeman and Company:New York, NY (1992); And the method for describing).The affinity difference of specific EPO mimetic CH 1 deleted mimetibodies-ligand interaction that different condition (for example salinity, pH) is measured down.Therefore, preferably adopt the standard solution of EPO mimetic CH 1 deleted mimetibodies and part and all standard buffers as described herein to measure affinity and other part incorporating parametric (K for example D, K a, K d).
Nucleic acid molecules
Utilize data provided herein, adopt and describe or methods known in the art herein, can obtain nucleotide sequence such as at least one part of the continuous amino acid of 90-100% at least of at least one sequence among the coding SEQ ID NO:1-42 and specific antibodies, wherein above-mentioned sequence is used as the Pep sequence of molecular formula (I) and inserts, can obtain EPO mimetic CH 1 deleted mimetibodies of the present invention, further comprise its specific fragment, variant or concensus sequence, perhaps can obtain the nucleic acid molecules of the present invention that comprised the deposition carrier of at least a above-mentioned sequence and encoded at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant.
Nucleic acid molecules of the present invention can be a rna form, such as mRNA, hnRNA, tRNA or other any form, or dna form, include but not limited to cDNA and by clone or the synthetic genomic DNA that is obtained, or its combination in any.This DNA can be three chains, two strands or strand, or the combination in any of three kinds of chain forms.The arbitrary portion of at least one chain of DNA or RNA all can be coding strand, also is usually said sense strand, can be noncoding strand perhaps, is also referred to as antisense strand.
Isolated nucleic acid molecule of the present invention can comprise and contain an open reading-frame (ORF), and randomly has the nucleic acid molecules of one or more introns, contains the nucleic acid molecules of the coded sequence of EPO mimetic CH 1 deleted mimetibodies or specific part or variant; With contain the nucleotide sequence different basically with above-mentioned nucleotide sequence, but because the degeneracy of genetic code, the nucleic acid molecules of at least a EPO mimetic CH 1 deleted mimetibodies described herein and/or known in the art of still encoding.Certainly, this genetic code is well known in the art.Therefore, to those skilled in the art, the above-mentioned degeneracy nucleic acid variant that the preparation code book is invented specific EPO mimetic CH 1 deleted mimetibodies or specific part or variant is routine work.See, for example above-mentioned Ausubel, et al., and above-mentioned nucleic acid variant also is included among the present invention.
So point out in the place, and the nucleic acid molecules of the present invention that comprises the nucleic acid of coding EPO mimetic CH 1 deleted mimetibodies or specific part or variant can include but not limited to, the nucleic acid molecules of the segmental aminoacid sequence of self coding EPO mimetic CH 1 deleted mimetibodies; The coded sequence of complete EPO mimetic CH 1 deleted mimetibodies or its part; The coded sequence of EPO mimetic CH 1 deleted mimetibodies, fragment or part, and other sequence, the coded sequence of or fusogenic peptide leading such as at least a signal, has or do not have above-mentioned other coded sequence, such as at least one intron, together with other non-coding sequence, include but are not limited to non-coding 5 ' and 3 ' sequence, such as transcribe, mRNA processing, comprise in montage and the polyadenylation signal and playing a role the transcribing of (for example-mRNA ribosome combination and stability), non-translated sequence; Other aminoacid of encoding is such as other functional amino acid whose other coded sequence can be provided.Therefore, the sequence of coding EPO mimetic CH 1 deleted mimetibodies or specific part or variant can be merged to labelled sequence, such as the sequence of coding particular peptide, this peptide helps purification to contain fusion EPO mimetic CH 1 deleted mimetibodies or the specific part or the variant of EPO mimetic CH 1 deleted mimetibodies fragment or part.
Polynucleotide with polynucleotide selective cross described herein
The invention provides can be under the selective cross condition with disclosed polynucleotide herein or comprise its specific variants or the isolating nucleic acid of other multi-nucleotide hybrid of part.Therefore, the polynucleotide of this embodiment can be used to separate, detect and/or quantitatively contain the nucleic acid of these polynucleotide.
Low or medium stringency hybridization condition typically but exclusively is not applied to compare with complementary series the sequence that sequence homogeneity is lowered.Medium and high stringency can randomly be used to the sequence of higher homogeneity.Low stringency can realize having the selective cross of the sequence of about 40-99% sequence homogeneity, and can be used to identify directly to homology or symbiosis homologous sequence.
Randomly, the coded EPO mimetic CH 1 deleted mimetibodies of the polynucleotide described from here of polynucleotide codified of the present invention or at least one part of specific part or variant.Polynucleotide of the present invention have comprised the nucleotide sequence that can be used to invent with code book the polynucleotide selective cross of EPO mimetic CH 1 deleted mimetibodies or specific part or variant.See, for example, above-mentioned Ausubel; Above-mentioned Colligan, all herein as a reference by complete introducing.
The structure of nucleic acid
Isolating nucleic acid of the present invention can adopt (a) well known in the art recombination method, (b) synthetic technology, (c) purification technique, or the combination of centralized way and being prepared.
This nucleic acid can comprise the sequence except that polynucleotide of the present invention easily.For example, the multiple clone site that comprises one or more endonuclease restriction sites can be inserted in this nucleic acid, to help separating these polynucleotide.Equally, but translation sequences can be inserted in this nucleic acid, to help separating translation polynucleotide of the present invention.For example, six histidine mark sequences provide purification the present invention proteinic conventional route.Nucleic acid of the present invention randomly also can be used as the carrier, conjugant or the joint that are used to clone and/or express polynucleotide of the present invention except that can be used as coded sequence.
Other sequence also can be added into above-mentioned clone and/or expressed sequence, optimizing it in the function aspect clone and/or the expression, to help separating polynucleotide of the present invention, or impels polynucleotide of the present invention to be imported into cell.The purposes of cloning vehicle, expression vector, conjugant and joint all is well known in the art.See for example above-mentioned Ausubel; Or above-mentioned Sambrook.
Make up the recombination method of nucleic acid
Utilize multiple cloning process well known by persons skilled in the art, all can from biological origin, obtain isolating nucleic acid compositions of the present invention, such as RNA, cDNA, genomic DNA or its combination in any.In some embodiment, adopted under suitable stringency can with the oligonucleotide probe of polynucleotide selective cross of the present invention, in cDNA or genome dna library, to identify needed sequence.The structure of the separation of RNA, cDNA and genomic library all is that those of ordinary skills know.(see for example above-mentioned Ausubel; Or above-mentioned Sambrook).
Be used to make up the synthetic method of nucleic acid
Isolating nucleic acid of the present invention also can be by known direct chemical synthetic method preparation (seeing for example above-mentioned Ausubel, et al.).Chemosynthesis generates single stranded oligonucleotide usually, by with complementary sequence hybridization, or by being template, adopt archaeal dna polymerase to carry out polymerization with this strand, this single stranded oligonucleotide can be converted into double-stranded DNA.This area any technical staff all will appreciate that, though the chemosynthesis of DNA can be limited to about 100 or the sequence of polybase base more, longer sequence can obtain by the connection of shorter sequence.
Recombinant expressed assembly
The present invention further provides the recombinant expressed assembly that comprises nucleic acid of the present invention.Nucleotide sequence of the present invention, for example the cDNA or the genome sequence of code book invention EPO mimetic CH 1 deleted mimetibodies or specific part or variant can be used to make up specific recombinant expressed assembly, and this assembly can be imported in a kind of at least ideal host cell.Recombinant expressed assembly will typically comprise can be operatively connected the polynucleotide of the present invention of transcribing starting adjusting sequence to specific, and wherein this is transcribed starting adjusting sequence and will guide polynucleotide of the present invention transcribing in specifying host cell.Allos and non-allos (promptly endogenous) promoter all can be used to guide expression of nucleic acids of the present invention.
In certain embodiments, the isolating nucleic acid that serves as promoter, enhancer or other element can be imported on the correct position of the non-allos form of the present invention polynucleotide (in upstream, downstream or the intron), regulates the expression of polynucleotide of the present invention with plus or minus.For example, as known in the art, endogenesis promoter can and/or substitute in vivo or external being changed by sudden change, disappearance.Polynucleotide of the present invention can have justice or antisense orientation to be expressed by specified.Should be understood that, can directly influence observable feature the control that justice or antisense orientation gene expression are arranged.The another kind of method that suppresses is to have justice to suppress.The nucleic acid that importing is set to sense orientation has been proved to be to block a kind of effective way that target gene is transcribed.
Carrier and host cell
The present invention also relates to comprise the carrier of isolated nucleic acid molecule of the present invention, via the host cell of this recombinant vector genetic modification, and by the preparation of recombinant technique known in the art at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant.See for example above-mentioned Sambrook, et al.; Above-mentioned Ausubel, et al., all herein as a reference by complete introducing.
Polynucleotide of the present invention can randomly be connected to specific support, but this carrier contains the selected marker that is useful in host's internal breeding.Usually, adopt suitable known method, with the plasmid vector transfered cell, other known method comprises utilizing and can be used as sedimentary carrier such as methods such as electroporations, as the calcium phosphate precipitation thing, or forms complex with electrically charged lipid.If this carrier is a virus, can adopts suitable package cell line with its external packing, and subsequently it be imported host cell.
DNA inserts should be connected to being operated property suitable promoter.This expression construct can further contain and optional be used at least a transcription initiation, termination, and is positioned at the site of transcriptional domain, and the ribosome binding site that is used to translate.The termination codon that the coded portion of the expressed mature transcript of this construct will preferably include the translation starting of beginning and suitably be positioned at mRNA end to be translated (for example, UAA, UGA or UAG), and the UAA and the UAG that are preferred for mammal or eukaryotic cell expression.
Expression vector can be preferred, but but also can randomly comprise at least a selected marker.Such labelling comprises, for example, but is not limited only to methotrexate (MTX), dihydrofolate reductase (DHFR, US Pat.Nos.4,399,216 that eukaryotic cell is cultivated; 4,634,665; 4,656,134; 4,956,288; 5,149,636; 5,179,017), ampicillin, neomycin (G418), mycophenolic acid or glutamine synthetase (GS, US Pat.Nos.5,122,464; 5,770,359; 5,827,739) tetracycline or the ampicillin resistance gene (above-mentioned patent all by complete introducing herein as a reference) cultivated at escherichia coli and other antibacterial or prokaryote of resistance and being used for.The suitable culture medium and the condition that are used for above-mentioned host cell are known in the art.Suitable carriers should be that those of skill in the art know.With the vector construction body import transfection that host cell can be by calcium phosphate transfection, the mediation of DEAE-glucosan, cation lipid mediation transfection, electroporation, transduce, infect or other known method is realized.This area is such as above-mentioned Sambrook, 1-4 and 16-18 chapter; Above-mentioned Ausubel has described these methods in 1,9,13,15,16 chapters.
At least a EPO mimetic CH 1 deleted mimetibodies of the present invention or specific part or variant all can be expressed by modified forms, such as fused protein, and can not only comprise secretion signal, also comprise other allos functional areas.For example, the additional aminoacid in a zone, especially charge residue can be added into the N-terminal of EPO mimetic CH 1 deleted mimetibodies or specific part or variant, thereby improves this EPO mimetic CH 1 deleted mimetibodies or specific part or variant in host cell, during the purification or stability and persistency between subsequent treatment and storage life.Equally, EPO mimetic CH 1 deleted mimetibodies of the present invention or specific part or variant also can add peptide moiety, thereby help its purification.Before final preparation EPO mimetic CH 1 deleted mimetibodies or its at least one fragment, above-mentioned zone can be removed.Many standard laboratory handbooks, such as above-mentioned Sambrook, 17.29-17.42 and 18.1-18.74 chapter; Above-mentioned Ausubel has all described said method in 16,17 and 18 chapters.
Those of ordinary skill in the art knows and can be used for expressing many expression systems that code book is invented proteinic nucleic acid.
The cell culture example useful to the preparation of analogue body of the present invention, specific part or its variant is mammalian cell.The mammal cell line system is generally the cell monolayer form, but also can adopt mammalian cell suspension or bioreactor.But this area has been developed a large amount of suitable host cell lines that the The expressed glycosylated protein, comprise COS-1 (for example ATCC CRL1650), COS-7 (for example ATCC CRL-1651), HEK293, BHK21 (for example ATCCCRL-10), (for example ATCC CRL 1610 for CHO, DG-44) and BSC-1 (for example ATCCCRL-26) cell line, the hepG2 cell, P3X63Ag8.653, SP2/0-Ag14,293 cells, HeLa cell etc., all can be easily from for example, American Type CultureCollection (U.S. typical case DSMZ), Manassas, Va obtains.Preferred host cell comprises the cell in lymph source, such as myeloma and lymphoma cell.Especially preferred host cell is P3X63Ag8.653 cell (ATCC numbers CRL-1580) and SP2/0-Ag14 cell (ATCC numbers CRL-1851).
The expression vector that is used for above-mentioned cell can comprise one or more following expression control sequencs, for example, but is not limited only to origin of replication; Promoter (for example latter stage or early stage SV40 promoter, CMV promoter (for example United States Patent(USP) Nos. 5,168,062; 5,385,839), HSV tk promoter, pgk (phosphoglyceric kinase) promoter, EF-1 α promoter (for example U.S. Patent No. 5,266,491), at least a human immunoglobulin promoter); Enhancer and/or machining information site, for example ribosome binding site, RNA splice site, polyadenylation site (for example the big T Ag of SV40 polyadenylic acid adds the angle of striking) and tanscription termination subsequence.See, for example, above-mentioned Ausubel et al.; Above-mentioned Sambrook, et al..To nucleic acid of the present invention or other useful cell of proteinic preparation all is known, and/or can from, for example American Type Culture Collection Catalogue of Cell Lines andHybridomas (U.S. typical case DSMZ's cell line and hybridoma catalogue) (www.atcc.org) or other known or commercial source obtain.
When using eukaryotic host cell, be that polyadenylation or transcription terminator typically are integrated in the above-mentioned carrier.An example of terminator sequence is the polyadenylation sequence in bovine growth hormone gene source.Also can comprise the sequence that is used for accurate montage transcript.An example of montage sequence is the VP1 intron (Sprague, et al., J.Virol.45:773-781 (1983)) in SV40 source.In addition, as known in the art, the gene order of duplicating in the control host cell can be integrated in the above-mentioned carrier.
The purification of EPO mimetic CH 1 deleted mimetibodies or specific part or its variant
EPO mimetic CH 1 deleted mimetibodies or specific part or variant can be by the methods of knowing, comprise, but be not limited only to A protein purification, ammonium sulfate or ethanol precipitation, acid extraction, anion or cation-exchange chromatography, phosphocellulose chromatography, hydrophobic interaction chromatography, affinity chromatography, hydroxyapatite chromatography and agglutinin chromatograph, from reconstitution cell, reclaimed and purification.Also can use high performance liquid chromatography (" HPLC ") and carry out purification.See Colligan for example, CurrentProtocols in Immunology, or Current Protocols in ProteinScience, John Wiley ﹠amp; Sons, NY, NY, (1997-2002), 1,4,6,8,9,10 chapters for example, all herein as a reference by complete introducing.
Analogue body of the present invention or specific part or variant comprise natural purification product, chemosynthesis operation product and by recombinant technique from eukaryotic cell, comprise the product that obtains in for example yeast, higher plant, insecticide and the mammalian cell.According to the host who uses in the reorganization preparation manipulation, EPO mimetic CH 1 deleted mimetibodies of the present invention or specific part or variant can be glycosylations or nonglycosylated, and be preferably glycosylated.Many standard laboratory handbooks, such as above-mentioned Sambrook, the 17.37-17.42 joint; Above-mentioned Ausubel, 10,12,13,16,18 and 20 chapters, above-mentioned Colligan, Protein Science has all described said method in the 12-14 chapter, all by complete introducing herein as a reference.
Analogue body, specific fragment and/or variant
Separation simulation body of the present invention comprises EPO mimetic CH 1 deleted mimetibodies or the specific part or the variant of any one polynucleotide encoding of the present invention of more complete argumentation from here, or the EPO mimetic CH 1 deleted mimetibodies of any separation or preparation or specific part or its variant.
Preferred EPO mimetic CH 1 deleted mimetibodies or ligand binding moiety or variant combine at least one epo protein part or receptor, thereby and provide respective egg white matter or its segmental at least a EPO biologic activity.Multiple all is well known in the art at the protein that has an important function aspect treatment or the diagnosis, and this proteinic suitable assay method or biologic activity also are well known in the art.
The limiting examples of for the purpose of the present invention, suitable EPO simulating peptide sees the following form 1.These peptides can be by open and/or methods known in the art preparation.Adopted the single-letter amino acid abbreviations in most of situation.In these sequences (and in the whole text in this description, in particular instance as do not have do not indicate in addition) X refer in 20 kinds of naturally occurring aminoacid any one, or known amino acid residue or its known derivant that may exist, or its any known modified amino acid.In these peptides any several all can have or jointless situation under series connection link to each other (promptly continuously), and provide several series connection continuous example in this table.Joint is listed in " A ", and can be arbitrary joint described herein.Series connection repeats and joint is separated with clear demonstration by dash.Any peptide that contains cysteinyl residue all can be randomly crosslinked with another peptide that contains cysteine, one of them or the two all can be connected on the carrier.Several crosslinked examples are provided in this table.Any peptide with an above cysteine residues all can form disulfide bond in the peptide; See, for example, the EPO simulating peptide in the table 1.Some examples of the peptide that connects by disulfide bond in the peptide have been listed in this table.In these peptides any one all can provide some derivatization examples by method described herein by derivatization in this table.For the derivant that carboxyl terminal can be added a cover by amino group, add a cover amino group and be represented as-NH 2For amino acid residue by the alternate derivant of non-amino acid residue part, should substitute and be represented as δ, represent any part known in the art, for example by complete introducing as a reference Bhatnagar et al. herein, (1996), J.Med.Chem.39:3814-9 and Cuthbertson et al. (1997), the part of describing among the J.Med.Chem.40:2876-82.J substitutes base and Z substitutes basic (Z defined in as a reference the U.S. Patent No. 5,608,035,5,786,331 and 5,880,096 by complete introducing herein 5, Z 6... .Z 40).For EPO simulated series (table 1), substituent X 2-X11 and integer " n " have been defined by the complete WO that is incorporated herein by reference 96/40772.The residue that runic shows is a D-aminoacid, but also can randomly be L-aminoacid.Without otherwise indicated, all peptides all connect by peptide bond.Abbreviation is listed in the latter end of this description.In " SEQ ID NO. " hurdle, " NR " refers to this particular sequence be need not sequence list.
Table 1-EPO-simulating peptide sequence
Sequence/structure;
SEQ ID NO：
YXCXXGPXTWXCXP 1
YXCXXGPXTWXCXP-YXCXXGPXTWXCXP 2
YXCXXGPXTWXCXP-A-YXCXXGPXTWXCXP 3
GGTYSCHFGPLTWVCKPQGG 5
GGDYHCRMGPLTWVCKPLGG 6
GGVYACRMGPITWVCSPLGG 7
VGNYMCHPGPITWVCRPGGG 8
GGLYLCRFGPVTWDCGYKGG 9
GGTYSCHFGPLTWVCKPQGG-10
GGTYSCHPGPLTWVCKPQGG-Δ-GGTYSCHFGPLTWVCKPQGG 11
GGTYSCHFGPLTWVCKPQGGSSK 12
GGTYSCHFGPLTWVCKPQGGSSK 13
GGTYSCHFGPLTWVCKPQGGSSK 14
GGTYSCHFGPLTWVCKPQGGSSK-Δ-GGTYSCHFGPLTWVCKPQGGSSK
Figure A0382011000322
GGTYSCHFGPLTWVCKPQGGSSK (Δ-biotin) 16
CX 4X 5GPX 6TWX 7C 17
GGTYSCHGPLTWVCKPQGG 18
VGNYMAHMGPITWVCRPGG 19
GGPHHVYACRMGPLTWIC 20
GGTYSCHFGPLTWVCKPQ 21
GGLYACHMGPMTWVCQPLRG 22
TIAQYICYMGPETWECRPSPKA 23
YSCHFGPLTWVCK 24
YCHFGPLTWVC 25
X 3X 4X 5GPX 6TWX 7X 8 26
YX 2X 3X 4X 5GPX 6TWX 7X 8 27
X 1YX 2X 3X 4X 5GPX 6X 7X 8X 9X 10X 1128
X 1YX 2CX 4X 5GPX 6TWX 7CX 9X 10X 11 29
GGLYLCRFGPVTWDCGYKGG 30
GGTYSCHFGPLTWVCKPQGG 31
GGDYHCRMGPLTWVCKPLGG 32
VGNYMCHFGPITWVCRPGGG 33
GGVYACRMGPITWVCSPLGG 34
VGNYMAHMGPITWVCRPGG 35
GGTYSCHFGPLTWVCKPQ 36
GGLYACHMGPMT％AIVCQPLRG 37
TIAQYICYMGPETWECRPSPKA 38
YSCHFGPLTWVCK 39
YCHFGPLTWVC 40
SCHFGPLTWVCK 41
(AX 2) nX 3X 4X 5GPX 6TWX 7X 8 42
The EPO biologic activity is well known in the art.See, for example the Erythropoietin has a mitegenic andpositive chemotactic effect on endothelial cells. (erythropoietin to endotheliocyte have mitogenesis and positive chemotaxis effect) of people in Proceedings of the National Academy of Science (USA) 87:5978-82 (1990) such as Anagnostou A; Fandrey J and the Jelkman WE Interleukin 1and tumor necrosisfactor-alpha inhibit eryt hropoietin production in vitro. (interleukin and tumor necrosis factor-alpha suppress the generation of external erythropoietin) in Annals of the New York Academy of Science628:250-5 (1991); The Recombinant human erythropoietin:A multipotentialhemopoietic growth factor in vivo and in vitro. of people such as Geissler K in Contrib.Nephrol.87:1-10 (1990) (recombinant human erythropoietin: in the body and external pluripotency hemopoietic growth factor); The Erythropoietin sensitivity as a differentiation marker in thehemopoietic system.Studies of three erythropoietic colonyresponses in culture. of Gregory CJ in Journal of Cellular Physiology 89:289-301 (1976) is (in hemopoietic system as the erythropoietin sensitivity of differentiation marker.To three kinds of researchs that the erythropoietin colony is replied in cultivating.) the Monokinesinhibiting erythropoietin production in human hepatomacultures and in isolated perfused rat kidneys. (suppress human hepatocytes tumor culture and separate perfusion kidney of rats in the monokine of generation erythropoietin) of people such as Jelkman W in Life Sci.50:301-8 (1992); The Human recombinant erythropoietindirectlystimulates Bcell immunoglobulin production and proliferationin serum-free medium.s (human recombinant erythropoietin directly stimulate B cell immunoglobulin generation and propagation in serum-free medium) of people in Clinical and Experimental Immunology85:151-6 (1991) such as Kimata H; The Erythropoietin enhances immunoglobulin production andproliferation by human plasma cells in a serum-free medium.s (erythropoietin in serum-free medium improved human plasmacytic immunoglobulin output and hypertrophy) of people in Clin.Immunology Immunopathol.59:495-501 (1991) such as Kimata H; The Effect of recombinant humanerythropoietin on human IgE production in vitro. (influence that recombinant human erythropoietin to external human IgE generate) of people such as Kimata H in Clinical and ExperimentalImmunology 83:483-7 (1991); Koury MJ and the BondurantMC Erythropoietin retards DNAbreakdown and prevents programmed cell death in erythroidprogenitor cells. (erythropoietin has postponed the DNA decomposition, and has stoped the programmed cell death in the erythrocyte CFU-GM) in Science 248:378-81 (1990); The Effect of recombinant humanerythropoietin on renal function in humans.s (recombinant human erythropoietin to the influence of human body renal function) of people in KidneyInternational 37:131-6 (1990) such as Lim VS; The Autocrinestimulation by erythropoiet in and autonomous growth of humanerythroid leukemic cells in vitro.s (autocrine stimulation of erythropoietin and human erythrocyte leukaemia in external autonomous growth) of people in Journal ofClinical Investigation 88:789-97 (1991) such as Mitjavila MT; The Performance of animmunoradiometric assay of erythropoietin and results forspecimens from anemic and polycythemic patients. of people such as Andre M in Clinical Chemistry 38:758-63 (1992) (performance of immunoradiometric assay erythropoietin and to the originate analysis result of sample of anemia and erythrocytosis patient); Erythropoietin-dependent anderythropoietin-producing cell lines.Implications forresearch and for leukemia therapy. (erythropoietin dependency and the erythropoietin cellulation system of people such as Hankins WD in Annals of the New York Academy ofScience 554:21-8 (1989).Meaning to research and leukemia treating); The Storage andpreparation of samples for erythropoietin radioimmunoassay.s (be used for preservation and the preparation of the sample of erythropoietin radioimmunoassay, RIA) of people in Clin.Lab.Haematology 13:189-96 (1991) such as Kendall RGT; The Comparison ofrelevant biological assays for the determination ofbiological active erythropoietin. (measure the comparison of the erythropoietin used associated biomolecule assay method of biologically active) of people such as KrumviehD in Dev.Biol.Stand.69:15-22 (1988); The Assessment of anEIA for measuring human serum erythropoietin as compared withRIA and an in-vitro bioassay. of people such as Ma DD in BritishJournal of Haematology 80:431-6 (1992) (comparing with vitro bioassay with RI A) to the assessment of the EIA that measures human serum erythropoietin; The A sensitivesandwich ELISA for measuring erythropoietin in human serum.s (a kind of sensitive sandwich ELISA method that be used for measure human serum erythropoietin) of people in BritishJournal of Haematology 80:285-92 (1992) such as Noe G; People such as Pauly JU are at Behring Institut Mitteilungen 90:112-25, (1991) the Highly specific and highly sensitive enzyme immunoassaysfor antibodies to human interleuk in 3 in, (IL3) and humanerythropoietin, (EPO) in serum., (at being situated between plain 3 between human leucocyte in the serum, (IL3) and human erythropoietin, the high specific of antibody (EPO) and high sensitivity enzyme immunoassay); Improved microbioassay for plasma erythropoietinbased on CFU-E colony formation. in Ann.Hematology 64:224-30 (1992) of Sakata S and Enoki Y (forming improvement microbiological assay) to the blood plasma erythropoietin based on the CFU-E colony; People such as Sanengen T are at Acta Physiol.Scand.135:11-6, (1989) the Immunoreactive erythropoietin anderythropoiesis stimulating factor in, (s) in plasma fromhypertransfused neonatal and adult mice.Studies with aradioimmunoassay and a cell culture assay for erythropoietin., (immunoreactivity erythropoietin and erythropoietic-stimulating factor in highly transfusion new life and the ripe mice source blood plasma.Adopt radioimmunoassay and measure the research of carrying out) at the cell culture of erythropoietin; A sensitive and specificerythropoietin immunoprecipitation assay:application topharmacokinetic studies. (a kind of sensitivity and the specificity erythropoietin immune elutriation: application in pharmacokinetics research) of people such as WidnessJA in Journal of Lab.Clin.Med.119:285-94 (1992); The individual cells system that other data adopts in also measuring referring to individual organisms.Above-mentioned list of references all by complete introducing herein as a reference.By using the cell line that to reply this factor,, can measure EPO such as HCD57, NFS-60, TF-1 and UT-7.By in colony-forming test, measuring the amount of medullary cell source CFU-E, also can evaluate the EPO activity.The RT-PCR quantitative method that a kind of optional and diverse detection method is a cytokine.
EPO mimetic CH 1 deleted mimetibodies or specific part or its variant can be in conjunction with specified protein or fragment parts, thereby and provide at least a activity, this activity is by making protein and at least a protein ligands or receptors bind, or rely on or mediation mechanism and being mediated in addition above-mentioned EPO mimetic CH 1 deleted mimetibodies or specific part or its variant part ground or preferably provide at least a above-mentioned specified protein basically or segmental at least a biological activity by other protein.Term used herein " EPO mimetic CH 1 deleted mimetibodies activity " refers to can be according to assay method with about 20-10,000%, preferably with about at least 60,70,80,90,91,92,93,94,95,96,97,98,99,100,110,120,130,140,150,160,170,180,190,200,250,300,350,400,450,500,550,600,700,800,900,1000,2000,3000,4000,5000,6000,7000,8000,9000% or regulate more at high proportion or to cause that at least a protein relies on active.
EPO mimetic CH 1 deleted mimetibodies or specific part or variant can provide at least a protein to rely on active ability and preferably be evaluated by at least a suitable protein biology mensuration described herein and/or known in the art.Human EPO mimetic CH 1 deleted mimetibodies of the present invention or specific part or variant can be similar to the immunoglobulin (IgG, IgA, IgM etc.) or the isotype of any kind of, and can comprise the k or the lambda light chain of at least one part.In a kind of embodiment, human EPO mimetic CH 1 deleted mimetibodies or specific part or variant have comprised IgG weight chain variable fragment, hinge region, CH2 and CH3, for example, and at least a isotype, IgG1, IgG2, IgG3 or IgG4.
At least a EPO mimetic CH 1 deleted mimetibodies of the present invention or specific part or variant combine has specific at least a specific ligand at least a protein, subunit, fragment, part or this combination in any of several.At least a EPO simulating peptide of at least a EPO mimetic CH 1 deleted mimetibodies of the present invention, specific part or variant can be randomly in conjunction with at least one specific ligand epi-position of this part.This can comprise the combination in any of at least a specific amino acids sequence in conjunction with epi-position, the contained aminoacid of 1-3 at least of this specific amino acids sequence, and the complete specific part of continuous amino acid is selected from protein ligands, such as one of EPO receptor or its part.
This analogue body can be by following method preparation, promptly adopt known technology that the different piece of the EPO mimetic CH 1 deleted mimetibodies of formula (I) is linked together, by adopting known recombinant DNA technology or other any appropriate method, such as at least a (being one or more) nucleic acid molecules of chemical synthesis preparation and this EPO mimetic CH 1 deleted mimetibodies of expression coding.
Adopt appropriate method, such as phage display (Katsube, Y., et al., Int.JMol.Med, 863-868 (1998)) or the method for applying transgene animal known in the art and/or described herein 1 (5):, can prepare and human protein's part or receptors bind, and comprise the analogue body of determining heavy or variable region of light chain.Just can express this EPO mimetic CH 1 deleted mimetibodies, specific part or variant by utilizing EPO mimetic CH 1 deleted mimetibodies, specific part or variant code nucleic acid or its part in the suitable host cell.
Preferably, this analogue body or its part binding fragment can with human EPO part or receptor high-affinity combine and (for example, be less than or equal to about 10 -7The K of M D).Comprise with the essentially identical aminoacid sequence of sequence described herein and to contain conservative amino acid replacement, and the sequence of aminoacid deletion and/or insertion.Conservative amino acid replacement refers to that first aminoacid is had second amino acid replacement of chemistry similar to it and/or physical characteristic (for example electric charge, structure, polarity, hydrophobicity/hydrophilic).Conservative substitution is included in down in the group aminoacid scope, and a seed amino acid is by another kind of amino acid replacement, that is: lysine (K), arginine (R) and histidine (H); Aspartic acid (D) and glutamic acid (E); Agedoite (N), glutamine (Q), serine (S), threonine (T), tyrosine (Y), K, R, H, D and E; Alanine (A), valine (V), leucine (L), isoleucine (I), proline (P), phenylalanine (F), tryptophan (W), methionine (M), cysteine (C) and glycine (G); F, W and Y; C, S and T.
Amino acid code
Forming the aminoacid of analogue body of the present invention or specific part or variant is abridged usually.By indicating this aminoacid by amino acid whose single-letter code, trigram code, title or three nucleotide codons well known in the art, just can express this aminoacid title and (see Alberts, B., et al., Molecular Biology of The Cell, Third Ed., GarlandPublishing, Ino., New York, 1994):
The single-letter code The trigram code Title The trinucleotide codon
A Ala Alanine GCA，GCC，GCG，GCU
C Cys Cysteine UGC，UGU
D Asp Aspartic acid GAC，GAU
E Glu Glutamic acid GAA，GAG
F Phe Phenylalanine UUC，UUU
G Gly Glycine GGA，GGC，GGG，GGU
H His Histidine CAC，CAU
I Ile Isoleucine AUA，AUC，AUU
K Lys Lysine AAA，AAG
L Leu Leucine UUA，UUG，CUA，CUC， CUG，CUU
M Met Methionine AUG
N Asn Agedoite AAC，AAU
P Pro Proline CCA，CCC，CCG，CCU
Q Gln Glutamine CAA，CAG
R Arg Arginine AGA，AGG，CGA，CGC， CGG，CGU
S Ser Serine AGC，AGU，UCA，UCC， UCG，UCU
T Thr Threonine ACA，ACC，ACG，ACU
V Val Valine GUA，GUC，GUG，GUU
W Trp Tryptophan UGG
Y Trp Tyrosine UAC，UAU
EPO mimetic CH 1 deleted mimetibodies of the present invention or specific part or variant can comprise defined derive from natural mutation or artificial one or more amino acid replacements, disappearance or the interpolation of handling herein.
Certainly, those of skill in the art should determine the quantity of amino acid replacement according to the many factors that comprise above-mentioned factor.In general, should not surpass 40,30,20,19,18,17,16,15,14,13,12,11,10,9,8,7,6,5,4,3,2,1 aminoacid to the amino acid replacement of at least a EPO mimetic CH 1 deleted mimetibodies, insertion or disappearance quantity, as specified 1-30 or any range wherein or numerical value herein.
In EPO mimetic CH 1 deleted mimetibodies of the present invention or specific part or variant, can be identified by methods known in the art the requisite aminoacid of function, such as site-directed mutagenesis or alanine scanning mutagenesis (for example above-mentioned Ausubel, 8,15 chapters; Cunninghamand Wells, Science 244:1081-1085 (1989)).All imported the single alanine sudden change on a kind of each residue that operates in the molecule in back.Subsequently, the biological activity of the check mutating molecule that obtains for example, but is not limited only to illustrate or at least a protein-related activity known in the art herein.Critical sites for EPO mimetic CH 1 deleted mimetibodies or specific part or variant combination also can be by being identified (Smith such as the structured analysis method of crystallization, nuclear magnetic resonance, NMR or photoaffinity labeling, et al., J.Mol.Biol.224:899-904 (1992) and de Vos, et al., Science 255:306-312 (1992)).
Analogue body of the present invention or specific part or variant can comprise the Pep part in the molecular formula (I), but are not limited only at least one part, the sequence of at least one sequence among the SEQ ID NO:1-42 or are selected from 3 combinations that sequence arrives all sequences among the SEQ ID NO:1-42.Can strengthen or keep at least a above-mentioned active non-limiting variant to comprise, but be not limited only to, arbitrary aforementioned polypeptides, further comprise at least a sudden change, at least a displacement, the insertion of this sudden change and the suitable biological activity of not appreciable impact EPO mimetic CH 1 deleted mimetibodies of the present invention or function or lack corresponding.
EPO mimetic CH 1 deleted mimetibodies or specific part or variant can further randomly comprise at least one funtion part as at least a polypeptide of Pep part in the molecular formula (I), and at least one sequence has the homogeneity of 90-100% among this funtion part and the SEQ ID NOS:1-42 ").The EPO mimetic CH 1 deleted mimetibodies can further randomly comprise the Pep part that can be used as in the molecular formula (I), and is selected from the aminoacid sequence of the one or more sequences among the SEQ ID NO:2-41.
In a kind of embodiment, the corresponding aminoacid sequence of the appropriate section of at least one sequence has about 90-100% homogeneity (promptly 90,91,92,93,94,95,96,97,98,99,100 or any range wherein or numerical value) among Pep aminoacid sequence or its part and the SEQ ID NO:1-42.Preferably, utilize suitable computerized algorithm known in the art to determine 90-100% aminoacid homogeneity (promptly 90,91,92,93,94,95,96,97,98,99,100 or any range or numerical value wherein).
Analogue body of the present invention or specific part or variant can comprise any amount continuous amino acid residue that derives from EPO mimetic CH 1 deleted mimetibodies of the present invention or specific part or variant, and wherein this quantity is selected from by continuous one of group of integers of forming of the 10-100% of residue quantity in the EPO mimetic CH 1 deleted mimetibodies.Randomly, the subsequence of continuous amino acid contains about 2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19,20,21,22,23,24,25,26,27,28,29,30,40,50,60,70,80,90,100,110,120,130,140,150,160,170,180,190,200,210,220,230,240,250 or amino acids more at least on length, or contains the aminoacid of any range wherein or numerical value.In addition, the quantity of this subsequence can be the arbitrary integer that is selected from 1-20, such as at least 2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19,20 or bigger integer.
The technical staff it should be understood that EPO mimetic CH 1 deleted mimetibodies of the present invention or specific part or the variant that the present invention includes at least a biologically active.The given activity of the analogue body of biologically active or specific part or variant is active at least 20%, 30% or 40% of natural (non-synthetic), endogenous or relevant and known insertion or fusion rotein or specific part or a variant, be preferably at least 50%, 60% or 70%, most preferably at least 80%, 90% or 95%-100%.The method of analysis and quantitation of enzyme activity and substrate specificity is well known to those skilled in the art.
On the other hand, the present invention relates to described herein by covalent attachment organic moiety and adorned human analogue body and part binding fragment.This modification can produce and have the improvement PK (pharmacokinetic) profile EPO mimetic CH 1 deleted mimetibodies or the part binding fragment of (for example serum half-life in the body of Yan Changing).Organic moiety can be line style or branch's hydrophilic polymeric group, fatty acid group or fatty acid ester group.In the particular, the hydrophilic polymeric group can have about 800-about 120,000 daltonian molecular weight, and can be polyalkane glycol (for example Polyethylene Glycol (PEG), polypropylene glycol (PPG)), glycopolymers, amino acid polymer or polyvinylpyrrolidone, and fatty acid or fatty acid ester group can comprise about 40 carbon atoms of about 8-.
Modification analogue body of the present invention and part binding fragment can comprise one or more and EPO mimetic CH 1 deleted mimetibodies or specific part or the covalently bound directly or indirectly organic moiety of variant.With EPO mimetic CH 1 deleted mimetibodies of the present invention or bonded each organic moiety of part binding fragment can independently be hydrophilic polymeric group, fatty acid group or fatty acid ester group all.Term " fatty acid " used herein " comprise mono carboxylic acid and two carboxylic acids.Term used herein " hydrophilic polymeric group " refers to organic polymer more easily molten in water than in octane.For example, polylysine is easier to be molten in water than in octane.Therefore, the present invention includes the EPO mimetic CH 1 deleted mimetibodies of being modified by the covalent attachment polylysine.Be applicable to that the hydrophilic polymer of modifying analogue body of the present invention can be line style or branch-like, and comprise, for example the polymer (for example polylysine, poly arginine, poly-aspartate etc.) of polyalkane glycol (for example PEG, mono methoxy-Polyethylene Glycol (mPEG), PPG etc.), saccharide (for example glucosan, cellulose, oligosaccharide, polysaccharide etc.), hydrophilic amino acid, poly-epoxyalkane (for example poly(ethylene oxide), poly(propylene oxide) etc.) and polyvinylpyrrolidone.Preferably, when the hydrophilic polymer of modifying EPO mimetic CH 1 deleted mimetibodies of the present invention is the unimolecule entity form, have about 800-about 150,000 daltonian molecular weight.For example, can adopt PEG 2500, PEG 5000, PEG 7500, PEG 9000, PEG 10000, PEG 12500, PEG 1500And PEG 20,000, wherein be designated as dalton's mean molecule quantity of this polymer down.
The hydrophilic polymeric group can be by 1 to about 6 alkyl, fatty acid or fatty acid ester group displacement.Can be by fatty acid or the metathetical hydrophilic polymer of fatty acid ester group by using suitable method preparation.For example, the polymer that comprises amine groups can be coupled on the carboxyl of fatty acid or fatty acid ester, and the activated carboxyl on fatty acid or the fatty acid ester (for example via N, the activation of N-carbonyl dimidazoles) can be coupled to the oh group on the polymer.
Be applicable to that fatty acid and the fatty acid ester of modifying analogue body of the present invention can be saturated one or more unsaturated units that maybe can contain.Be applicable to that the fatty acid of modifying analogue body of the present invention comprises, for example, n-dodecylic acid (C 12, lauric acid), n-four capric acid (C 14, myristic acid), n-eight capric acid (C 18, stearic acid), n-arachic acid (C 20, arachic acid), n-behenic acid (C 22, mountain Yu acid), n-melissic acid (C 30), n-tetracontane acid (C 40), suitable-Δ 9-octadecanoid acid (C 18, oleic acid), all suitable-Δs 5,8,11,14-eicosatetraenoate (C 20, arachidonic acid), suberic acid, four decanedioic acid, eight decanedioic acid, docosandioic acid etc.The suitable fatty acids ester comprises the monoesters of the dicarboxylic acids that contains line style or branch's low-grade alkyl group.Can comprise 1 to about 12 than low-grade alkyl group, preferred 1 to about 6 carbon atoms.
Human analogue body of modifying and part binding fragment can adopt such as, by with the appropriate method preparation of one or more dressing agents reactions.Term used herein " dressing agent " refers to comprise the suitable organic group (for example hydrophilic polymer, fatty acid, fatty acid ester) of activated group." activated group " refers to can be under proper condition and the reaction of second chemical group, thus between above-mentioned dressing agent and second chemical group chemical part or the functional group of formation covalent bond.For example, the reactive activated group of amine comprises electrophilic group, such as tosylate, methanesulfonates, halogenation (chlorine, bromine, fluorine, iodine), N-hydroxyl succinimide ester (NHS) etc.Can comprise with the activated group of thiol reactant, for example maleimide, iodacetyl, acryloyl, pyridyl disulfide, 5-thiol-2-nitrobenzoic acid mercaptan (TNB-mercaptan) etc.Aldehyde functional group group can be coupled to contain amine-or the molecule of hydrazides on, and azido group can with the three valent phosphors radical reaction, thereby form phosphoramidate or phosphamide key.The appropriate method that activated group is imported molecule is (seeing Hermanson for example, G.T., Bioconjugate Techniques, AcademicPress:San Diego, CA (1996)) known in the art.Activated group can directly be bonded to organic group (for example hydrophilic polymer, fatty acid, fatty acid ester), or by blank area, for example bivalence C 1-C 12Group is realized combination, and wherein one or more carbon atoms can be replaced by the hetero atom such as oxygen, nitrogen or sulfur.Suitable blank area comprises, for example TEG ,-(CH 2) 3-,-NH-(CH 2) 6-NH-,-(CH 2) 2-NH-and-CH 2-O-CH 2-CH 2-O-CH 2-CH 2-O-CH-NH-.The dressing agent that comprises blank area can be by following method preparation, even single Boc (tert-butyl group)-alkyl diamine (for example single Boc-ethylene diamine, single Boc-diamino hexane) under the existence condition of 1-ethyl-3-(3-two methyl aminopropyls) carbodiimides (EDC) and fatty acid response, thereby between free amine and fatty acid carboxylate, form amido link.Can from product, remove the Boc blocking group by following method; promptly adopt trifluoroacetic acid (TFA) to handle; with expose can with described another carboxylic acid coupling; or can with the primary amine of maleic anhydride reaction, and make the obtained product cyclisation, generate the maleimide derivatives that this fatty acid is activated and (see; Thompson for example; et al., WO92/16221, its complete content is incorporated herein by refere).
Modification analogue body of the present invention can be generated by making the reaction of human EPO mimetic CH 1 deleted mimetibodies or part binding fragment and dressing agent.For example, by using the reactive dressing agent of amine, for example the NHS ester of PEG can not have the locus specificity mode organic moiety is bonded to the EPO mimetic CH 1 deleted mimetibodies.Human analogue body of modifying or part binding fragment also can be prepared by the disulfide bond (for example intrachain disulfide bond) of reduction EPO mimetic CH 1 deleted mimetibodies or part binding fragment.Subsequently, can make the reaction of reductive EPO mimetic CH 1 deleted mimetibodies or part binding fragment and thiol-reactive dressing agent, to obtain modification EPO mimetic CH 1 deleted mimetibodies of the present invention.By adopting appropriate method, such as reverse Proteolytic enzyme (Fisch et al., Bioconjugate Chem., 3:147-153 (1992); Werlen et al., Bioconjugate Chem., 5:411-417 (1994); Kumaran et al., ProteinSci.6 (10): 2233-2241 (1997); Itoh et al., Bioorg.Chem., 24 (1): 59-68 (1996); Capellas et al., Biotechnol.Bioeng., 456-463 (1997)) and Hermanson 56 (4):, G..T., BioconjugateTechniques, Academic Press:San Diego, the described method of CA (1996), can prepare human analogue body of the modification that contains specific organic moiety and part binding fragment, wherein this specific organic moiety is incorporated in to the specific site of EPO mimetic CH 1 deleted mimetibodies of the present invention or specific part or variant.
EPO mimetic CH 1 deleted mimetibodies compositions
The present invention also provides at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant compositions, said composition has comprised described herein and/or known in the art, at least a, at least two kinds, at least three kinds, at least four kinds, at least five kinds, at least six kinds or more analogue bodies or specific part or its variant that exist compositions, mixture or form to be provided with non-natural.The ratio of said composition is by weight, volume, concentration, molar concentration, and the molar concentration of liquid known in the art or described herein or dry solution, mixture, suspension, Emulsion or colloidal form is represented.
Said composition can comprise the weight of 0.00001-99.9999%, volume, concentration, molar concentration liquid known in the art or described herein, gas or dry solution, mixture, suspension, the molar concentration of Emulsion or colloidal form, or any range wherein or numerical value "; for example, but be not limited only to 0.00001; 0.00003; 0.00005; 0.00009; 0.0001,0.0003,0.0005,0.0009,0.001,0.003,0.005,0.009,0.01,0.02,0.03,0.05,0.09,0.1,0.2,0.3,0.4,0.5,0.6,0.7,0.8,0.9,1.0,1.1,1.2,1.3,1.4,1.5,1.6,1.7,1.8,1.9,2.0,2.1,2.2,2.3,2.4,2.5,2.6,2.7,2.8,2.9,3.0,3.1,3.2,3.3,3.4,3.5,3.6,3.7,3.8,3.9,4.0,4.3,4.5,4.6,4.7,4.8,4.9,5,6,7,8,9,10,15,20,25,30,35,40,45,50,55,60,65,70,71,72,73,74,75,76,77,78,79,80,81,82,83,84,85,86,87,88,89,90,91,92,93,94,95,96,97,98,99,99.1,99.2,99.3,99.4,99.5,99.6,99.7,99.8,99.9%.Therefore, said composition of the present invention include but not limited to 0.00001-100mg/ml and/or 0.00001-100mg/g.
Said composition can randomly further comprise at least a chemical compound or the protein of effective amount of drug such as being selected from least a anti-infectives, cardiovascular (CV) system medicine, central nervous system (CNS) medicine, autonomic nervous system (ANS) medicine, respiratory drugs, gastrointestinal (GI) tract drug, hormonal medicaments, the medicine that is used for body fluid or electrolyte balance, blood substance, antitumor, immunoregulation medicament, eye, ear or nose medication, local application, nutritional drugs.Said medicine all is well known in the art, comprises prescription, indication, the dosage of listed each medicine herein and uses and (see Nursing 2001Handbook of Drugs for example, 21 StEdition, Springhouse Corp., Springhouse, PA, 2001; HealthProfessional ' s Drug Guide 2001, ed., Shannon, Wilson, Stang, Prentice-Hall, Inc, Upper Saddle River, NJ; PharmcotherapyHandbook, Wells et al., ed., Appleton ﹠amp; Lange, Stamford, CT, all herein as a reference) by complete introducing.
Anti-infectives can be at least a medicine that is selected from amebacide or at least a antiprotozoal, anthelmintic, antifungal agent, antimalarial, antitubercular agent or at least a antileprotic, aminoglycoside medicine, penicillin, cephalosporin, tetracycline, sulfanilamide, fluoroquinolone, antiviral agents, macrolide anti-infective, the comprehensive anti-infective.The CV medicine can be at least a medicine that is selected from variable force medicine, antiarrhythmics, anti-angina pectoris medicine, antihypertensive, antilipemic, the comprehensive cardiovascular drugs.The CNS medicine can be at least a medicine that is selected from non-narcotic analgesics, or be selected from least a medicine in antipyretic, on-steroidal anti-inflammatory medicaments, the anesthetics, or at least a opium sample analgesic, calmness-hypnotic, anticonvulsant, antidepressants, antianxiety drugs, psychosis, central nervous system's stimulant, antiparkinsonism drug, comprehensive medicine for central nervous system.The ANS medicine can be at least a medicine that is selected from cholinergic drug (parasympathomimetic agent), anticholinergic, beta adrenergic agent (sympathomimetic), adrenergic blocking drug (sympatholytic), skeletal muscle relaxant, the neuromuscular blocking agents.Respiratory drugs can be at least a medicine that is selected from hydryllin, bronchodilatation medicine, the expectorant or at least a cough medicine, comprehensive respiratory medications.The GI tract drug can be at least a medicine that is selected from antacid, or at least a adsorbent, or at least a antiflatulent, digestive enzyme or at least a cholelithiasis solubilizing agent, diarrhea, laxative, antiemetic, antiulcerative.Hormonal medicaments can be at least a medicine that is selected from corticosteroids, the androgen, or at least a anabolic steroids, estrogen or at least a progesterone, promoting sexual gland hormone, antidiabetic medicine or at least a glucagon, thyroid hormones, thyroid hormones antagonist, pituitary hormone, class parathyroid gland medicine.The medicine that is used for balance body fluid and electrolyte can be at least a medicine or at least a substitutional solution, acidulant or at least a basifier that is selected from diuretic, electrolyte.Blood substance can be at least a medicine that is selected from hematonic, anticoagulant, blood derivatives, the streptokinase.Antitumor drug can be the antitumor drug that is selected from alkylation medicine, antimetabolite, antibiotics antitumor drug, can changes hormonal balance, at least a medicine in the comprehensive antitumor drug.Immunoregulation medicament can be at least a medicine that is selected from immunosuppressant, the vaccine, or at least a toxoid, antitoxin or at least a venom, immune serum, biological respinse modifier.Eye, ear and nose medication can be at least a medicine that is selected from ophthalmology anti-infective, ophthalmology anti-inflammatory agent, miotic, mydriatic, ocular angiogenesis contracting agent, comprehensive eye, ear, the nose medication.Local application can be at least a medicine that is selected from local anti-infective agent, the antiscabietic or at least a antipedicular agent, local corticosteroids.Nutritional drugs can be at least a medicine that is selected from vitamin, mineral or the caloic.See the content among for example above-mentioned Nursing 2001Drug Handbook.
Above-mentioned at least a amebacide or antiprotozoal can be at least a medicine that is selected from atovaquone, chloroquine hydrochloride, chloroquine phosphate, metronidazole, hydrochloric acid metronidazole, the pentamidine isethionate.Above-mentioned at least a anthelmintic can be at least a medicine that is selected from mebendazole, pamoic acid quinoline-pyrimidine, the thiabendazole.Above-mentioned at least a antifungal can be at least a medicine that is selected from amphotericin B, amphotericin B cholesteryl sulphate complex, amphotericin B lipid complex, AM Bison, fluconazol, flucytosine, microsize griseofulvin, ultra micro small size griseofulvin, itraconazole, ketoconazole, nystatin, the terbinafine HCl.Above-mentioned at least a antimalarial can be and be selected from chloroquine hydrochloride, chloroquine phosphate, doxycycline, sulphuric acid hydroxylation chloroquine, hydrochloric acid mefloquine, primaquine phosphate, Pyrimethamine, contains at least a medicine in the Pyrimethamine of sulfamethoxine.Above-mentioned at least a antitubercular agent or antileprotic can be at least a medicine that is selected from clofazimine, cycloserine, dapsone, ebutol, isoniazid, pyrazinamide, rifabutin, rifampicin, rifapentine, the streptomycin sulfate.Above-mentioned at least a aminoglycoside medicaments can be at least a medicine that is selected from amikacin sulfate, sulfate gentamicin, polygynax, streptomycin sulfate, the tobramycin sulfate.Above-mentioned at least a penicillin can be and is selected from amoxicillin/potassium clavulanate, BRL-2333, ampicillin, sodium ampicillin, Ampicillin Trihydrate, sodium ampicillin/sulbactam sodium, Cloxacillin, dicloxacillin, mezlocillin sodium, sodium ethoxynaphthamidopenicillanate, toluene isoxazolyl penicillin sodium, benzathine penicillin G, scotcil, aquacillin, penicillin G sodium, potassium v calcium, piperacillin sodium, piperacillin sodium/sodium-tazobactam, the ticarcillin disodium, at least a in ticarcillin disodium/potassium clavulanate.Above-mentioned at least a cephalosporin can be and is selected from least a cefaclor, cefadroxil, the cefonicid sodium element, cefdinir, cefepime hydrochloride, cefixime, cyanogen azoles methoxy cefonicid sodium element, cefonicid sodium, oxygen piperazine oxybenzene azoles head sodium, cefotaxime sodium, the Cefotetan disodium, thiophene methoxy cefonicid sodium element, Cefpodoxime Proxetil, cefprozil, ceftazidime, ceftibuten, ceftizoxime sodium, ceftriaxone sodium, CEFUROXIME AXETIL, Cefuroxime Sodium, cefalexin hydrochloride, one hydration cefalexin, Cefradal, at least a in the Loracarbef.Above-mentioned at least a tetracycline can be and is selected from least a in demeclocycline hydrochloride, doxycycline calcium, doxycycline hyclate, doxycycline hydrochloride, a hydration doxycycline, minocycline hydrochloride, the quadracycline.Above-mentioned at least a sulfanilamide can be and is selected from least a in compound sulfamethoxazole methyl isoxazole, sulfadiazine, sulfamethoxazole, sulfanilamide Sulfafurazole, the acetyl sulfisoxazole.Above-mentioned at least a fluoroquinolone can be and is selected from least a in my trovafloxacin of methanesulfonic acid, ciprofloxacin, enoxacin, levofloxacin, lomefloxacin hydrochloride, nalidixan, norfloxacin, ofloxacin, Sparfloxacin, the trovafloxacin mesilate.Above-mentioned at least a antiviral drugs can be and is selected from abacavir sulfate, Acyclovir Sodium, amantadine hydrochloride, amprenavir, cidofovir, methanesulfonic acid dilazep dimension is fixed, didanosine, Yi Feiweilun, famciclovir, Fomivirsen sodium, foscarnet sodium, ganciclovir, indinavir sulfate, lamivudine, lamivudine/neat many furans pyridine, nelfinavir mesilate, interior Wella is flat, oseltamivir phosphate, ribavirin, rimantadine hydrochloride, ritonavir, saquinavir, the methanesulfonic acid saquinavir, take charge of his furan pyridine, valaciclovir hydrochlordide, zalcitabine, Zha Namiwei, at least a in neat many furans pyridine.Above-mentioned at least a macrolide anti-infective can be and is selected from least a in azithromycin, clarithromycin, dirithromycin, erythromycin substrate, erythromycin propionate lauryl sulfate, SAI NENG SHA, erythromycin lactobionate, the erythromycin octadecanoate.Above-mentioned at least a comprehensive anti-infective can be and is selected from aztreonam, bacitracin, chloromycetin sodium succinate, clindamycin hydrochloride, clindamycin palmitate hydrochloride, clindamycin phosphate, imipenum.And Cilastatin Sodium,, at least a in the meropenem, nitrofurantoin megacryst, nitrofurantoin microcrystal, quinupristin/dalfopristin, spectinomycin hydrochloride, trimethoprim, Lyphocin (Fujisawa).(seeing, for example the 24-214 page or leaf among the Nursing 2001Drug Handbook).
Above-mentioned at least a variable force medicine can be and is selected from least a of the upright agricultural middle school of lactic acid AMR, digoxin, lactic acid rice.Above-mentioned at least a anti-arrhythmic can be and is selected from adenosine, Amiodarone Hydrochloride, atropine sulfate, the toluenesulfonic acid bretylium tosylate, the hydrochloric acid DILTIAZEM HCl, norpace (disopyramide), disopyramide phosphate, esmolol hydrochloride, the acetic acid flecainide, Ibolite fumarate, lidocaine hydrochloride, hydrochloric acid Mai Xileiding, Moracizine Hydrochloride, phenytoin, phenytoin Sodium, procamide, propafenone hydrochloride, propranolol hydrochloride, the bisulphate chinidine, the gluconic acid chinidine, the polygalacturonic acid chinidine, quinidine sulfate, Suo Daluo, tocainide hydrochloride, at least a in the hydrochloric acid verapamil.Above-mentioned at least a anti-angina pectoris medicine can be and is selected from least a in Amlodipine Besylate Tablet, amyl nitrite, bepridil hydrochloride, hydrochloric acid DILTIAZEM HCl, isosorbide dinitrate, an isosorbidi dinitras, nadolol, Licardipine Hydrochloride, nifedipine, nitroglycerin, phloride, verapamil, the hydrochloric acid verapamil.Above-mentioned at least a antihypertensive can be and is selected from Acebutolol Hydrochloride, benzenesulfonic acid A Muluohe, atenolol, benazepril hydrochloride, hydrochloric acid Beta Luo Er, the bisoprolol fumarate, candesartan Cilexetil, captopril, the hydrochloric acid galutedin, carvedilol, clonidine, dichloranilino imidazolin, diazoxide, the hydrochloric acid DILTIAZEM HCl, Carclura, enalaprilat, enalapril maleate, the methanesulfonic acid Eprosartan, felodipine, Fenoldopam Mesylate, fosinopril sodium, the acetic acid guanabenz, guanadrel sulfate, hydrochloric acid paddy ammonia process is new, hydralazine hydrochloride, irbesartan, isradipine, Labetalol Hydrochloride, lisinopril, Losartan Potassium, methyldopa, Aldomet Ester Hydrochloride salt, the succinic acid metoprolol, the tartaric acid metoprolol, Minoxidil, CI-925, nadolol, Licardipine Hydrochloride, nifedipine, nisin, Nitroprusside sodium, penbutolol sulfate, perindopril erbumine, the fragrant Chlorazene of methanesulfonic acid, pindolol, minipress, propranolol hydrochloride, quinapril hydrochloride, ramipril, telmisartan, terazosin hydrochloride, timolol maleate, trandolapril, valsartan, at least a in the hydrochloric acid verapamil.Above-mentioned at least a antilipemic can be and is selected from least a in Atorvastatin calcium, cerivastatin sodium, Cholestyramine, colestipol hydrochloride, fenofibrate (micronization), Fluvastatin Sodium, gemfibrozil, lovastatin, nicotinic acid, pravastatin sodium, the simvastatin.Above-mentioned at least a comprehensive CV medicine can be and is selected from abciximab., at least a in the Alprostadil, hydrochloric acid arbutamine, cilostazol, two Clopidogrel Hydrogensulfates, dipyridamole, Eptifibatide, Midodrine Hydrochloride, pentoxifylline, ticlopidine hydrochloride, tirofiban hydrochloride.(seeing, for example the 215-336 page or leaf among the Nursing 2001Drug Handbook).
Above-mentioned at least a non-narcotic analgesics or antipyretic can be and be selected from least a in acetaminophen, aspirin, Choline magnesium trisalicylate, diflunisal, the salicylic acid enzyme.Above-mentioned at least a on-steroidal anti-inflammatory medicaments can be and is selected from least a in celecoxib, Ciclofenaziae potassium, Ciclofenaziae sodium, Yi Tuoduo thunder, fenopron, flurbiprofen, ibuprofen, indomethacin, three hydration indomethacin sodium, ketone propanoic acid, ketorolac tromethamine, nabumetone, naproxen, naproxen sodium, promazine, piroxicam, rofecoxib, the sulindac.Above-mentioned at least a anesthetics or opium sample analgesic can be and be selected from Fentanyl, buprenorphin hydrochloride, butorphanol tartrate, codeine phosphate, codeine sulfate, fentanyl citrate, fentanyl is through dermal system, fentanyl is striden mucosa, dihydro-morphinone hydrochloride, pethidine, methadone hydrochloride, morphine hydrochloride, morphine sulfate, morphine tartrate, nalbuphlne hydrochloride, oxycodone hydrochloride, pectination 1,4-hydroxyl dihydrocodeinone, the hydrochloric acid oxymorphone, pentazocine hydrochloride, pentazocine hydrochloride and naloxone hydrochloride, pentazocine lactate, PROPOXY PHEN HCL, the LOMAR PWA EINECS 246-676-2 dextropropoxyphene, remifentanil hydrochloride, sufentanil citrate, at least a in the tramadol hydrochloride.Above-mentioned at least a analgesic-hypnotic can be and is selected from least a in chloral hydrate, triazole nitrogen, flurazepam hydrochloride, pentobarbital, pentobarbital sodium, sodium phenobarbital, barbose, dicoumarol, triazole benzene phenodiazine, zaleplon, the Zolpidemtar Trate.Above-mentioned at least a anticonvulsant can be and is selected from acetazolamide sodium, carbamazepine, clonazepam, chlorine nitrogen dipotassium, stable, divalproex sodium, ethosuximide, fosphenytoin sodium, gabapentin, lamotrigine, magnesium sulfate, phenobarbital, sodium phenobarbital, phenytoin, phenytoin Sodium, phenytoin Sodium (dilution), primidone mysoline, hydrochloric acid and replaces and add at least a in guest, topiramate, sodium valproate, the valproic acid.Above-mentioned at least a antidepressants can be and are selected from amitriptyline hydrochloride, amitriptyline embonate, chlorine piperazine oxygen, bupropion hydrochloride, citalopram hydrobromate, Clomipramine Hydrochloride, the hydrochloric acid desipramine, adapin, fluoxetine Hydrochloride, imipramine hydrochloride, imipramine embonate, mirtazapine, nefazodone hydrochloride, psychostyl, paroxetine hydrochloride, W-1544a, sertraline hydrochloride, tranylcypromine sulfate, stangyl, at least a in the wanlafacine hydrochloride.Above-mentioned at least a antianxiety drugs can be and is selected from least a in alprazolam, buspirone hydrochloride, chlordiazepoxide, hydrochloric acid chlordiazepoxide, chlorine nitrogen dipotassium, stable, adapin, hydroxyzine pamoate, hydroxyzine hydrochloride, hydroxyzine pamoate, tavor, mephrobamate, midazolam hydrochloride, the oxazepam.Above-mentioned at least a psychosis can be and is selected from chlorpromazine hydrochloride, clozapine, the capric acid fluphenazine, the enanthic acid fluphenazine, the hydrochloric acid fluphenazine, haloperidol, haloperidol decanoate, the lactic acid haloperidol, the hydrochloric acid loxapine, loxitane, mesoridazine benzenesulfonate, the hydrochloric acid molindone, olanzapine, perphenazine, pimozide, prochlorperazine, quetiapine fumarate, risperidone, mellaril, thiothixene, thiothixene hydrochloride, at least a in the hydrochloric acid trifluoperazine.Above-mentioned at least a central nervous system's stimulant can be and is selected from least a in amphetamine sulfate, caffeine, sulphuric acid dexamphetamine, hydrochloric acid doxapram, methamphetamine hydrochloride, hydrochloric acid methylphenidate, modafinil, pemoline, the phentermine hydrochloride.Above-mentioned at least a antiparkinsonism drug can be and is selected from least a in virofral, methanesulfonic acid benzetropine, biperiden hydrochloride, biperiden lactate, bromocriptine methanesulfonate, carbidopa and levodopa, entacapone, levodopa, pergolide mesylate, two hydrochloric acid pramipexoles, ropinirole hydrochloride, SelegilineHydrochloride, tolcapone, the Cyclodol.Above-mentioned at least a comprehensive medicine for central nervous system can be and is selected from bupropion hydrochloride, donepezil hydrochloride, Droperidol, fluvoxamine maleate, lithium carbonate, Lithium Citrate de, hydrochloric acid and receives at least a in thunder Qu Tan, nicotine polacrilex (a kind of active component of smoking give-up chewing gum), nicotine transdermal system, propofol, Lizakuputan benzoate, Sibutramine hydrochloride, Sumatriptan Succinate, hydrochloric acid tacrine, the Zolmitriptan.(seeing, for example the 337-530 page or leaf among the Nursing 2001Drug Handbook)
Above-mentioned at least a cholinergic agent (for example parasympathomimetic agent) can be and is selected from least a in bethanecholchloride, edrophonium chloride, Neostigmine, neostigmine methylsulfate, physostigmine salicylate, bromination 3-formyl oxygen dimethylamino-1 picoline.Above-mentioned at least a anticholinergic can be and is selected from least a in atropine sulfate, bentrl hydrothloride, robinul, Daturine, sulphuric acid Daturine, probanthine, scopolamine, butyl bromination scopolamine, the hydrogen scotropin.Above-mentioned at least a beta adrenergic agent (sympathomimetic) can be and is selected from least a in dobutamine hydrochloride, dopamine hydrochloride, acid Metaraminol Bitartrate, acid Noradenaline Bitatrate, phenylephrine hydrochloride, pseudoephedrine hydrochloride, the pseudoephedrine sulfate.Above-mentioned at least a adrenergic blocking drug (sympatholytic) can be and is selected from least a in agit, gynergen, desernil, the propranolol hydrochloride.Above-mentioned at least a skeletal muscle relaxant can be and is selected from least a in baclofen, carisoprodol, benzoflex, cyclobenzaprine hydrochloride, dantrolene sodium, methocarbamol, the tizanidine hydrochloride.Above-mentioned at least a neuromuscular blocking agents can be and is selected from least a along in atracurium, doxacurium chloride, mivacurium chloride, pancuronium bromide, pipecuronium bromide, thunder storehouse bromine ammonium, Rocuronium Bromide, succinylcholine chloride, tubocurarine chloride, the vecuronium bromide of benzenesulfonic acid atracurium, benzenesulfonic acid.(seeing, for example the 531-84 page or leaf among the Nuring 2001DrugHandbook).
Above-mentioned at least a hydryllin can be and is selected from least a in brompheniramine maleate, cetirizine hydrochloride, chlorphenamine, fumaric acid clematine, cyproheptadine, diphhydramine hydrochloride, fexofenadine hydrochloride, loratadine, promethazine hydrochloride, promethazine teoclate, the triprolidine hydrochloride.Above-mentioned at least a bronchodilator can be and is selected from least a in salbutamol, Salbutamol, aminophylline, atropine sulfate, ephedrine sulfate, epinephrine, hydrogen tartrate epinephrine, adrenalin hydrochloride, ipratropium bromide, isoproterenol, hydrochloric acid isoproterenol, sulphuric acid isoproterenol, albuterol hydrochloride, metaproterenol sultate, Oxtriphylline, pirbuterol acetate, carbonaphthoic acid salmaterol, sulphuric acid Arubendol, the theophylline.Above-mentioned at least a expectorant or cough medicine can be and be selected from least a in benzonatate dew, codeine phosphate, codeine sulfate, dextromethorphan hydrobromide, diphhydramine hydrochloride, guaifenesin, the dihydro-morphinone hydrochloride.Above-mentioned at least a comprehensive respiratory medications can be and is selected from acetylcysteine, beclomethasone dipropionate, beractant, budesonide, calfactant, sodium cromoglicate, streptodornase α, prostacyclin sodium, 9-and goes at least a in fluorine fluocinonide, fluticasone propionate, Menglusitena, sodium nedocromil, palivizumab, the third scorching pine, zafirlukast, the zileuton.(seeing the 585-642 page or leaf among the Nursing 2001Drug Handbook)
Above-mentioned at least a antacid, adsorbent or antiflatulent can be and be selected from least a in aluminium carbonate, aluminium hydroxide, calcium carbonate, magaldrate, magnesium hydroxide, magnesium oxide, dimethicone, the sodium bicarbonate.Above-mentioned at least a digestive enzyme or cholelithiasis solubilizing agent can be and be selected from least a in pancreatin, pancreatic lipase, the ursodeoxycholic acid.Above-mentioned at least a diarrhea can be and is selected from least a in Attagel, basic bismuth salicylate, WL-140, diphenoxylate hydrochloride or atropine sulfate, loperamide, octreotide acetate, tinctura opii, the tinctura opii (containing Camphora).Above-mentioned at least a antidiabetic drug or glucagon can be and be selected from least a in acarbose, chlorpropamide, glutathion, glipizide, glucagon, glyburide, insulin, hydrochloric acid metformin, miglitol, pyrrolidine hydrochloride row ketone, repaglinide, rosiglitazone maleate, the troglitazone.Above-mentioned at least a thyroxin can be and is selected from least a in levothyroxine sodium, Cynomel, liotrix, the dried thyroid.Above-mentioned at least a thyroxin antagonist can be and is selected from the thin imidazoles of first, potassium iodide, potassium iodide (saturated solution), propylthiouracil (PTU), radioiodine (sodium iodide 131I), at least a in the Lugol's solution.Above-mentioned at least a pituitary hormone can be and is selected from least a in thyroliberin, cosyntropin, desmophressin acetate, the bright dried meat Li Te of acetic acid, long-acting thyroliberin, somatrem, growth hormone, the vassopressin.Above-mentioned at least one kind parathyroid gland medicine can be and is selected from least a in calcifediol, calcitonin (mankind), calcitonin (salmon), calitriol, dihydrotachysterol, the etidronate.(seeing, for example the 696-796 page or leaf among the Nursing 20001Drug Handbook)
Above-mentioned at least a diuretic can be and is selected from least a in acetyl azoles (sulphur) amine, acetyl azoles (sulphur) amine sodium, hydrochloric acid amiloride, bumetanide, chlortalidone, ethacrynate sodium, acidum ethacrynicum, furosemide, Hydrochlorothiazide, indapamide, mannitol, Mei Latuo ancestor, spironolactone, torasemide, traimterene, the urea.Above-mentioned at least a electrolyte or substitutional solution can be and be selected from calcium acetate, calcium carbonate, calcium chloride, calcium citrate, calcium glubionate, calcium glucoheptonate, calcium gluconate, calcium lactate, calcium phosphate (binary), calcium phosphate (ternary), glucosan (high molecular), dextran (low-molecular-weight), hetastarch, magnesium chloride, magnesium sulfate, potassium acetate, potassium bicarbonate, potassium chloride, potassium gluconate, the RingerShi injection, RingersShi injection (lactic acidization), at least a in the sodium chloride.Above-mentioned at least a acidulant or basifier can be and be selected from least a in sodium bicarbonate, sodium lactate, the tromethane.(seeing, for example the 797-833 page or leaf among the Nursing 2001Drug Handbook)
Above-mentioned at least a hematonic can be and is selected from least a in ferrous fumarate, ferrous gluconate, ferrous sulfate, ferrous sulfate (drying), iron dextran, Iron Sorbitex, polysaccharide-iron complex, the Ferrlecit.Above-mentioned at least a anticoagulant can be and is selected from Ardeparin Sodium., at least a in the dalteparin sodium, Danaparoid sodium, Enoxaparin Sodium, calciparine, heparin sodium, tintorane.Above-mentioned at least a blood derivatives can be and is selected from least a in albumin 5%, albumin 25%, antihemophilic factor, counter inhibitor coagulant complex, Antithrombin III (mankind), the IX factor (mankind), IX factor complex, the plasma protein fraction.Above-mentioned at least a thrombolytic can be and is selected from least a in alteplase, Ah Buddhist nun's chain enzyme, reteplase (reorganization), streptokinase, the urokinase.(seeing, for example the 834-66 page or leaf among the Nursing2001Drug Handbook)
Above-mentioned at least a alkylation medicine can be and is selected from least a in busulfan, carboplatin, carmustine, chlorambucil, cisplatin, cyclophosphamide, ifosfamide, lomustine, mustine hydrochlcride, melphalan, hydrochloric acid melphalan, streptozotocin, temozolomide, the thiotepa.Above-mentioned at least a antimetabolite can be and is selected from least a in capecitabine, cladribine, cytosine arabinoside, fluorine glycosides, fludarabine phosphate, fluorouracil, hydroxyurea, purinethol, methotrexate, methotrexate sodium, the thioguanine.Above-mentioned at least a antibiotic antitumor drug can be and is selected from least a among bleomycin sulfate, actinomycin D, citric acid daunorubicin liposome, daunorubicin hydrochloride, doxorubicin hydrochloride, hydrochloric doxorubicin liposome, epirubicin hydrochloride, idarubicin hydrochloride, mitomycin, pentoside, plicamycin, the valrubicin.The above-mentioned at least a antitumor drug that changes hormonal balance can be and is selected from least a in Anastrozole, bicalutamide, estramustine phosphate sodium, exemestane, flutamide, acetic acid goserelin, letrozole, the bright dried meat Li Te of acetic acid, megestrol acetate, nilutamide, TAMOXIFEN CITRATE, testolactone, the Toremifene Citrate.Above-mentioned at least a comprehensive antitumor drug can be and is selected from asparaginase, bacillus Calmette-Guerin (BCG) (live body intravesical), dacarbazine, Docetaxel, etoposide, the phosphoric acid etoposide, gemcitabine hydrochloride, irinotecan hydrochloride, mitotane, mitoxantrone hydrochloride, paclitaxel, Pegaspargase, porfimer sodium, hydrazine under the hydrochloride methyl, Rituximab, teniposide, topotecan hydrochloride, Herceptin, retin-A, Vinblastine Sulfate, the sulphuric acid vincristin, Vinorelbine tartrate, at least a.(seeing, for example the 867-963 page or leaf among the Nursing 2001Drug Handbook)
Above-mentioned at least a immunosuppressant can be and is selected from least a in azathioprine, basiliximab, ciclosporin, daclizumab, lymphocyte immune globulin, muromonab-CD3, mycophenolic acid morpholine ethyl ester, hydrochloric acid mycophenolic acid morpholine ethyl ester, sirolimus, the tacrolimus.Above-mentioned at least a vaccine or toxoid can be and be selected from BCG vaccine, cholera vaccine, diphtheria and tetanus toxoid (absorbent-type), diphtheria and tetanus toxoid and acellular pertussis vaccine absorbent-type, diphtheria and tetanus toxoid and whole cell pertussis vaccine, b type haemophilus combined vaccine, Hepatitis A Vaccine (deactivation), hepatitis B vaccine (reorganization), influenza virus vaccine 1999-2000 triple type A﹠amp; B (surface antigen of purification), influenza virus vaccine 1999-2000 triple type A﹠amp; B (the subvirus body of subvirus body or purification), influenza virus vaccine 1999-2000 triple type A﹠amp; B (totivirus body), Japanese encephalitis virus vaccine (deactivation), Lyme borrelia burgdorferi disease vaccine (reorganization OspA), measles and parotitis and rubella virus vaccine (work), measles and parotitis and rubella virus vaccine (deactivation), measles virus vaccines (deactivation), the meningococcus polysaccharide vaccine, mumps virus vaccine (work), pestilence vaccine, Pnu-Imune 23 (multi-joint), poliovirus vaccine (deactivation), poliovirus vaccine (is lived, oral, three), rabies vaccine (absorbent-type), rabies vaccine (human diploid cell), rubella and mumps virus vaccine (work), rubella virus vaccine (is lived, deactivation), tetanus toxoid (absorbent-type), tetanus toxoid (liquid), antityphoid vaccine (oral), antityphoid vaccine (parenteral administration), Typhoid Vi Polysaccharide Vaccine, Varivax, at least a in the yellow fever vaccine.Above-mentioned at least a antitoxin or venom can be and be selected from latrodectus mactans's venom, Crotalidae venom (multi-joint), diphtheria antitoxin (horse), golden yellow Corallium Japonicum Kishinouye calmette's serum) at least a.Above-mentioned at least a immune serum can be and is selected from cytomegalovirus immune globulin (intravenous), hepatitis B immune globulin (mankind), the immunoglobulin intramuscular, the immunoglobulin intravenous, rabies immune globulin (mankind), respiratory syncytial virus immunoglobulin intravenous (mankind), Rho (D) immunoglobulin (mankind), Rho (D) immunoglobulin intravenous (mankind), tetanus immune globulin (mankind), at least a in the varicella-zoster immunoglobulin.Above-mentioned at least a biological respinse modifier can be and is selected from Ah Di Liujin, at least a according in pool Ai Ting α, Fei Lasiting, injection glatiramer acetate, interferon alfacon-1, Intederon Alpha-2a (reorganization), Interferon Alpha-2b (reorganization), interferon beta-1a, interferon beta-1b (reorganization), gamma interferon 1-b, levamisole hydrochloride, rHuIL-11, the Sargramostim.(seeing, for example the 964-1040 page or leaf among the Nursing 2001Drug Handbook)
Above-mentioned at least a ophthalmology anti-infective can be selected from least a in bacitracin, chloromycetin, ciprofloxacin, erythromycin, sulfate gentamicin, pyridine carboxylic acid 0.3%, aerosporin, sulphacetamide 10%, sulphacetamide 15%, sulphacetamide 30%, tobramycin, the vidarabine.Above-mentioned at least a ophthalmology anti-inflammatory agent can be and is selected from least a in dexamethasone, dexamethasone sodium phosphate, diclofenac sodium 0.1%, chloromethane dragon, flurbiprofen sodium, ketorolac tromethamine, predniso lone acetate (suspension), the prednisolone phosphate sodium (solution).Above-mentioned at least a miotic can be and is selected from least a in acetylcholine chloride, carbachol (ophthalmic), carbachol (part), ecostigmine, pilocarpine, pilocarpine hydrochloride, the pilocarpine nitrate.Above-mentioned at least a iridodilator can be and is selected from least a in atropine sulfate, cyclogyl hydrochloride, adrenalin hydrochloride, epinephrine borate, homatropine hydrobromide, phenylephrine hydrochloride, scopolamine hydrobromide, the N-ethyl-N-(.gamma.-picolyl)tropamide.Above-mentioned at least a eye vasoconstrictor can be and is selected from least a in naphazoline hydrochloride, oxymetazoline hydrochloride, the Visine.Above-mentioned at least a comprehensive medicament for the eyes can be and is selected from hydrochloric acid apraclonidine, betaxolol hydrochloride, brimonidine tartrate, hydrochloric acid galutedin, dipivefrine hydrochloride, hydrochloric acid and stops up beautiful at least a in Bu Lage, sodium chloride (height oozes), the timolol maleate of amide, fumaric acid emedastine, fluorescein sodium, Fumaric acid ketotifen, latanoprost, Levobunolol Hydrochorid, hydrochloric acid.Above-mentioned at least a ear medication can be and is selected from least a in boric acid, carbamide peroxide, chloromycetin, the triethanolamine oleate polypeptide-condensed fluid.Above-mentioned at least a nose medication can be and is selected from beclomethasone dipropionate, budesonide, ephedrine sulfate, adrenalin hydrochloride, 9-and goes at least a in fluorine fluocinonide, fluticasone propionate, naphazoline hydrochloride, oxymetazoline hydrochloride, phenylephrine hydrochloride, Visine, the third scorching pine, the xylometazoline hydrochloride.(seeing, for example the 1041-97 page or leaf among the Nursing 2001Drug Handbook)
Above-mentioned at least a local anti-infective agent can be and is selected from acycloguanosine, amphotericin B, Azelaic Acid butterfat, bacitracin, Nitric acid butoconazole, the p chloromethylbenzoic acid cillimycin, clotrimazole, econazole nitrate, erythromycin, sulfate gentamicin, ketoconazole, the acetic acid marfanil, metronidazole (part), the mould anti-azoles of nitric acid, mupirocin, naftifine hydrochloride, polygynax, nitro sugar hydrazone, nystatin, silver sulfadiazine, the special Binet phenol of hydrochloric acid, terconazole (triaconazole), quadracycline, tioconazole, at least a during tineatonsurans is moved back.Above-mentioned at least a antiscabietic or antipedicular agent can be and be selected from least a in crotamiton, gamma hch, permethrin, the pyrethrin.Above-mentioned at least a topical corticosteroid can be selected from two betamethasone dipropionates, betamethasone valerate, CBP, desonide, desoximetasone, dexamethasone, dexamethasone sodium phosphate. at least a in diflorasone diacetate, acetone fluocinonide, fluocinolone ester, flurandrenolide, fluticasone propionate, halcionide, hydrocortisone, hydrocortisone acetate, hydrocortisone butyrate, valeric acid hydrocortisone, furancarboxylic acid Mo Meitasong, the third scorching pine.(seeing, for example the 1098-1136 page or leaf among the Nursing2001Drug Handbook)
Above-mentioned at least a microorganism or mineral can be and be selected from vitamin A, compound vitamin B, cyanocobalamin, folic acid, hydroxocobalamine, calcium leucovorin, nicotinic acid, nicotiamide, pyridoxine hydrochloride, riboflavin, thiamine hydrochloride, vitamin C, vitamin D, cholecalciferol, ergocalciferol, novel vitamin D analogues, the degree ostelin, the sharp ostelin of handkerchief, vitamin E, the vitamin K analog, vitamin K1, sodium fluoride, sodium fluoride (part), trace element, chromium, copper, iodine, manganese, selenium, at least a in the zinc.Above-mentioned at least a caloic can be the aminoacid preserved material that is selected from aminoacid preserved material (crystalline), the dextrose, contain contain electrolytical aminoacid preserved material in electrolytical aminoacid preserved material, the dextrose, at the aminoacid preserved material of liver failure, at least a in the aminoacid preserved material of hypermetabolism pressure, the aminoacid preserved material, dextrose, fat emulsion, MCT Oil at renal failure.(seeing, for example the 1137-63 page or leaf among the Nursing 2001DrugHandbook)
CH1 of the present invention disappearance analogue body antibody or peptide composition can further comprise at least a in any particular composition suitable and/or effective dose or the pharmaceutical composition, this particular composition or pharmaceutical composition have comprised the above-mentioned adjusting of needs, the cell of handling or treating, tissue, organ, at least a CH1 disappearance analogue body albumen or antibody that animal or patient use, this particular composition or pharmaceutical composition have randomly further comprised and (for example have been selected from least a TNF antagonist, but be not limited only to TNF chemistry or protein antagonist, TNF monoclonal or polyclonal antibody or fragment, solvable TNF receptor (p55 for example, p70 or p85) or fragment, its fused polypeptide, or micromolecule TNF antagonist, for example conjugated protein I of TNF or II (TBP-1 or TBP-II), nerelimonmab, English monoclonal antibody of sharp former times, enteracept, CDP-571, CDP-870, Afelimomab, Lenercept. etc.), antirheumatic (for example, methotrexate, AF, aurothioglucose, azathioprine, Embrel, Kidon (Ono), sulphuric acid hydroxyl chloroquine, leflunomide, sulfasalazine), muscle relaxant, anesthetics, on-steroidal anti-inflammatory medicaments (NSAID), analgesic, anesthetis, tranquilizer, local anesthetic, neuromuscular blocking agents, antibacterial (aminoglycoside for example, antifungal, antiparasitic, antiviral drugs, carbamyl, cephalosporin, fluoroquinolone, macrolide, penicillin, sulfanilamide, tetracycline, other antibacterial), antipsoriatic, corticosteroid, anabolic steroid, the diabetes related drugs, mineral, nutrient substance, the thyroid medicine, vitamin, the calcium associated hormone, diarrhea, cough medicine, antiemetic, antiulcerative, aperient, anticoagulant, erythropoietin (for example according to pool Ai Ting α), Fei Lasiting (G-CSF for example, Neupogen), Sargramostim (GM-CSF, Leukine), immunity, immunoglobulin, immunosuppressant (basiliximab for example, cyclocyto polypeptide, daclizumab), growth hormone, hormone replaces medicine, the estrogen receptor instrumentality, iridodilator, cycloplegic, the alkylation medicine, antimetabolite, mitotic inhibitor, radiopharmaceutical, antidepressants, antimaniacal drugs, psychosis, antianxiety drugs, hypnotic, sympathomimetic, analeptic, donepezil, tacrine, asthma drug, the β synergist, the induction type steroid, the leukotriene inhibitor, methylxanthine, cromoglicic acid, epinephrine or analog, streptodornase α (Pulmozyme), at least a in cytokine or the cytokine antagonist.The limiting examples of above-mentioned cytokine include but not limited to any one among the IL-1-IL-23.Suitable dose is well known in the art.See Wells etal. for example, eds., Pharmacotherapy Handbook, 2 NdEdition, Appleton and Lange, Stamford, CT (2000); PDR Pharmacopoeia, Tarascon Pocket Pharmacopoeia 2000, Deluxe Edition, Tarascon Publishing, LomaLinda, CA (2000), above-mentioned list of references all herein by complete introducing, as a reference.
Above-mentioned composition also can comprise with at least a antibody of the present invention or polypeptide associate, combine, collaborative preparation or the collaborative lps molecule of using.This toxin can randomly play selectivity and kill pathological cells or tissue.This pathological cells can be cancerous cell or other cell.This toxin can be, but purification or the recombinant toxin or the toxin fragment at least one functional cell toxin district of particular toxin have been not limited only to comprise, this particular toxin for example is selected from, one of at least a ricin, diphtheria toxin, diphtherotoxin, venom toxin or bacteriotoxin.The term toxin also comprises by any can cause the pathological condition that comprises toxic shock in human and other mammal, and the endotoxin and the extracellular toxin that can cause dead natural existence, sudden change or recombinant bacteria or virus to be generated.This toxin can include but not limited to enterotoxigenic heat-labile enterotoxin of E, coli (LT), thermally-stabilised enterotoxin (ST), shigella dysenteriae cytotoxin, Aeromonas enterotoxin, toxic shock syndrome toxin-1 (TSST-1), staphylococcal enterotoxin A (SEA), B (SEB) or C (SEC), streptococcus enterotoxin etc.Above-mentioned antibacterial comprises, but be not limited only to enterotoxigenic escherichia coli (ETEC), cause enterorrhagia escherichia coli (for example bacterial strain of serotype 0157:H7), staphylococcus kind (staphylococcus aureus for example, staphylococcus pyogenes), shigella dysenteriae kind (dysentery bacterium for example, the Fu Shi bacillus dysenteriae, Shigella boydii and bacterium sonnei), salmonella strain (Bacillus typhi for example, Salmonella choleraesuls, Salmonella enteritidis), clostruidium kind (clostridium perfringens for example, difficult bacillus, bacillus botulinus), bent stick strain (campylobacter jejuni for example, embryo's Campylobacter), screw rod strain (for example helicobacter pylori), Aeromonas kind (for example, Aeromonas sobria, Aeromonas hydrophila, Aeromonas caviae), Plesiomonas shigelloides, Yersinia enterocolitica, vibrio kind (vibrio cholera for example, vibrio parahaemolyticus), the klebsiella bacillus kind, Pseudomonas aeruginosa and streptococcus.See Stein for example, ed., INTERNAL MEDICINE, 3 RdEd., pp1-13, Little, Brown and Co., Boston, (1990); Evanset al., eds., Bacterial Infections of Humans; Epidemiology andControl, 2d.Ed., pp 239-254, Plenum Medical Book Co., New York (1991); Mandell et al., Principles and Practice of InfectiousDiseases, 3d.Ed., Churchill Livingstone, New York (1990); Berkow et al., eds., The Merck Manual, 16 ThEdition, Merck andCo., Rahway, N.J., 1992; Wood et al., FEMS MicrobiologyImmunology, 76:121-134 (1991); Marrack et al., Science, 248:705-711 (1990), the content of above-mentioned list of references is all by complete introducing herein as a reference.
EPO mimetic CH 1 deleted mimetibodies of the present invention or specific part or variant compositions can further comprise at least a suitable arbitrarily adminicle, for example, but be not limited only to diluent, binding agent, stabilizing agent, buffer agent, salt, lipophilic solvent, antiseptic, adjuvant etc.It is preferred that materia medica can be accepted adminicle.The limiting examples of above-mentioned sterile solution and preparation method all are well known in the art, for example, but are not limited only to Gennaro, Ed., Remington ' sPharmaceutical Sciences, 18 ThEdition, Mack Publishing Co. (Eas ton, PA) 1990.Can select the pharmacological-acceptable carrier that matches with the method for application of EPO mimetic CH 1 deleted mimetibodies compositions, dissolubility and/or stability by well known and/or described herein conventional method.
Drug excipient and additive useful in the present composition comprise, but be not limited only to and exist separately or with combining form, comprise separately or the protein, peptide, aminoacid, lipid and the saccharide that in combination, exist with 1-99.99% weight or volume ratio (for example, sugar, comprise monosaccharide, disaccharide, trisaccharide, tetrose and oligosaccharide; Derived carbohydrate is such as sugar alcohol, alduronic acid, esterified saccharides etc.; Polysaccharide or carbohydrate polymer).The example of protein excipient comprises serum albumin, such as human serum albumin (HSA), recombinant human albumin (rHA), gelatin, casein etc.Also can bring into play the representative aminoacid/EPO mimetic CH 1 deleted mimetibodies of cushioning effect or specific part or variant component and comprise alanine, glycine, arginine, betanin, histidine, glutamic acid, aspartic acid, cysteine, lysine, leucine, isoleucine, valine, methionine, phenylalanine, aspartame etc.A kind of preferred amino acids is a glycine.
Be applicable to that saccharide excipient of the present invention comprises, for example, monosaccharide is such as fructose, maltose, galactose, glucose, D-mannose, sorbose etc.; Disaccharide is such as lactose, sucrose, trehalose, cellobiose etc.; Polysaccharide is such as Raffinose, melezitose, maltodextrin, glucosan, starch etc.; And sugar alcohol, such as mannitol, xylitol, maltose alcohol, lactose, xylitol, Sorbitol (glucitol), inositol etc.Be applicable to that preferred saccharide excipient of the present invention is mannitol, trehalose and Raffinose.
EPO mimetic CH 1 deleted mimetibodies compositions also can comprise buffer agent or pH regulator agent; Typical buffer agent be from organic acid or alkali preparation and salt.Representative buffer agent comprises acylate, such as the salt of citric acid, ascorbic acid, gluconic acid, carbonic acid, tartaric acid, succinic acid, acetic acid or phthalic acid; Tris, hydrochloric acid Tris or phosphate buffer.The preferred reducing that is applicable to the present composition is an acylate, such as citrate.
In addition, EPO mimetic CH 1 deleted mimetibodies of the present invention or specific part or variant compositions can comprise polymeric excipient/additive, such as polyvinylpyrrolidone, ficolls (a kind of polymerization sugar), dextrates (cyclodextrin for example, such as 2-hydroxy propyl-Beta cyclodextrin), Polyethylene Glycol, flavoring agent, antibacterial, sweeting agent, antioxidant, antistatic additive, surfactant (for example polysorbate, such as " TWEEN 20 " and " TWEEN 80 "), lipid (phospholipid for example, fatty acid), steroid (for example cholesterol) and chelating agen (for example EDTA).
Be applicable to that the above-mentioned of EPO mimetic CH 1 deleted mimetibodies compositions of the present invention and other known drug excipient and/or additive all are known in the art, for example at " Remington:TheScience﹠amp; Practice of Pharmacy ", 19 ThEd., Williams﹠amp; Williams, (1995) and in " Physician ' s Desk Reference ", 52 NdEd., MedicalEconomics, Montvale, listed drug excipient and/or additive among the NJ (1998), the disclosure of above-mentioned list of references is all by complete introducing herein as a reference.Preferred carrier or excipient materials are saccharide (for example saccharide and sugar alcohol) and buffer agent (for example citrate) or polymerizer.
Prescription
As mentioned above, the invention provides stabilized formulations, this prescription can preferably include the suitable buffer agent of brackish water or specific salts, with the optional antiseptic that contains, and the listerine and the prescription that are fit to the anticorrosion prescription of multipurpose of medicinal or veterinary purpose, and comprised at least a EPO mimetic CH 1 deleted mimetibodies or specific part or the variant that materia medica can be accepted the form of filling a prescription.Anticorrosion prescription contains at least a known antiseptic, or be selected from following optional antiseptic, be at least a phenol, m-cresol, p-cresol, o-cresol, chlorocresol, benzyl alcohol, nitrous acid benzene hydrargyrum, phenoxyethanol, formaldehyde, ammonia butanols, magnesium chloride (for example hexahydrate), alkyl P-hydroxybenzoic acid (methyl, ethyl, propyl group, butyl etc.), alkyldimethylbenzylammonium chloride, benzethonium chloride, dehydro sodium acetate and thimerosal, or the mixture of above-mentioned substance in aqueous diluent.Can be according to the concentration or the mixture of knowledge application of any suitable known in the art, such as 0.001-5%, or any range wherein or numerical value, for example, but be not limited only to 0.001,0.003,0.005,0.009,0.01,0.02,0.03,0.05,0.09,0.1,0.2,0.3,0.4,0.5,0.6,0.7,0.8,0.9,1.0,1.1,1.2,1.3,1.4,1.5,1.6,1.7,1.8,1.9,2.0,2.1,2.2,2.3,2.4,2.5,2.6,2.7,2.8,2.9,3.0,3.1,3.2,3.3,3.4,3.5,3.6,3.7,3.8,3.9,4.0,4.3,4.5,4.6,4.7,4.8,4.9 or any range wherein or numerical value.Limiting examples comprises, non-representational 0.1-2%m-cresol (for example 0.2,0.3,0.4,0.5,0.9,1.0%), 0.1-3% benzyl alcohol (for example 0.5,0.9,1.1,1.5,1.9,2.0,2.5%), 0.001-0.5% thimerosal (for example 0.005,0.01%), 0.001-2.0% phenol (for example 0.05,0.25,0.28,0.5,0.9,1.0%), 0.0005-1.0% alkyl P-hydroxybenzoic acid (for example 0.00075,0.0009,0.001,0.002,0.005,0.0075,0.009,0.01,0.02,0.05,0.075,0.09,0.1,0.2,0.3,0.5,0.75,0.9,1.0%) etc.
As mentioned above, the invention provides a kind of goods, these goods comprise that packaging material and at least one contain the bottle of particular solution, this particular solution contains at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant and above-mentioned buffer agent and/or antiseptic, and randomly be the aqueous diluents form, wherein above-mentioned packaging material comprise that having indicated above-mentioned solution can preserve and surpassed 1,2,3,4,5,6,9,12,18,20,24,30,36,40,48,54,60,66,72 hour or the label of longer time section.The present invention further comprises a kind of goods, these goods comprise packaging material, contain first bottle of at least a lyophilizing EPO mimetic CH 1 deleted mimetibodies or specific part or variant, second bottle with the aqueous diluents that contains above-mentioned buffer agent or antiseptic, wherein above-mentioned packaging material comprise a label, can how at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant be sneaked in the aqueous diluents again to patient explanation, thereby and form to preserve and surpassed 24 hours or the solution of longer time section.
At least a EPO mimetic CH 1 deleted mimetibodies of using according to the present invention or specific part or variant can be by the reorganization approach, comprise from mammalian cell or transgenic goods preparing that perhaps from then on the place is described or other biological origin purification acquisition known in the art.
In product of the present invention, the quantitative range of at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant is included under the situation in wet/dry system, restoring the amount that can obtain the about 1000mg/ml concentration of about 1.0ug/ml-on the basis, but low and higher concentration also is feasible, and depend on specific delivery vectors, for example solution formula is different from percutaneous medicine card, lung, strides mucosa, or infiltration or micro pump method.
Preferably, above-mentioned aqueous diluents comprises further that randomly materia medica can accept antiseptic.Preferred antiseptic comprises and is selected from following antiseptic, be phenol, m-cresol, p-cresol, o-cresol, chlorocresol, benzyl alcohol, alkyl P-hydroxybenzoic acid (methyl, ethyl, propyl group, butyl etc.), alkyldimethylbenzylammonium chloride, benzethonium chloride, dehydro sodium acetate and thimerosal, or the mixture of above-mentioned substance.The concentration of preservatives that is applied in the present invention's prescription is the concentration that is enough to produce anti-microbial effect.This concentration depends on selected antiseptic, and the technical staff can easily measure this concentration.
Other excipient, for example isotonic agent, buffer agent, antioxidant, preservative enhancers all can be chosen wantonly and preferably be added in the above-mentioned diluent.Isotonic agent as glycerol, adopts its concentration known usually.Physiology can tolerate buffer agent and preferably be added, to improve pH control.The present invention's prescription can cover large-scale pH, and such as the about pH10 of about pH4-, preferred range is the about pH9 of about pH5-, and highly preferred scope is the about pH8.0 of about pH6.0-.The preferred pH value of the present invention's prescription is approximately between the 6.8-about 7.8.Preferred reducing agents comprises phosphate buffer, most preferably sodium phosphate, especially phosphate buffered saline(PBS) (PBS).
Other additive, can accept solubilizing agent such as materia medica, as Tween 20 (polyethylene glycol oxide (20) Span-20), Tween 40 (polyethylene glycol oxide (20) sorbitan monopalmitate), Tween 80 (polyethylene glycol oxide (20) sorbitan monooleate), Fluronic F68 (polyethylene glycol oxide polyoxypropylene block copolymers) and PEG (Polyethylene Glycol) or non-ionic surface active agent, such as polysorbate20 or 80 or poloxalkol 184 or 188, Pluronic  polyls, other block copolymer, with chelating agen such as EDTA and EGTA, all can randomly be added in prescription of the present invention or the compositions, to reduce cohesion.When adopting pump or plastic containers to use the present invention when filling a prescription, above-mentioned additive is especially effective.The tendency of protein condenses has been lowered in the existence that materia medica can be accepted surfactant.
The present invention's prescription can prepare by ad hoc approach, this method comprise with at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant be selected from one of following antiseptic and mix, be phenol, m-cresol, p-cresol, o-cresol, chlorocresol, benzyl alcohol, alkyl P-hydroxybenzoic acid (methyl, ethyl, propyl group, butyl etc.), alkyldimethylbenzylammonium chloride, benzethonium chloride, dehydro sodium acetate and thimerosal, or the mixture of above-mentioned substance in aqueous diluents.At least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant and antiseptic are blended in the aqueous diluents, are to adopt conventional dissolving and married operation realization.For preparing suitable prescription, for example, at least a EPO mimetic CH 1 deleted mimetibodies or the appointment antiseptic in specific part or variant and the buffer accurately measured in the buffer are pressed the particular magnitude compatibility, and this particular magnitude is enough to provide the prescribed concentration of protein and antiseptic.Any those of ordinary skill in this area all should be approved the change form of this method.For example, add the order of component, whether adopt other additive, prepare the temperature and the pH of this prescription, thus be can optimisedly be applicable to whole factors of the concentration that adopts and route of administration.
The prescription of institute of the present invention prescription can be used as clear liquid or two doleiform formula is provided for the patient, wherein in two bottles, there is one bottle to contain at least a freeze dried EPO mimetic CH 1 deleted mimetibodies or specific part or variant, can utilize and contain water, antiseptic and/or excipient, preferably phosphoric acid salt buffer agent and/or saline and specific salts, and another bottle of formation aqueous diluents restores it.Need restorative single solution bottle or two bottle all can be repeated to utilize repeatedly, and be enough to satisfy single or many circulation treatments needs, thereby can provide than existing general treatment scheme Therapeutic Method more easily to the patient.
The goods of institute of the present invention prescription are applicable to from using to 24 hours or longer time section span immediately.Correspondingly, the goods of institute of the present invention prescription provide significant benefits for the patient.The present invention's prescription is safe storage under the about 40 ℃ temperature of about 2-randomly, and keep proteinic biological activity for a long time, thereby can on 6,12,18,24,36,48,72 or 96 hours or longer time period span, be saved and/or use indicating this solution on the packaging label.As adopt anticorrosion diluent, this label can comprise operating period until at least one month in 1-12 the middle of the month number, half a year, a year and a half and/or 2 years.
In the present invention, the solution of at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant can prepare by ad hoc approach, and this method comprises at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant are blended in the aqueous diluents.This mixing is to adopt conventional dissolving and married operation to realize.For preparing suitable diluent, for example, with in water or buffer, with at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant that the particular magnitude compatibility is accurately measured, this value is enough to provide the prescribed concentration of protein and optional antiseptic or buffer agent.Any those of ordinary skill in this area all should be approved the change form of this method.For example, add the order of component, whether adopt other additive, the temperature and the pH of preparation the present invention prescription, thereby be can optimisedly be applicable to whole factors of the concentration that adopts and route of administration.
The product of institute of the present invention prescription can be used as clear liquid or two doleiform formula is provided for the patient, wherein in two bottles, there is one bottle to contain at least a freeze dried EPO mimetic CH 1 deleted mimetibodies or specific part or variant, can utilizes another bottle of property of water-bearing diluent that it is restored.Need restorative single solution bottle or two bottle all can be repeated to utilize repeatedly, and be enough to satisfy single or many circulation treatments needs, thereby can provide than existing general treatment scheme Therapeutic Method more easily to the patient.
By providing the clear liquid of institute of the present invention prescription or the product of two doleiform formulas to pharmacy, clinic or other similar means and facility, it can be offered the patient indirectly, wherein in two bottles, there is one bottle to contain at least a freeze dried EPO mimetic CH 1 deleted mimetibodies or specific part or variant, can utilizes another bottle of property of water-bearing diluent that it is restored.Clear liquid in this situation can reach one liter or more volume, thereby need provide a kind of big reservoir, by its one or more times will more a spot of at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant solution shift in the into less bottle, this reservoir can be provided to its client and/or patient by pharmacy or clinic.
The generally acknowledged device that comprises above-mentioned single bottle system comprises and is used to send pen type-injection device of passing solution, such as Huma ject , NovaPen , B-D Pen, AutoPen And OptiPen The generally acknowledged device that comprises two bottles system comprises and can be used for freeze-dried drug is restored in cartridge case, and send pen type-injection device of passing this reconstituted solution, such as HumatroPen
The product of institute of the present invention prescription comprises packaging material.Except that the desired information of administrative organization, these packaging material also provide the condition of using this product.Packaging material of the present invention provide description to the patient, told about at product and be two bottles, wet/during dry form, restore at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant in the aqueous diluent, form the method for solution, and surpassed 2-24 hour or the span of longer time section on use the method for this solution.For single bottle of solution product, labeled marker this solution can be used above 2-24 hour or longer time section.The product of institute of the present invention prescription is useful aspect the human medicine product purpose.
The present invention's prescription can prepare by ad hoc approach, and this method comprises that with at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant and specific buffers the phosphate buffer of preferred brackish water or specific salts is mixed.At least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant and buffer agent are blended in the aqueous diluents, are to adopt conventional dissolving and married operation realization.For preparing suitable prescription, for example, at least a EPO mimetic CH 1 deleted mimetibodies or the appointment buffer agent in specific part or variant and the water accurately measured in water or the buffer agent are pressed the particular magnitude compatibility, this value is enough to provide the prescribed concentration of protein and buffer agent.Any those of ordinary skill in this area all should be approved the change form of this method.For example, add the order of component, whether adopt other additive, the temperature and the pH of preparation the present invention prescription, thereby be can optimisedly be applicable to whole factors of the concentration that adopts and route of administration.
Stable or the anticorrosion prescription of institute of the present invention prescription can clear liquid or two doleiform formula be provided for the patient, in this pair bottle, there is one bottle to contain at least a freeze dried EPO mimetic CH 1 deleted mimetibodies or specific part or variant, can utilizes the antiseptic of property of water-bearing diluent form or another bottle of buffer agent and excipient that it is restored.Need restorative single solution bottle or two bottle all can be repeated to utilize repeatedly, and be enough to satisfy single or many circulation treatments needs, thereby can provide than existing general treatment scheme Therapeutic Method more easily to the patient.
According to the present invention, at least a EPO mimetic CH 1 deleted mimetibodies in stable or anticorrosion prescription or the solution described herein or specific part or variant can be applied to the patient by the multiple method of passing of sending, and comprise SC or IM injection; Percutaneous, lung, stride mucosa, transplanting, osmotic pumps, cartridge case, micro pump or well known, and other approach of being understood by those of skill in the art.
Treatment is used
With regard to analogue body, the present invention also provides and has regulated or the treatment cell, tissue, organ, the method of animal or patient's anemia, this anemia comprises, but be not limited only at least a in any anemia, the treatment of cancer anemia of being correlated with, radiotherapy or the chemotherapy anemia of being correlated with, virus or the relevant anemia of bacterial infection treatment, renal anemia, precocious relevant anemia, department of pediatrics and/or the relevant anemia of one-tenth human cancer, with lymphoma, myeloma, the anemia that multiple myeloma is relevant, the AIDS anemia of being correlated with, this method comprises follows treatment to what the patient with following situation carried out, promptly there is or do not exist the patient who contributes the inferior time elective surgery of blood situation from body, before for the operation of operation and after the operation, accepted to contribute blood or blood transfusion from body, periodontitis treatment, suffer from cyclic neutropenia or Kostmann syndrome (congenital agranulocytosis), latter stage nephropathy, the anemia relevant with dialysis, chronic renal insufficiency, constitutional hemopoietic disease is such as congenital hypoplastic anemia, major thalaseemia or sickle cell disease, the patient of the vascular occlusion complication of sickle cell disease.Furman et al., Pediatrics 1992; 90:716-728, Goldberg Science.1988; 242:1412-1415; Paul et al., Exp Hematol.1984; 12:825-830; Erslev et al., Arch Intern Med.1968; 122:230-235; Ersley et al., Ann Clin Lab Sci.1980; 10:250-257; Jacobs et al., Nature.1985; 313:806-810; Linet al., Proc Natl Acad Sci USA.1985; 82:7580-7584; Law et al., Proc Natl Acad Sci USA.1986; 83:6920-6924; Goldwasser et al., J Biol Chem.1974; 249:4202-4206; Eaves et al., Blood.1978; 52:1196-1210; Sawyer et al., Blood.1989; 74:103-109; Winearls et al., Lancet.1986; 2:1175-1178; Eschbach et al., N Engl J Med.1987; 316:73-78; Eschbach et al., Ann Intern Med.1989; 111:992-1000, above-mentioned list of references are all by complete introducing herein as a reference.
Analogue body of the present invention also can be applied to by for example, the anemia of the non-kidney form that chronic infectious disease, inflammatory process, radiotherapy and cell growth inhibiting Drug therapy are brought out, and obtained challenging result is reported in non-renal anemia patient.See, for example Abels RI and the Rudnick SA Erythropoietin:evolving clinical appliactions. (erythropoietin: developing clinical practice) in Experimental Hematology 19:842-50 (1991); Graber SE and the Krantz SB Erythropoietin:biology and clinical use. (erythropoietin: biology and clinical application) in Hematology/Oncol.Clin.North Amer.3:369-400 (1989); Jelkman W and Gross AJ (eds) Erythropoietin.Springer, Berlin 1989; Koury MJ and the Bondurant MC The molecularmechanism of erythropoietin action. (molecular mechanism of erythropoietin effect) in EuropeanJournal of Biochemistry 210:649-63 (1992); The Erythropoietin (erythropoietin) of Krantz SB in Blood 77:419-34 (1991); The Erythropoietin.Biology and clinicalapplications. (erythropoietin of Tabbara IA in Archivesf Intemal MedicineI53:298-304 (1993), biology and clinical practice), above-mentioned list of references is all by complete introducing herein as a reference.
The present invention also provides and has regulated or treatment cell, tissue, organ, animal or patient's anemia or the conditions associated method of hemocyte, conditions associated and the following at least a disease association of wherein above-mentioned anemia or hemocyte, include but not limited at least a immune related diseases, cardiovascular diseases, infectious disease, pernicious and/or neuropathy.This method can randomly comprise cell, tissue, organ, animal or the patient who the particular composition of at least a effective dose or pharmaceutical composition is applied to this adjusting of needs, processing or treatment, and said composition or pharmaceutical composition contain at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant.
The present invention also provides and has regulated or the treatment cell, tissue, organ, the method of animal or patient's cancer/infectious disease, these diseases comprise, but be not limited only at least a acute or chronic bacterial infection, acute and chronic parasitism or infectious process, comprise antibacterial, virus and fungus transmission, HIV infection/HIV neuropathy, meningitis, hepatitis, septic arthritis, peritonitis, pneumonia, epiglottitis, colon bacillus 0157: h7, hemolytic uremic syndrome/thrombolytic thrombocytopenia purpura, malaria, the dengue fever hemorrhagic fever, leishmaniasis, leprosy, toxic shock syndrome, streptococcus myositis, gas gangrene, Mycobacterium tuberculosis, in the mycobacterium avium cell, pneumocystis carinii pneumonia, pelvic inflammatory disease, orchitis/epididymitis, Legionnella, Lyme disease, A type influenza, Epstein-Barr virus, the blood phagocyte syndrome relevant with vitals, fatal encephalitis/aseptic meningitis etc.; (ii) leukemia, acute leukemia, acute lymphatic leukemia (ALL), the B-cell, T-cell or FAB ALL, acute myelogenous leukemia (AML), chronic bone marrow leukemia (CML), chronic lymphocytic leukemia (CLL), hairy cell leukemia, myelodysplastic syndromes, lymphoma, Hokdkin disease, malignant lymphoma, non_hodgkin lymphoma, burkitt's lymphoma, multiple myeloma, kaposi's sarcoma, colon cancer, the pancreas pain, nasopharyngeal carcinoma, malignant histiocytosis, paraneoplastic syndrome/malignant hypercalcemia, solid tumor, adenocarcinoma, sarcoma, malignant melanoma etc.; Or (iii) nerve degeneration disease, multiple sclerosis, migraine, AIDS dull-witted compoundly levy, demyelinating disease, such as multiple sclerosis and acute transverse myelitis; Outer and the little brain disorder of cone is such as the infringement of cortex spinal cord system; The imbalance of ganglion basal or little brain disorder; The hyperkinesia dyskinesia is such as hungtington's chorea and senile chorea; The drug-induced dyskinesia is such as the dyskinesia that medicine brought out by sealing CNS dopamine receptor; The hypokinesia dyskinesia is such as parkinson; Carrying out property supranucleo paralysis; The structural infringement of cerebellum; Spinocerebellar degeneration is such as spinal ataxia, family ataxia, cerebellar cortex degeneration, multiple system degradation (Mencel, Dejerine-Thomas, Shi-Drager and Machado-Joseph); Systematicness imbalance (Refsum's disease, abetalipoproteinemia, ataxia, telangiectasis and the imbalance of mitochondrion multisystem); The imbalance of demyelination nuclear is such as multiple sclerosis, acute transverse myelitis; With the motor unit imbalance, such as nerve amyotrophy (anterior horn cell is degenerated, such as amyotrophic lateral sclerosis, werdnig-Hoffmann disease and juvenile Duchenne-Arandisease); Alzheimer; The middle age mongolism; Diffusion-type Lu Yishi body disease; Lu Yishi build alzheimer disease; The Wernicke-Korsakoff syndrome; Dipsorrhexia; The Creutzfeldt-Jakob disease; Subacute sclerosing panencephalitis, Hallerrorden-Spatz disease; With dementia pugilistica etc.This method can randomly comprise cell, tissue, organ, animal or the patient who the particular composition of effective dose or pharmaceutical composition is applied to this adjusting of needs, processing or treatment, and said composition or pharmaceutical composition contain at least a TNF antibody or specific part or variant.See the Merck Manual for example, 16 ThEdition, Merck ﹠amp; Company, Rahway, NJ (1992).
This method can randomly comprise cell, tissue, organ, animal or the patient who at least a particular composition of effective dose or pharmaceutical composition is applied to this adjusting of needs, processing or treatment, and said composition or pharmaceutical composition contain at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant.
The present invention also provides and has regulated or the treatment cell, tissue, organ, animal or at least a cardiovascular diseases's of patient method, this cardiovascular diseases comprises, but be not limited only at least a heart syndrome of fainting, myocardial infarction, congestive heart failure, apoplexy, ischemic stroke, hemorrhage, arteriosclerosis, atherosclerosis, the diabetes arteriosclerotic disease, hypertension, arterial hypertension, renal vascular hypertension, faint, shock, the syphilis of cardiovascular system, heart failure, pulmonary heart disease, primary pulmonary hypertension, arrhythmia, room ectopic beat, atrial flutter, auricular fibrillation, (continuing or paroxysmal), irregularity or many focuses atrial tachycardia, regular narrow QRS tachycardia, specific arrhythmia, ventricular fibrillation, His restraints arrhythmia, atrioventricular block, bundle branch block, the myocardial ischaemia sexual maladjustment, coronary heart disease, angina pectoris, myocardial infarction, cardiomyopathy, the dilatation and congestion cardiomyopathy, restrictive cardiomyopathy, valvular heart disease, endocarditis, pericardial disease, cardiac tumor, aorta and peripheral arterial aneurysm, aortic dissection, the aorta inflammation, ventral aorta and branched obturation thereof, the peripheral blood vessel imbalance, the imbalance of occlusive tremulous pulse, the peripheral arterial atheromatosis, thromboangiitis obliterans, the imbalance of functional peripheral tremulous pulse, Raynaud's phenomenon and disease, acrocyanosis, erythromelalgia, phlkebocholosis, thrombophlebitis, cirso-, arteriovenous fistula, lymphedema, lipedema, unstable angina, reperfusion injury, syndrome behind the pump, ischemia-reperfusion injury etc.This method can randomly comprise cell, tissue, organ, animal or the patient who the particular composition of effective dose or pharmaceutical composition is applied to this adjusting of needs, processing or treatment, and said composition or pharmaceutical composition contain at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant.
Arbitrary method of the present invention all can comprise cell, tissue, organ, animal or the patient who the particular composition of effective dose or pharmaceutical composition is applied to this adjusting of needs, processing or treatment, and said composition or pharmaceutical composition contain at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant.This method can randomly further comprise at treatment the collaborative of above-mentioned immunological disease to be used or therapeutic alliance, wherein above-mentioned at least a EPO mimetic CH 1 deleted mimetibodies, using of specific part or its variant further is included in before it, simultaneously and/or afterwards, use and be selected from following at least a medicine, be at least a TNF antagonist (for example, but be not limited only to TNF antibody or fragment, soluble TNF acceptor or fragment, its fusion rotein or micromolecule TNF antagonist), antirheumatic, muscle relaxant, anesthetics, on-steroidal anti-inflammatory medicaments (NSAID), analgesic, anesthetis, tranquilizer, local anesthetic, neuromuscular blocking agents, antibacterial (for example, aminoglycoside, antifungal, antiparasitic, antiviral agents, carbamyl, cephalosporin, fluoroquinolone, macrolide, penicillin, sulfanilamide, tetracycline, other antibacterial), antipsoriatic, corticosteroid, anabolic steroid, the diabetes related drugs, mineral, nutrient, the thyroid medicine, vitamin, the calcium associated hormone, diarrhea, cough medicine, antiemetic, antiulcerative, aperient, anticoagulant, erythropoietin (for example according to pool Ai Ting α), Fei Lasiting (G-CSF for example, Neupogen), Sargramostim (GM-CSF, Leukine), immunity, immunoglobulin, immunosuppressant (basiliximab for example, ciclosporin, daclizumab), growth hormone, hormone replaces medicine, the estrogen receptor instrumentality, mydriatic, cycloplegic, alkylating agent, antimetabolite, mitotic inhibitor, radiopharmaceutical, antidepressants, antimaniacal drugs, psychosis, antianxiety drugs, hypnotic, sympathomimetic, analeptic, donepezil, tacrine, asthma drug, the β synergist, suck steroid, the leukotriene inhibitor, methylxanthine, cromoglicic acid, epinephrine or analog, streptodornase α (Pulmozyme), cytokine or cytokine antagonist.Proper dosage is well known in the art.See Wells et al. for example, eds., PharmacotherapyHandbook, 2 NdEdition, Appleton and Lange, Starnford, CT (2000); PDR Pharmacopoeia, Tarascon Pocket Pharmacopoeia 2000, DeluxeEdition, Tarascon Publishing, Loma Linda, CA (2000), each list of references are all by complete introducing herein as a reference.
Analogies also can be applied to external, cultivate such as autologous bone marrow.In brief, before chemotherapy, in patient's body, take out bone marrow, and adopt TPO and/or EPO to handle, randomly in conjunction with analogue body, randomly in conjunction with one or more other cytokines.Subsequently, treated bone marrow is returned to through chemotherapeutical patient, to quicken the recovery of bone marrow.In addition, TPO can use separately and with EPO analogue body and/or EPO use in conjunction, also can be used to external expansion bone marrow or peripheral blood CFU-GM (PBPC).Before chemotherapy, can adopt stem cell factor (SCF) or G-CSF to stimulate bone marrow, so that early progenitor cell is discharged in the peripheral circulation.Randomly, from peripheral blood, collect and concentrate these CFU-GM, and adopt TPO and analogue body to handle its culture subsequently, randomly in conjunction with one or more other cytokines, comprise, but be not limited only to SCF, G-CSF, IL-3, GM-CSF, IL-6 or IL-11, making its differentiation and hypertrophy is high density megalokaryocyte culture, subsequently it is randomly returned in the patient's body after the too high dose chemotherapy.The TPO dosage range that is used for extracorporeal treatment bone marrow should be 100pg/ml-10ng/ml, preferably 500pg/ml-3ng/ml.The dosage of analogue body should be equal to EPO on activity, adoptable dosage range is 0.1 unit/ml-20 unit/ml, 0.5 unit/ml-2 unit/ml preferably, or any range wherein or numerical value.
Be applicable to the present composition, therapeutic alliance, collaboratively use, the TNF antagonist of device and/or method (further comprising at least a antibody of the present invention, specific part or its variant) comprises, but be not limited only to anti-TNF antibodies, its part binding fragment and with the bonded acceptor molecule of TNF specificity; Prevention and/or inhibition TNF are synthetic, TNF discharges or its chemical compound to the effect of target cell, such as thalidomide, tenidap, phosphodiesterase inhibitor (for example pentoxifylline and rolipram), A2b adenosine receptor antagonists and A2b adenosine receptor reinforcing agent; Stop and/or suppress the chemical compound of TNF receptor signalling, such as mitogen-activated protein(MAP) (MAP) inhibitors of kinases; Retardance and/or the cracked chemical compound of inhibition film TNF are such as metalloprotein alcohol inhibitor; Retardance and/or the active chemical compound of inhibition TNF are such as angiotensin converting enzyme (ACE) inhibitor (for example captopril); Generate and/or synthetic chemical compound with retardance and/or inhibition TNF, such as map kinase inhibitor.
Reductions such as " tnf antibody " used herein, " TNF antibody ", " TNF Alpha antibodies " or fragment, block, suppress, eliminate or disturbed external, original position and/or preferred intravital TNF alpha active.For example, suitable TNF human antibodies of the present invention can be in conjunction with TNF α, and comprises anti-TNF antibodies, its Fab and specified mutant or itself and the bonded zone of TNF alpha specific.Suitable TNF antibody or fragment also can reduce, block, eliminate, disturb, stop and/or suppress TNF RNA, DNA or protein synthesis, TNF release, the signalling of TNF receptor, film TNF cracking, TNF activity, TNF generation and/or synthesize.
Chimeric antibody cA2 is by in the high-affinity and mouse anti human TNF alpha IgG1 antibody, and promptly the antigen of A2 is in conjunction with variable region and IgG 1, and promptly the constant region of K immunoglobulin is formed.This IgG 1Fc has improved the alloantibody effector function in the district, has prolonged the circulation serum half-life of this antibody, and has reduced its immunogenicity.The affinity of chimeric antibody cA2 and epitope specificity derive from the variable region of murine antibody A 2.In a kind of particular, for the nucleic acid of these murine antibody A 2 variable regions of encoding, preferred source is the A2 hybridoma cell line.
Chimeric A2 (cA2) is with dosage dependence mode the two the cellulotoxic effect of natural and recombinant human TNF α that neutralized.From combining the test of chimeric antibody cA2 and recombinant human TNF α, the affinity costant of extrapolating chimeric antibody cA2 is 1.04 * 10 10M -1The method for optimizing that suppresses mensuration monoclonal antibody specificity and affinity by competition can be with reference to Harlow, et al., Antibodies:A Laboratory Manual, Cold Spring Harbor Laboratory Press, ColdSpring Harbor, New York, 1988; Colligan etal., eds., CurrentProtocols in Immunology, Greene Publishing Assoc.and WileyInterscience, New York. (1992-2002); Kozbor et al., Immunol.Today, 4:72-79 (1983); Ausubel et al., eds.Current Protocolsin Molecular Biology, Wiley Interscience, New York (1987-2002); And Muller, Meth.Enzymol., 95:589-601 (1983), these lists of references are all by complete introducing herein as a reference.
In a kind of particular, murine monoclonal antibody A2 is generated by the cell line that is named as c134A.Chimeric antibody cA2 is generated by the cell line that is named as c168A.
This area has been described and can be applicable to other monoclonal anti-TNF antibodies example of the present invention and (see that for example U.S. Patent No. 5,231,024; M ller, A.et al., Cytokine2 (3): 162-169 (1990); U. S. application No.07/943,852 (JIUYUE was submitted on the 11st in 1992); Rathjen et al., International Publication No.WO91/02078 (publication on February 21st, 1991); Rubin etal., EPO PatentPublication No.0218868 (publication on April 22nd, 1987); Yone et al., EPO Patent Publication No.0288088 (on October 26th, 1988); Liang, et al., Biochem.Biophys.Res.Comm.137:847-854 (1986); Meager, et al., Hybridoma 6:305-311 (1987); Fendly et al., Hybridoma 6:359-369 (1987); Bringman, et al., Hybridoma6:489-507 (1987); And Hirai, et al., J.Immunol.Meth.96:57-62 (1987), these lists of references are all by complete introducing herein as a reference).
The TNF acceptor molecule
The TNF acceptor molecule that the present invention preferably adopts is that those combine with TNF α high-affinity and (see Feldmann et al. for example, International Publication No.WO92/07076 (publication on April 30th, 1992); Schall et al., Cell 61:361-370 (1990); With Loetscher et al., Ce1161:351-359 (1990), these lists of references are all by complete introducing herein as a reference), and randomly have the TNF acceptor molecule of reduced immunogenicity.55kDa (p55TNF-R) and 75kDa (p75TNF-R) TNF cell surface receptor are particularly useful in the present invention.The clipped form of these receptors comprises the extracellular domain (ECD) of this receptor or its funtion part (seeing Corcoran et al. for example, Eur.J.Biochem.223:831-840 (1994)), and is also useful in the present invention.Detected the clipped form that this TNF receptor comprises ECD in urine and serum, promptly the TNF α of 30kDa and 40kDa suppresses conjugated protein (Engelmann, H.et al., J.Biol.Chem.265:1531-1536 (1990)).TNF receptor multimeric molecule and TNF immunity receptor fusion molecule, and derivant and fragment or part are other TNF acceptor molecule examples that can be applicable to the inventive method and compositions.Can be applicable to TNF acceptor molecule of the present invention and be characterised in that it has the long-term treatment patient, and can well arrive the ability of relief of symptoms admirably, and toxicity is low.Reduced immunogenicity and/or high-affinity, and other not clear and definite feature all may work to the therapeutic effect that is obtained.
Useful in the present invention TNF receptor multimeric molecule comprises via one or more peptide linkers or other non-peptide linker, such as Polyethylene Glycol (PEG), and whole or funtion part of the ECD of the two or more TNF receptors that link together.This multimeric molecule can further comprise the signal peptide of secreted protein, to guide the expression of this multimeric molecule.U. S. application No.08/437 has described these multimeric molecules and preparation method thereof in 533 (submissions on May 9 nineteen ninety-five), and the content of this list of references by complete introducing herein as a reference.
Useful TNF immunity receptor fusion molecule comprises at least one part of one or more immunoglobulin molecules and whole or funtion part of one or more TNF receptors in the inventive method and compositions.These immunity receptor fusion molecule can fitted to be monomer, or heteromultimers or homology polymer.This immunity receptor fusion molecule also can be unit price or multivalence.An example of this TNF immunity receptor fusion molecule is TNF receptor/IgG fusion rotein.This area has been described TNF immunity receptor fusion molecule and preparation method thereof (Lesslauer et al., Eur.J.Immunol.21:2883-2886 (1991); Ashkenazi et al., Proc.Natl.Acad.Sci.USA 88:10535-10539 (1991); Peppel et al., J.Exp.Med.174:1483-1489 (1991); Kolls et al., Proc.Natl.Acad.Sci.USA 91:215-219 (1994); Butler et al., Cytokine 6 (6); 616-623 (1994); Baker et al., Eur.J.Immunol.24:2040-2048 (1994); Beutler et al., U.S. Patent No. 5,447,851; With U. S. application No.08/442,133 (submissions on May 16 nineteen ninety-five), these lists of references are all by complete introducing herein as a reference).The method for preparing the immunity receptor fusion molecule also can be with reference to Capon et al., U.S. Patent No. 5,116,964; Capon et al., U.S. Patent No. 5,225,538; With Capon et al., Nature 337:525-531 (1989), these lists of references are all by complete introducing herein as a reference.
The function equivalent of TNF acceptor molecule, derivant, fragment or district refer to the specific part of this TNF acceptor molecule, or the part of the TNF acceptor molecule of encoding in this TNF acceptor molecule sequence, its size and sequence be enough to make its on function to can be applicable to TNF acceptor molecule of the present invention similar (for example, with TNF α high-affinity combine, and have reduced immunogenicity).The function equivalent of TNF acceptor molecule be also included within on the function to can be applicable to TNF acceptor molecule of the present invention similar (for example, with TNF α high-affinity combine, and have reduced immunogenicity) modification TNF acceptor molecule.For example, the function equivalent of TNF acceptor molecule can contain " SILENT " codon or one or more amino acid replacement, disappearance or interpolation (for example are replaced into another acidic amino acid with an acidic amino acid; Or with one the coding identical or different hydrophobic amino acid codon be replaced into another the coding hydrophobic amino acid codon).Referring to Ausubel et al., Current Protocols in Molecular Biology, Greene PublishingAssoc.and Wiley-Interscience, New York (1987-2002).
Cytokine include but not limited to all known cytokines.See, for example CopewithCytokines.com.Cytokine antagonist include but not limited to any antibody, fragment or analogue body, any solvable receptor, fragment or analogue body, any micromolecule antagonist, or the combination in any of above-mentioned substance.
Any method of the present invention all can comprise the method for the imbalance that is used for the treatment of protein mediation, this method comprises cell, tissue, organ, animal or the patient who the particular composition of effective dose or pharmaceutical composition is applied to this adjusting of needs, processing or treatment, and said composition or pharmaceutical composition contain at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant.This method can randomly further comprise at treatment the collaborative of immunological disease to be used or therapeutic alliance, the using of wherein above-mentioned at least a EPO mimetic CH 1 deleted mimetibodies, specific part or its variant further be included in before it, simultaneously and/or afterwards, use and be selected from least a other cytokine, such as IL-3 ,-6 and-11; Stem cell factor; At least a material among G-CSF and the GM-CSF.
Typically, treatment to pathologic condition is to realize by at least a EPO mimetic CH 1 deleted mimetibodies compositions of using effective dose or dosage, promptly according to the contained given activity of said composition, the scope of each dosage amounts to average out to, the per kilogram weight in patients is used at least approximately at least a EPO mimetic CH 1 deleted mimetibodies or the specific part or the variant of 0.01-500 milligram, and preferably per kilogram weight in patients single or multiple is used at least approximately 0.1-100 milligram EPO mimetic CH 1 deleted mimetibodies or specific part or variant.Alternatively, effectively serum-concentration can comprise that single or multiple uses the 0.1-5000ug/ml serum-concentration that is obtained.Proper dosage is that the doctor is known, and depends on specified disease situation, the given activity of the compositions of using and the particular patient of just receiving treatment natch.Under some situation,, be necessary to carry out repetitive administration, promptly use separately, and before obtaining appointed date dosage or effect, repeat this and use separately with the repetition of specifically monitored dosage or dosing for obtaining the TA amount.
Preferred dose can randomly comprise uses 0.01 at every turn, 0.02,0.03,0.04,0.05,0.06,0.07,0.08,0.09,0.1,0.2,0.3,0.4,0.5,0.6,0.7,0.8,0.9,1,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19,20,21,22,23,24,25,26,27,28,29 and/or 30mg/kg, or any range wherein, numerical value or mark, or use by single or multiple and to obtain 0.1,0.5,0.9,1.0,1.1,1.2,1.5,1.9,2.0,2.5,2.9,3.0,3.5,3.9,4.0,4.5,4.9,5.0,5.5,5.9,6.0,6.5,6.9,7.0,7.5,7.9,8.0,8.5,8.9,9.0,9.5,9.9,10,10.5,10.9,11,11.5,11.9,12,12.5,12.9,13.0,13.5,13.9,14.0,14.5,15,15.5,15.9,16,16.5,16.9,17,17.5,17.9,18,18.5,18.9,19,19.5,19.9,20,20.5,20.9,21,22,23,24,25,26,27,28,29,30,35,40,45,50,55,60,65,70,75,80,85,90,96,100,200,300,400,500,600,700,800,900,1000,1500,2000,2500,3000,3500,4000,4500 and/or the 5000ug/ml serum-concentration, or any range wherein, numerical value or mark.
Alternatively, can be according to known facts, such as the drug effect characteristic of particular agent, and mode of administration and approach; Receiver's age, health condition and body weight; The frequency of the nature and extent of symptom, the kind of combined treatment, treatment and Expected Results change application dosage.Usually, the dosage of effective ingredient can be per kilogram of body weight and uses about 0.1-100 milligram.General dose is 0.1-50, and preferred dose is that per kilogram of body weight is used the 0.1-10 milligram at every turn, or obtains Expected Results effectively by continuous releasing pattern.
A limiting examples is, can be to the treatment of the mankind or animal by disposable or periodically use at least a EPO mimetic CH 1 deleted mimetibodies of the present invention or specific part or the variant dosage of 0.0-100mg/kg, single such as adopting, infusion or repeated doses, the 1st, 2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19,20,21,22,23,24,25,26,27,28,29,30,31,32,33,34,35,36,37,38, at least one sky in 39 or 40 days, or alternatively 1,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18, every day at least one cycle in 19 or 20 weeks, or use 0.5 the every day in the combination in any cycle of above-mentioned natural law and all numbers, 0.9,1.0,1.1,1.5,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19,20,21,22,23,24,25,26,27,28,29,30,40,45,50,60,70,80,90 or at least a EPO mimetic CH 1 deleted mimetibodies of the present invention or specific part or the variant of 100mg/kg.
The common per unit of the dosage form that use suitable inside (compositions) or each container contain about 0.0001 milligram-about 500 milligrams of effective ingredient.In these pharmaceutical compositions, the amount of effective ingredient is calculated by weight the about 0.5-95% that should be the said composition gross weight usually.
For parenteral administration, EPO mimetic CH 1 deleted mimetibodies or specific part or variant can be configured to solution, suspension, emulsion or lyophilized powder with the acceptable parenteral excipient of materia medica or in the mode that provides respectively.The example of this excipient is water, saline, RingerShi solution, glucose solution and 5% human serum albumin.Also can adopt liposome and non-aqueous excipient, such as expressed oi.Excipient or lyophilized powder can contain the additive of keeping isotonicity (for example sodium chloride, mannitol) and chemical stability (for example buffer agent and antiseptic).This prescription is by known or suitable technique sterilization.
Suitable pharmaceutical carrier canonical reference document in the field, promptly Remington ' sPharmaceutical Sciences all describes in the latest edition of A.Osol to some extent.
Therapeutic administration
According to the present invention, can adopt many known and mode of administration of having researched and developed, be used at least a EPO mimetic CH 1 deleted mimetibodies of the present invention or the specific part or the variant of administering therapeutic effective dose.Though adopted pulmonary administration in the following content, still can adopt other mode of administration, and obtain suitable effect according to the present invention.
EPO mimetic CH 1 deleted mimetibodies of the present invention can form solution, emulsion, colloid or suspension in carrier, or with powder type, by utilize multiple be applicable to suck or described herein or apparatus and method that other known mode of this area is used in any one, passed and send.
Parenteral prescription and using
The prescription that is used for parenteral administration can contain usual excipients, sterilized water or saline, such as the oil of the ployalkylene glycol of Polyethylene Glycol, plant origin, hydrogenated naphthalene etc.According to known method, can prepare the aqueous or the oiliness suspension of injection by adopting suitable emulsifying agent or humidizer and suspending agent.The medicament of injection can be a diluent nontoxic, can not be oral, such as the aqueous solution that forms in solvent or sterile injectable solution or suspension.Effectively excipient or solvent can adopt water, RingerShi solution, isotonic saline solution etc.; Usual vehicle or suspended solvents then can adopt aseptic expressed oi.With regard to these purposes, can adopt the expressed oi and the fatty acid of any kind of, comprise natural synthetic or semisynthetic fatty oil or fatty acid; Natural synthetic or semisynthetic monoglyceride or diester or three esters.Parenteral administration is known in the art, comprise, but be not limited only to conventional injecting pathway, U.S. Patent No. 5,851, air-pressure type needleless injection device and the U.S. Patent No. 5,839 described in 198, the laser beam perforation apparatus of describing in 446, these two patents all by complete introducing herein as a reference.
Can select and recommend and pass
The invention further relates to by parenteral, subcutaneous, intramuscular, intravenous, pill, vagina, rectum, oral cavity, Sublingual, intranasal or percutaneous approach and use at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant.Protein, EPO mimetic CH 1 deleted mimetibodies or specific part or variant compositions can be produced and be used for parenteral (subcutaneous, intramuscular or intravenous) and use, and are good with liquid solution or suspensions especially; When being used for vagina or rectal administration,, be good such as paste and suppository then especially with semi-solid form; When oral cavity or sublingual administration, be good with tablet or capsule form especially then; During intranasal administration, be good with powder, nasal drop or aerosol or particular agent form especially; Applied dermally is good with gel, ointment, lotion, suspensions especially then, or contain the chemical intensifier that can change skin texture or improve percutaneous medicine card Chinese medicine concentration, such as dimethyl sulfoxine (Junginger, et al. is at " Drug PermeationEnhancement "; Hsieh, D.S., Eds., pp59-90 (complete introducing herein as a reference for Marcel Dekker, Inc.New York 1994)), or contain to make and contain the oxidant (WO 98/53847) that protein and peptide prescription are applied to skin, maybe can apply electric field, and cause the of short duration approach of passing that send, such as electroporation, maybe can improve the mobility that electrically charged medicine passes skin, such as ionotherapy, but or the using ultrasound ripple, import method (U.S. Patent No. 4 such as ultrasound wave, 309,989 and 4,767,402) medicine card send delivery system (above-mentioned publication and patent are all by complete introducing as a reference) herein.
Lung/nasal administration
For pulmonary administration, preferably send at least a EPO mimetic CH 1 deleted mimetibodies or the specific part or the variant compositions of passing granular size, so that it arrives at the respiratory tract of lung or hole bottom effectively.According to the present invention, can send by any one of multiple suction that is used for suction-type administering therapeutic medicament known in the art or Nasal delivery devices and pass at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant.These devices that atomizing prescription is deposited in patient's hole chamber or the alveolar comprise metered dose inhaler, nebulizer, dry powder generator, aerosol apparatus etc.Other device that is applicable to guiding EPO mimetic CH 1 deleted mimetibodies or specific part or variant lung or nasal administration also is known in the art.Above-mentioned all devices all can be used the prescription that is fit to use, so that EPO mimetic CH 1 deleted mimetibodies or specific part or variant are distributed in the aerosol.This aerosol can be made up of solution (aqueous and non-aqueous) or solid particle.Metered dose inhaler is as Yentolion Metered dose inhaler typically adopts propulsive gas, and need start (see that for example WO 94/16970, WO 98/35888) in intake period.Diskus is as Turbuhaler TM(Astra), Rotahaler (Glaxo), Diskus (Glaxo), Spiros TMInhaler (Dura), the device of selling by Inhale Therapeutics, and Spinhaler Powder inhalator (Fisons) all sucks mixed-powder (US4668218Astra, EP 237507Astra by respiratory activity, WO 97/25086Glaxo, WO 94/08552Dura, US 5458135Inhale, WO 94/06498Fisons, complete introducing is herein as a reference).Nebulizer such as AERx TMAradigm, Ultravent Nebulizer (Mallinckrodt) and Acorn II Nebulizer (Marquest MedicalProducts) (US 5404871Aradigm, WO 97/22376), can generate aerosol by solution, inhaler such as metered dose inhaler, Diskus then can produce the granule aerosol, and above-mentioned list of references all by complete introducing herein as a reference.Commerce can provide the above-mentioned particular instance of suction apparatus only category of the present invention not to be construed as limiting the representative of putting into practice specific device of the present invention for being applicable to.Preferably, send by Diskus or aerosol apparatus and pass the compositions that comprises at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant.Be applicable to that the suction apparatus of using at least a EPO mimetic CH 1 deleted mimetibodies of the present invention or specific part or variant has some gratifying features.For example, send the advantage of passing to be reliably, can to reappear by suction apparatus and accurately.This suction apparatus can randomly send passs little dried granule, and for example for good inhalable, this granule is preferably about 1-5 μ m less than about 10 μ m.
Use EPO mimetic CH 1 deleted mimetibodies or specific part or variant compositions with Sprayable
Comprise that the proteic spraying of EPO mimetic CH 1 deleted mimetibodies or specific part or variant compositions can be prepared by nozzle by suspension or the solution that forces at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant under pressure.The size of this nozzle and configuration, applying pressure and liquid feed rate all can be selected to obtain the output and the particle size of expection.Electron spray can pass through, and for example electric field and capillary tube or nozzle material-feeding obtain.Send the advantage of passing at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant compositions protein body to be by aerosol apparatus, particle size is less than about 10 μ m, preferable range is the about 5 μ m of about 1um-, and highly preferred scope is the about 3 μ m of about 2 μ m-.
Be applicable to that at least a EPO mimetic CH 1 deleted mimetibodies of aerosol apparatus or specific part or variant compositions protein prescription typically comprise EPO mimetic CH 1 deleted mimetibodies or the specific part or the variant compositions albumen of aqueous solution form, its concentration is at least a EPO mimetic CH 1 deleted mimetibodies or specific part or the variant compositions albumen that every ml soln contains the about 20mg of about 1mg-.This prescription can comprise such as excipient, buffer agent, isotonic agent, antiseptic, surfactant and preferred zinc.This prescription also can comprise and is used to make protein stabilized excipient of EPO mimetic CH 1 deleted mimetibodies or specific part or variant compositions or reagent, such as buffer agent, Reducing agent, filling albumen or carbohydrate.Can be used for preparing the proteic filling albumen of EPO mimetic CH 1 deleted mimetibodies or specific part or variant compositions and comprise albumin, protamine etc.Can be used for preparing the proteic typical saccharide of EPO mimetic CH 1 deleted mimetibodies or specific part or variant compositions and comprise sucrose, mannitol, lactose, trehalose, glucose etc.This EPO mimetic CH 1 deleted mimetibodies or specific part or variant compositions protein prescription also can comprise surfactant, the EPO mimetic CH 1 deleted mimetibodies that atomizing caused or specific part or the proteic spatial induction cohesion of variant compositions in the time of can reducing or prevent to form aerosol by solution.Multiple conventional surfactants all can be employed, such as polyoxyethylene fatty acid ester and ethanol and polyoxyethylene sorbitol fatty acid ester.Its application quantity scope should be the 0.001-14% of formulation weight usually.For the especially preferred surfactant of application of the present invention is Tween-81, polysorbate80, polysorbate20 etc.Proteinic other reagent that is used to prepare such as analogue body or specific part or variant is known in the art, also is included in the prescription.
Use EPO mimetic CH 1 deleted mimetibodies or specific part or variant compositions by nebulizer
EPO mimetic CH 1 deleted mimetibodies or specific part or variant compositions albumen can pass through nebulizer, such as blast atomizer or soniclizer and used.Typically, in blast atomizer, adopt compressed air source to form the high-speed air injection stream by nozzle.When gas when nozzle diffuses out, formed low-pressure area, EPO mimetic CH 1 deleted mimetibodies or specific part or the proteic solution of variant compositions are sucked, and make its capillary tube by linking to each other with liquid reservoir.When being derived from liquid stream capillaceous and leaving this capillary tube, be cut into unsettled filamentous and drop, just formed aerosol.Can use a series of structure, flow velocity and flow deflector type, from the particular spray nebulizer, to obtain the performance characteristic of expection.In soniclizer, adopt high-frequency electrical energy to form the mechanical energy of concussion, typically applying piezoelectric formula transducer.This energy directly or by means of coupling liquid is transferred to EPO mimetic CH 1 deleted mimetibodies or specific part or variant compositions protein prescription, has just formed to comprise this EPO mimetic CH 1 deleted mimetibodies or specific part or the proteic aerosol of variant compositions.Send the advantage of passing EPO mimetic CH 1 deleted mimetibodies or specific part or variant compositions protein body to be by nebulizer, particle size is less than about 10 μ m, preferable range is the about 5 μ m of about 1 μ m-, and highly preferred scope is the about 3 μ m of about 2 μ m-.
Be applicable to and spray or play EPO mimetic CH 1 deleted mimetibodies or specific part or the variant compositions albumen that at least a EPO mimetic CH 1 deleted mimetibodies of acoustic wave type nebulizer or specific part or variant prescription typically comprise the aqueous solution form that its concentration is at least a EPO mimetic CH 1 deleted mimetibodies or specific part or the misfolded proteins that every ml soln contains the about 20mg of about 1mg-.This prescription can comprise such as excipient, buffer agent, isotonic agent, antiseptic, surfactant and preferred zinc.This prescription also can comprise and is used to the excipient or the reagent that make at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant compositions protein stabilized, such as buffer agent, Reducing agent, filling albumen or carbohydrate.Can be used for preparing the proteic filling albumen of at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant compositions and comprise albumin, protamine etc.The typical saccharide that can be used for preparing at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant comprises sucrose, mannitol, lactose, trehalose, glucose etc.This at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant prescription also can comprise surfactant, at least a EPO mimetic CH 1 deleted mimetibodies that atomizing caused in the time of can reducing or prevent to form aerosol by solution or the spatial induction cohesion of specific part or variant.Multiple conventional surfactants all can be employed, such as polyoxyethylene fatty acid ester and ethanol and polyoxyethylene sorbitol fatty acid ester.Its application quantity scope should be the 0.001-4% of formulation weight usually.For the especially preferred surfactant of application of the present invention is Tween-81, polysorbate80, polysorbate20 etc.Other reagent that is used to prepare such as at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant proteins is known in the art, also is included in the prescription.
Use EPO mimetic CH 1 deleted mimetibodies or specific part or variant compositions by metered dose inhaler
In metered dose inhaler (MDI), propellant, at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant and any excipient or other additive all are inclusive in the canister, have formed the mixture that comprises liquified compressed gas.Starting of metering valve discharges this mixture with aerosol form, this aerosol preferably contains the granule of size range less than about 10um, and the preferred sizes scope is the about 5 μ m of about 1 μ m-, and highly preferred size range is the about 3 μ m of about 2 μ m-.By using by several different methods well known by persons skilled in the art, comprise EPO mimetic CH 1 deleted mimetibodies or the specific part or the variant compositions protein prescription of method preparations such as jet grinding, spray drying, critical point condensation, can obtain ideal aerosol particle size.Preferred metered dose inhaler comprises the metered dose inhaler of being produced by 3M or Glaxo, and has used hydrogenation fluorohydrocarbon propellant.
Being applicable to that at least a EPO mimetic CH 1 deleted mimetibodies of metered dose inhaler device or specific part or variant prescription generally include contains at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant, and can in non-aqueous media, form the attritive powder of suspension, for example, be suspended in the propellant by means of surfactant.This propellant can be any conventional material that can be used for this purposes, such as chlorine fluorohydrocarbon, hydrogenated chloride fluorine for hydrocarbon, hydrogenation fluorohydrocarbon or hydrocarbon, comprise Arcton 11, dichlorodifluoromethane, dichloro-tetrafluoro ethanol and 1,1,1,2-tetrafluoroethane, HFA-134a hydrofluoroalkane-134a ")), HFA-227 hydrogen fluoroalkane-227 etc.Preferred propellant is the hydrogenation fluorohydrocarbon.Can select surfactant, to stablize at least a EPO mimetic CH 1 deleted mimetibodies or the specific part or the variant of suspended form in this propellant, to protect this activating agent not by chemical degradation etc.Suitable surfactant comprises sorbitan trioleate, soybean lecithin, oleic acid etc.In some situation, the solution aerosol preferably adopts such as alcoholic acid solvent.Known in the artly can be used for preparing protein, also can be included in the present invention's prescription such as proteinic other reagent of the present invention.
Any those of ordinary skill in this area will be appreciated that all the inventive method can be achieved by at least a EPO mimetic CH 1 deleted mimetibodies of pulmonary administration or specific part or variant compositions by the device of not describing herein.
Mucosa prescription and using
For absorption via mucomembranous surface, the compositions of at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant and application process comprise contain a large amount of submicron particles, mucosa adheres to the peptide of macromole, biologically active and the emulsion of aqueous continuous phase, can adhere to by the mucosa of realizing this emulsion granules and promote its (U.S. Patent No. 5 of absorption via mucomembranous surface, 514,670).The mucomembranous surface that is fit to application emulsion of the present invention can comprise the route of administration of cornea, conjunctiva, oral cavity, Sublingual, nose, vagina, lung, stomach, intestinal and rectum.The prescription that is used for vagina or rectal administration, for example suppository can contain excipient, for example ployalkylene glycol, vaseline, cocoa butter etc.The prescription that is used for intranasal administration can be solid, and contains excipient, and for example lactose perhaps can be the aqueous or the oily solution of nasal drop.For oral, excipient comprises (U.S. Patent No. 5,849,695) such as sugar, calcium stearate, magnesium stearate, pre-gelatinized starches.
Formula of oral and using
Be used for collaborative the using that oral prescription depends on adjuvant (for example resorcinol and nonionic surfactant, such as polyoxyl 10 oleyl ether and n-cetyl polyvinylether), permeability with artificial raising intestinal wall, this prescription also depends on collaborative the using (for example trypsin inhibitor, diisopropyl fluorophosphate (DFP) (DFF) and aprotinin) of enzyme inhibitor, degrades with inhibitory enzyme.The activity composition chemical compound that is used for oral solid type dosage form can mix with at least a additive, this additive comprises sucrose, lactose, cellulose, mannitol, trehalose, Raffinose, maltose alcohol, glucosan, starch, agar, arginine, chitin, chitosan, pectin, gum tragacanth, Radix Acaciae senegalis, gelatin, collagen protein, casein, albumin, synthetic or semi synthetic polymer, and glyceride.These dosage forms also can contain the additive of other type, for example torpescence diluent, lubricant, such as magnesium stearate, P-hydroxybenzoic acid, such as the antiseptic of sorbic acid, ascorbic acid, alpha-tocopherol, such as the antioxidant of cysteine, disintegrating agent, binding agent, thickening agent, buffer agent, sweeting agent, flavoring agent, aromatizing agent etc.
Tablet and pill can be further processed in the enteric coating preparation.Be used for the pharmaceutical solutions that oral liquid preparation comprises Emulsion, syrup, elixir, suspensoid and can be used for medical usage.These preparations can contain the torpescence diluent that is usually used in this field, as water.It is the drug delivery system (U.S. Patent No. 4,239,754) of insulin and heparin that liposome also once was described to.More recently be that the microsphere of the artificial polymer of kilnitamin (albuminoid) once was used to send drug delivery (U.S. Patent No. 4,925,673).In addition, U.S. Patent No. 5,879,681 and U.S. Patent No. 5,5,871,753 in the carrier compound described be used to oral sending and pass bioactivator known in the art.
Percutaneous prescription and using
For applied dermally, this at least a EPO mimetic CH 1 deleted mimetibodies or specific part or variant are sealed in the delivery device, such as liposome or polymerization nanoparticle, microgranule, microcapsule or microsphere (without otherwise indicated, can be called microgranule jointly).It is known that many suitable devices are arranged, comprise by synthetic polymer, as polyhydroxy acid, as polylactic acid, polyglycolic acid and copolymer thereof, poe, poly-anhydride and poly-polyphosphazene, and natural polymer, such as collagen protein, polyamino acid, albumin and other protein, alginate and other polysaccharide, and the microgranule (U.S. Patent No. 5 made of the combination of above-mentioned substance, 814,599).
Chronic administration and prescription
Some the time, making The compounds of this invention cross over the long time period, to be applied to the patient again be ideal, for example, begins a thoughtful year section from single administration.Can utilize multiple slow release, storage or transplant dosage form.For example, dosage form can contain the acceptable nontoxic salts of materia medica of specific compound, this chemical compound has low solubility in body fluid, for example, (a) has polyprotic acid, such as the acid-addition salts of acid such as phosphoric acid, sulphuric acid, citric acid, tartaric acid, tannic acid, pamoic acid, alginic acid, polyglutamic acid, LOMAR PWA EINECS 246-676-2 or naphthalenedisulfonic acid, Poly Gal A Galacturonan; (b) have multivalent metal cation,, or have, the organic cations salt that N '-diphenyl-ethylenediamine or ethylenediamine form by for example N such as zinc, calcium, bismuth, barium, magnesium, aluminum, copper, cobalt, nickel, cadmium etc.; (c) (a) with (b) combination, for example tannic acid zinc salt.In addition, The compounds of this invention, or preferred, insoluble salt as above-mentioned salt, can be formulated into gel relatively, for example is applicable to injection, and for example contains, the monostearate alumina gel of Oleum sesami.Especially preferred salt is zinc salt, tannic acid zinc salt, embonate etc.Other type slow release of injection is stored to fill a prescription to contain and is disperseed to be sealed in slow degraded, nontoxic, nonantigenic polymer, such as U.S. Patent No. 3,773, and chemical compound or salt in 919 described polylactic acid/polyglycolic acid polymer.This chemical compound or, preferred insoluble salt relatively also can be formulated in the cholesterol substrate silicone ball as above-mentioned salt, especially is applied to animal.Other slow release, storage or transplanting prescription, for example gas or liquid fatty body can be referring to this area list of references (United States Patent(USP) Nos.s 5,770,222 and " Sustained and Controlled Release Drug Delivery Systems " (continue and controlled release drug send delivery system), J.R.Robinson ed., Marcel Dekker, Inc., N.Y., 1978).
Above summarized the present invention, will be more readily understood the present invention, and following example only is used for illustration, the present invention is not construed as limiting by following example.
Cloning and the expression of embodiment 1:EPO mimetic CH 1 deleted mimetibodies in mammalian cell
Typical case's mammalian expression vector contains the promoter element that at least one can mediate the mRNA transcription initiation, EPO mimetic CH 1 deleted mimetibodies or specific part or variant coded sequence, and tanscription termination and the needed signal of transcription product polyadenylation.Other element comprises enhancer, Kozak sequence and the both sides intervening sequence with RNA shearing donor and acceptor site.Efficiently transcribe and to realize as the long terminal repeat (LTRS) in RSV, HTLVI, HIVI source and the early promoter in cytomegalovirus (CMV) source by adopting early stage and late promoter, the retrovirus in SV40 source.But also can adopt cell element (for example human actin promoter).Can be used for realizing that suitable expression vector of the present invention comprises, for example, pIRESlneo, pRetro-Off, pRetro-On, PLXSN, or pLNCX (Clonetech Labs, PaloAlto, CA), pcDNA3.1 (+/-), pcDNA/Zeo (+/-) or pcDNA3,1/Hygro (+/-) (Invitrogen), PSVL and PMSG (Pharmacia, Uppsala, Sweden), pRSVcat (ATCC 37152), pSV2dhfr (ATCC 37146) and pBC12MI (ATCC67109).The mammalian cell that may be utilized comprises human Hela 293, H9 and Jurkat cell, mouse NIH 3T3 and C127 cell, Cos1, Cos7 and CV1, Carnis Coturnicis japonicae QC1-3 cell, mouse Lcell and Chinese hamster ovary (CHO) cell.
Alternatively, gene can be expressed in the specific stable cell lines, and this cell line contains the said gene that is be integrated in its chromosome.Have selectable marker, such as the cotransfection of dhfr, gpt, neomycin or hygromycin can realize to the evaluation of transfectional cell with separate.
Rotaring redyeing gene also can be amplified, with EPO mimetic CH 1 deleted mimetibodies or the specific part or the variant of expressing a large amount of codings.DHFR (dihydrofolate reductase) be marked with help develop carried be concerned about the hundreds of of gene or even the cell line of thousands of copies.Other useful selected marker is glutamine synthase (GS) (Murphy, et al., Biochem.J.227:277-279 (1991); Bebbington, et al., Bio/Technology 10:169-175 (1992)).Utilize these labellings, can in selective medium, cultivate mammalian cell, and select to have the cell of high resistance.These cell lines contain the amplification gene that is be integrated in its chromosome.Chinese hamster ovary (CHO) and NSO cell are normally used for generating EPO mimetic CH 1 deleted mimetibodies or specific part or variant.
Expression vector pC1 and pC4 contain strong promoter (LTR) (the Cu11enet al. of rous sarcoma virus, Molec.Cell.Biol.5:438-447 (1985)), add a fragment (Boshart, et al., Cell 41:521-530 (1985)) of CMV-enhancer.Multiple clone site for example has restriction endonuclease cracking site BamHI, XbaI and Asp718, help be concerned about the cloning of gene.This carrier also contains the polyadenylation and the termination signal of rat proinsulin protogene except that containing 3 ' intron.
Cloning in Chinese hamster ovary celI and expression
Carrier pC4 is used to express EPO mimetic CH 1 deleted mimetibodies or specific part or variant.Plasmid pC4 is the derivant (ATCC is numbered No.37146) of plasmid pSV2-dhfr.This plasmid contains the mice DHFR gene that is subjected to the control of SV40 early promoter.By at the selective medium that is supplemented with the chemotherapeutant methotrexate (α minus MEM for example, LifeTechnologies, Gaithersburg, MD) middle cultured cell, can filter out by these plasmid transfections, and lack the active Chinese hamster ovary cell of dihydrofoilic acid or other cell.Many lists of references have all write down the amplification of DHFR gene in the cell that methotrexate (MTX) is had resistance and (have seen F.W.Alt for example, etal., J.Biol.Chem.253:1357-1370 (1978); J.L.Hamlin and C.Ma, Biochem.et Biophys.Acta1097:107-143 (1990); With M.J.Pa ge and M.A.Sydenham, Biotechnology 9:64-68 (1991)).The cell of growing in progressive concentration MTX causes the excessive generation of target enzyme DHFR by gene-amplification DHFR gene, thereby has produced the resistance to medicine.If second gene is connected to this DHFR gene, then this second gene is usually by common amplification and overexpression.This method known in the art can be used to exploitation and carry the cell line that surpasses 1,000 copy of this amplification gene.Subsequently, when extracting methotrexate, can obtain to contain the cell line of amplification gene, this amplification gene has been integrated in one or more chromosomes of host cell.
Plasmid pC4 contains the strong promoter (Cullen that is useful on the rous sarcoma virus long terminal repeat (LTR) of gene that expression is concerned about, et al., Molec.Cell.Biol.5:438-447 (1985)), add the fragment (Boshart of separation from human cytomegalovirus (CMV) immediate early gene enhancer, et al., Cell 41:521-530 (1985)).The downstream of this promoter is BamHI, XbaI and the Asp718 restriction endonuclease cracking site that can realize that said gene is integrated.Behind these cloning sites, this plasmid contains the polyadenylation site of 3 ' intron and rat proinsulin protogene.Other efficient promoter also can be used to express, and for example human b-actin promoter, SV40 is early stage or late promoter or other retrovirus, as the long terminal repeat in HIV and HTLVI source.The Tet-Off of Clontech and Tet-On gene expression system and similar system can be used to express EPO (M.Gossen, and H.Bujard, Proc.Natl.Acad.Sci.USA89:5547-5551 (1992)) with the scalable approach in mammalian cell.For the polyadenylation of mRNA, also can adopt other signal, for example from the signal of human growth hormone or globulin gene.Adopting selectable marker, on the basis such as gpt, G418 or the common transfection of hygromycin, also can select carried be concerned about the stable cell lines of gene, this gene has been integrated in its chromosome.Adopt at first that selectable marker has superiority more than a kind, for example G418 adds methotrexate.
By operational approach known in the art, adopt restriction endonuclease digested plasmid pC4, and adopt calf enteral phosphatase dephosphorylation subsequently.From 1% agarose gel, isolate this carrier subsequently.
According to the known method step, adopt the DNA sequence of the complete EPO mimetic CH 1 deleted mimetibodies of coding or specific part or variant, sequence shown in the SEQ ID NOS:13,14,15,16,17,18 for example, this DNA sequence is corresponding with the HC and the LC variable region of EPO mimetic CH 1 deleted mimetibodies of the present invention.The isolating nucleic acid of the suitable human constant region of coding (being HC and LC district) also can be provided in (as providing among the carrier p1351) this construct.
Make subsequently that the separation of coding DNA is variable to be connected with the T4DNA ligase with the dephosphorylation carrier with constant region.Subsequently, transfection Escherichia coli HB101 or XL-1Blue cell, and adopt, for example the restriction enzyme analysis method is identified among the plasmid pC4 and is contained the segmental antibacterial of above-mentioned insertion.
Adopt Chinese hamster ovary (CHO) cell that lacks active DHFR gene to be used for transfection.Adopt transfection reagent lipofectin, by 0.5ug plasmid pSV20-neo cotransfection 5 μ g expression plasmid pC4.Plasmid pSV2neo contains dominant selectable marker and derives from the neo gene of Tn5, and this gene code can make it to one group of antibiotic, comprises that G418 has the enzyme of resistance.This cell inoculation is nourished among the α minus MEM that is filled with 1ug/ml G418.After 2 days, this cell of trypsinization, and inoculation advance hybridoma cloning flat board (Greiner, Germany) in, this flat board added be supplemented with 10,25 or the 50ng/ml methotrexate add the α minus MEM of 1ug/ml G418.About after 10-14 days, the single clone of trypsinization, and inoculate the 6 hole culture dishs or the 10ml that advance to contain variable concentrations methotrexate (50nM, 100nM, 200nM, 400nM, 800nM) subsequently and shake in the bottle.Subsequently, the clone who grows under the maximum concentration methotrexate condition is transferred in the new 6 hole flat boards that contain taller concentration methotrexate (1mM, 2mM, 5mM, 10mM, 20mM).Repeating same steps as can be till the clone who grows under the 100-200mM concentration conditions until acquisition.Analyze the expression of expection gene outcome, for example, by SDS-PAGE and Western blotting, or by the reversed-phase HPLC analysis.
Embodiment 2: the limiting examples of CH1 disappearance analogue body of the present invention
Background:EMP-1 (EPO simulating peptide-1) is the peptide that 20 amino acid longs are arranged, and (HuEPO) do not have sequence homology with human erythropoietin, but it has ability (as dimer) (the Wrighton et al. of activation EPO receptor, 1996, Science, vol.273,458-463).But, its low relatively activity (lower by 10 than HuEPO, 000-100,000 times) and short half-life (vitro half-lives in 50% serum is 8 hours, half-life the unknown in the body) have been limited its effectiveness aspect treatment.Therefore, need a kind of give this peptide than the long half-lift, do not disturb it to tire, and may promote the method that it is tired.For realizing this target, people make some trials, by stablizing the dimerization of this peptide, and with the activity of raising EMP-1, or by this peptide is integrated into than in the macrostructure, to prolong its half-life.People such as Wrighten (1997, Nature Biotechnology, vol.15 1261-65) with biotin labeling EMP-1 and Succ-PEG-DSPE associating, thereby makes dimerization stable.They find that activity has improved 100 times in the body outer cell proliferation experiment.They have also adopted anti-biotin antibodies, stablizing this peptide dimer, but find that activity has only improved 10 times.They have also prepared and chemically are being defined as dimeric EMPP-1.In this situation, the activity in vivo that is observed has improved 100 times.Another research group is attempted by making EMP-1 and Polyethylene Glycol (PEG) covalently bound, to improve activity (Johnson et al., 1997, the Chem.﹠amp of EMP-1; Bio., vol.4 (12), 939-50).The tiring and improved of their report up to 1000 times, but find this construct in mice, have immunogenicity (this antibody is at this peptide) (Dana Johnson, Personalcommunications).People such as Kuai (2000, J.Peptide Res., vol.56 59-62) inserts the EMP-1 peptide in the sequence of plasminogen activator inhibitor (PAI-1).After finding that EMP-1 is inserted in this supporting structure, both can stablize dimerization, but also prolong half-life.In the experiment, tiring of this construct is found than 2500 times of this heights of EMP-1 in vivo.Should be pointed out that and adopted different experiment in vitro and body inner model in the above-mentioned research, and report tire each other or therewith the place introduce the result and can not compare.
EMP-1CH1 disappearance analogue body of the present invention
A specific limiting examples of the present invention is the EMP-NfusCG1 construct, wherein V is a natural junior three aminoacid that has antibody, Pep is single copy of the EMP-1 peptide of biologically active, Flex is that the series connection of Gly-Gly-Gly-Ser flexible joint repeats, V2 is the J district of the natural IgG of existence, pHinge is complete IgG1 hinge region, and CH2﹠amp; CH3 belongs to IgG1 isotype subclass.This structure is considered to the peptide with constraint EMP-1, but also has enough flexibilities, can make as the dimerization of the peptide of this assembling homodimer part stable.In the body outer cell proliferation experiment, the specific activity EMP-1 peptide of EMP-NfusCG1 is high 550 times approximately, and only than the low 4-6 of reorganization HuEPO (rHuEPO) doubly, thereby confirmed above-mentioned viewpoint.In addition, it should be rHuEPO or the manyfold of EMP-1 peptide half-life itself that the half-life of this construct is considered to, and approximate with the half-life of I gG.Consistent therewith, compare with the mice of handling via the normal mouse that EMP-Nfus CG1 handles via rHuEPO, the former has obtained remarkable higher maximum hematocrit, and when one timing of isoreactivity unit, the rising level has been kept the long period.As if this construct is secreted from cell effectively, and suitably folded; Overcome a difficult problem relevant with first generation analogue body.
Except that above-mentioned base structure, this paper has also described the variant with potential favourable biological property.The construct that these variants comprise that tendency may reduce from connecting, immune effector miopragia or immunogenicity reduce.The present invention has also looked forward to other can give special characteristic, such as the configuration of the peptide that can improve biologically active and the modification that can cross over blood brain barrier.This suggestion variant and modification can be combined in any way, to obtain construct with given activity.
Utilize the recombinant DNA method, the EMP-1 peptide is inserted in the intermediate carrier, between immunoglobulin signal peptide and human J sequence.Complementary synthetic oligonucleotide has been adopted in this operation, and the BstI 107I in its end and this carrier is compatible with the KpnI restriction site
(5 ' TACAGGCCCAGATCCAGGGCGGTACCTACAGCTGCCACTTCGGGCCCCTCACGTGG GTGTGCAAGCCCCAGGGCGGCGGAAGCGGGGGAGGCTCCGGTAC3 ' (SEQ IDNO:43) and 3 ' CGGAGCCTCCCCCGCTTCCGCCGCCCTGGGGCTTGCACACCCACGTGAGGGGCCCG AAGTGGCAGCTGTAGGTACCGCCCTGGATCTGGGCCTGTA5 ') (SEQID NO:44).These oligonucleotide contain coded sequence, the EMP-1 peptide in the total site of signal peptidase and the flexible joint that is repeated to form by two Gly-Gly-Gly-Ser.The XbaI restricted fragment that contains function element is transferred in the expression vector subsequently.This carrier contains coded sequence, HC constant region 2 (CH2) and the constant region 3 (CH3) of anti-CD4 immunoglobulin promoter and enhancer, IgG 1 hinge region, also contain plasmid replication in the antibacterial and screening, and for the necessary element of screening of stably express in the mammalian cell.
This plasmid is by line styleization, and is imported in the NSO mouse myeloma cell line by electroporation.The screening resisting cell, and, identify high expressed of EMP-NfusCG1 by the elisa assay culture supernatant.Obtain this construct by standard A albumen affinity chromatography purification from cell culture supernatant.Under degeneration and reducing condition, make purified product by containing the polyacrylamide gel of SDS, have desired size to confirm this purified product.Further identify this purifying protein by mass spectral analysis and N-terminal order-checking.
The aminoacid sequence of EMP-NfusCG1 is as follows.Functional domain is the coded sequence of above-mentioned peptide.Three amino acid signal peptide consensus sequences are corresponding with junior three aminoacid of naturally occurring immunoglobulin.Thereby these aminoacid are considered to facilitate effective removal to signal peptide by the signal peptidase in the endoplasmic reticulum.After the EMP-1 coded sequence is located immediately at this sequence.2 C-terminal aminoacid of EMP-l sequence combine with 6 aminoacid of adjacency, have formed and have been characterized as the multiple flexible joint of Gly-Gly-Gly-Ser.Human (J) region sequence that connects is immediately following thereafter.The J sequence is considered to the flexibility that can provide bigger, makes the EMP-1 dimer present correct configuration, and makes this dimer outstanding from the spherical structure of immunoglobulin, and deeply to the breach between two EPO receptors.This construct also comprises the HC hinge region, and the J district is located immediately at thereafter.Contain three cysteine in the IgG1 hinge region (highlighted part).First cysteine usually matches with light chain immunoglobulin (LC), and two other cysteine has then participated in the interchain key between two HC.The remainder of this sequence is by CH2﹠amp; The CH3 district forms, and has constituted this proteic filling part.It is the bonded ability of itself and FcRn that immunoglobulin is considered to have one of reason of long serum half-life, and this FeRn can be by returning to extracellular space with the pinocytosis immunoglobulin, thereby prolong serum half-life.The junction in the binding site of FeRn and CH2 and CH3 district overlapping (Sheilds et al., 2O01, J.Bio.Chem., vol.276 (9), 6591-6604).
The peptide sequence of BMP-NfusCG1 has shown the important function territory.
Signal peptidase
Consensus sequence J district
～～～joint～～～～～～～～
The EMP-1 peptide～～～～～～the huG1 hinge
～～～～～～～～～～～～～～～～～～～～
～～～～～～～～～～～～～～～～～～～～～～～
1QIQGGTYSCHFGPLTWVCKPQGGGSGGGSGTLVTVSSEPKSCDKTHTCPPCPAPELLGGP
IgG1CH2
61SVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNS
IgG1CH2
～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～IgG1CH3
～～～～～～～～～～～～～～～～～
TYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRD
IgG1CH3
～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～
181
TKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSR
WQ
IgG1CH3
～～～～～～～～～～～～～～～
241QGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO：50)
As everyone knows, two IgG heavy chains during cell processing by hinge region in disulfide bond between two cysteine be assembled in together, formed homodimer.It is believed that this phenomenon also can occur between the modified peptides, thereby form the EMP-NfusCG1 construct of assembling.In addition, it is believed that between two cysteine in the EMP-1 peptide and also will form interchain disulfide bond.The expected structure of EMP-NfusCG1 contains two EMP-1 peptides.This peptide is in the spatial arrangements of N-terminal, and the flexibility of contiguous sequence makes this peptide form the dimer of biologically active.
At first in the external biological determination of activity, check the activity of EMP-NfusCG1.For this mensuration, adopted acute megakaryoblastic leukemia patient source EPO dependency UT-7/EPO cell line (Komatsu et al., 1993, Blood, vol.82 (2), 456-464).Take out these cells after 48-72 hour from the culture medium that is supplemented with rHuEPO, they just begin to experience programmed cell death.If under rHuEPO shortage condition, cultivated 24 hours the cell process rHuEPO or the processing of EPO synergist, just can be saved.EMP-NfusCG1 is added in the cell hungry under the no rHuEPO condition, and adopting tetrazotized zole compound MTS (CellTiter 96Aqueous One Solution, Promega) handled back 48 hours, measure cell viability, wherein this tetrazotized zole compound MTS can be by the living cells metabolism, and produces the product that can measure absorbance.The result of model experiment shows that the potency ratio EMP-1 peptide of mole foundation EMP-NfusCG1 is high 500 times, than low 5 times of rHuEPO.In addition, adopt EMP-NfusCG1 to stimulate above-mentioned same cell, and by making cell lysate carry out polyacrylamide gel electrophoresis, with range estimation tyrosine phosphorylation pattern.The shown pattern of EMP-NfusCG1 is similar to rHuEPO, and it is similar to rHuEPO to illustrate that EMP-NfusCG1 acts on the mechanism of above-mentioned cell.
Carry out studying in the body in normal mouse, relatively the two half-life of EMP-NfusCG1 and rHuEPO, and comparison, the two was to erythropoietic influence.Preliminary study shows uses continuous two days dosage but not during single dose to mice, more remarkable to replying of rHuEPO.Find also that in preliminary study when rHuEPO dosage was 2000U/kg, the hematocrit of mice rose to maximum at the 4th day, then fell near the baseline values in the 7th day.For the two difference on effect of relatively EMP-NfusCG1 and rHuEPO, test the activity of being measured, the dosage of normalization EMP-NfusCG1 based on the external biological determination of activity; Because the activity of EMP-NfusCG1 is lower than 5 times in this experiment, thereby gives the EMP-NfusCG1 of about 5 times of volumes.Also considered the difference on the molecular weight (rHuEPO=30,400 dalton, EMP-NfusCG1=62,200 dalton).When based on above-mentioned consideration, when mice was used same dose, EMP-NfusCG1 had caused higher maximum and has replied, and compares with rHuEPO, and this is replied and has been extended.
Measure the serum-concentration of rHuEPO and EMP-NfusCG1 by ELISA.In to the sensitive mensuration of rHuEPO concentration that is low to moderate 2.5mU/ml,, in sample, do not find positive signal with dilution in 1: 8 since the 4th day.This result is consistent with the 2-8 hour half-life that rHuEPO is expected in rodent.(PRI Export，Part IC2：T0xico-Pharmacologica1Documentation)。By comparison, the 4th day detected EMP-NfusCG1 level is up to 1.5ug/mL, and this high level is retained to the research end.The clearance rate scattergram is linear unusual among Fig. 6.May be because first time point was injected back 3 days the last time, before getting first sample, just reach Cmax (C Max), therefore do not observe the peak value of exponential curve.Absorption that another kind of explanation is this novel constructs and removing feature make its clearance rate scattergram present linearity.Based on these data, can calculate its half-life and be about 10.8 days, be the several times of rHuEPO half-life.
Except EMP-NfusCG1, the present invention has also prepared three similar constructs, and the three is at the isotype in hinge CH2 and CH3 district, or there are differences aspect the flexible number of elements in that the EMP-1 peptide is had.One of them construct is called as EMP-NfusCG4, and except that its hinge and constant region belong to the G4 hypotype, this construct remainder is all consistent with EMP-NfusCG1.Specifically, the XbaI restricted fragment that will be used for making up the EMP-NfusCG1 expression plasmid shifts into specific support, this carrier provides coded sequence, CH2 and the CH3 of anti-CD4 immunoglobulin promoter and enhancer, IgG 4 hinge regions, and to the necessary element of screening of stably express in plasmid replication and screening and the mammalian cell in the thin mattress.The expression of this plasmid is similar to EMP-NfusCG1 with the purification mode.The coded sequence of EMP-NfusCG4 is as follows.
The HuG4 hinge
The sequence consistent with BMP-NfusCG1～～～～～～～～～～～～～～～～～～～～
～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～
l QIQGGTYSCHFGPLTWVCKPQGGGSGGGSGTLVTVSSESKYGPPCPSCPAPEFLGGPSVF
IgG4CH2
～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～
61LFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEOFNSTYR
IgG4CH3
IgG4CH2
～～～～～～～～～～～～～～～～～～～～～
～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～
12l WSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYTLPPSQEEMTKN
IgG4CH3
～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～～
～～
181QVSLTCLVKGFYPSDIAVEWESNGOPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGN
IgG4CH3
～～～～～～～～～～～～～～～～～～～～～～～～～～
24l VFSCSVMHEALHNHYTQKSLSLSLGK SEQ ID NO：51
Hereinafter shown the comparative result between the some examples of CH1 disappearance analogue body:
QIQGGTYSCHFGPLTWVCKPQGGGSGGGSGTLVTVSSEPKSCDKTH-TCPPCP APELL
MG1---......................------------.....................
CG4......................................S.YGPP----..S.....F.
MG4---......................------------.S.YGPP----..S.....F.
muCG2a.......................................RGPTIKPCPPCK....N..
Significant differences between IgG1 and the IgG4 hypotype is the quantity of cysteine in the hinge region.Similar with the G1 hypotype, there are two cysteine that participated in disulfide bond between the heavy chain in the IgG4 hinge.Do not contain the cysteine that is usually directed to the disulfide-bonded of LC in the G4 hinge; This cysteine is arranged in the G4CH1 district that above-mentioned construct lacks.The IgG4 hinge region not as the IgG1 hinge pliable and tough, this may influence the dimeric potential configuration of EMP-NfusCG1 (Dangl et al., 1997, EMBO Journal, vol 7 (7), 1989-94).In addition, two isotypes there are differences aspect the ability of the cytotoxicity (ADCC) of its mediation complement dependent form cytotoxicity (CDC) and antibody dependent type cell.The G1 hypotype is the strong inducer of complementary cascade, and the G4 hypotype almost non-activity (Dangl et al., 1997, EMBO Journal, vol 7 (7), 1989-94).In addition, the G1 hypotype combines with high-affinity with the Fc receptor, thereby has facilitated cell-mediated cytotoxicity, and faint (the Allan ﹠amp of the adhesion of G4 hypotype; Seed, 1989, Science, vol.423,378-80).Owing to need make target cell propagation, so the G4 hypotype is ideal in the relative nullity aspect the activating effect subfunction.The binding site of FcRn scavenger receptor is present on the IgG4, and since its in conjunction with aspect as if do not have any IgG subclass difference, so its pharmacokinetics is considered to similar (West etal. to the G1 hypotype, 2000, Biochem., vol 39,9698-9708).
The same with the situation of G1, thus construct EMP-NfusCG4 is considered to and can forms homodimer by means of two cysteine in the hinge region.As mentioned above, do not contain the 3rd cysteine in the G4 hinge, thereby negated the probability that forms the 3rd disulfide bond between two chains.The dimeric configuration of EMP-1 also may be because the difference on the size of hinge region and the flexibility and different.Consider above-mentioned difference, it is similar to EMP-NfusCG4 that the universal architecture of EMP-NfusCG4 is considered to.
As previously mentioned, in external biological determination of activity experiment, check EMP-NfusCG4.Experiment carries out twice, and the result is similar.During this is measured, low 13 times of the potency ratio EMP-NfusCG1 of EMP-NfusCG4, but mean difference is about 10 times.Because low 5 times of the potency ratio rHuEPO of EMP-NfusCG1, so the difference between EMP-NfusCG4 and the rHuEPO is considered to should be about 50 times.Actual variance in this experiment is 60 times.Difference in this experiment between EMP-NfusCG4 and the EMP-1 is 40 times, but because the EC of EMP-1 50Low 2 times of numeric ratio history average is so the actual variance between EMP-NfusCG4 and the EMP-1 may be bigger.The activity that EMP-NfusCG4 is observed is lower, but attribution is the difference in the hinge region, and this difference makes the EMP-1 dimer can not form optimum configuration.Checked the activity of EMP-NfusCG4 so far not yet in vivo.
Other two kinds of constructs are designed to study active necessary functional domain.Therefore, human J district and a Gly-Gly-Gly-Ser repetition between EMP-1 sequence and hinge region, have been ignored.Difference between above-mentioned " complete " version and these minimal version is isotype subclass (G1 contrasts G4), CH2 and the CH3 district of hinge region.
Signal peptidase
Consensus sequence J district
～～～joint～～～～～～～～
The EMP-1 peptide～～～～
～～～～～～～～～～～～～～～～～～～～
Complete QIQGGTYSCHFGPLTWVCKPQGGGSGGGSGTLVTVSS..HingeSEQ ID NO:52
Minimum QIQGGTYSCHFGPLTWVCKPQGGGS..Hinge SEQ ID NO:53
For being designated as the minimal version of EMP-NfusMG1 and EMP-NfusMG4, the structure of expression plasmid is to be achieved by synthetic oligonucleotide is inserted intermediate carrier, its method identical with the method at the expression plasmid of EMP-NfusCG1 and EMP-NfusCG4 of making up ( 5 'TACAGGCCCAGGGCGGTACCTACAGCTGCCACTTCGGGCCCCTCACGTGGGTGTGC AAGCCCCAGGGCGGCGGATCAGGTAAGTT 3 '(SEQ ID NO:45) and 3 'CTAGAACTTACCTGATCCGCCGCCCTGGGGCTTGCACACCCACGTGAGGGGGCCCG AAGTGGCAGCTGTAGGTACCGCCCTGGGCCTGTA 5 ') (SEQIDNO:46).Coded sequence, EMP-1 peptide and single Gly-Gly-Gly-Ser that above-mentioned synthetic oligonucleotide contains the total site of signal peptidase repeat.But, in this case, oligonucleotide 3 ' terminal compatible with the XbaI restriction site, different with the oligonucleotide that is used to make up full release.In the Xba I site in downstream, intermediate carrier J district, ignored the coded sequence in J district by clone's carry.The structure of final expression plasmid is to adopt as realizing at EMP-NfusCG1 and the described method of EMP-NfusCG4.Because technical problem, so far, only EMP-NfusMG4 has obtained to express and purification.The structure of EMP-NfusMG4 be considered to have the EMP-1 peptide from the spherical structure of this protein form outstanding, should outstanding be considered to not as giving prominence to significantly in the EMP-NfusCG4 situation, owing to this reason, the EMP-1 peptide dimer may not be deeply to the breach between the EPO receptor.In addition, the flexibility that helps the EMP-1 peptide to form best dimer configuration may be lowered.This phenomenon particularly important in people's such as Livnah observed result (1998, Nat.Strust.Biol., vol.5,993-1004).They find that peptide (EMP-33) can make two EPO receptor dimerizationizations, but can not activate this receptor.By the two crystal structure of relatively EMP-33/EPO receptor complex and EMP-1/EPO receptor complex, they find that at the fine difference on the dimerization receptor orientation this peptide being activated on its receptor ability there are differences.Therefore, if the EMP-1 dimer can not form optimum configuration, it also may not make the EPO receptor dimerizationization, perhaps receptor can not be brought in the activation configuration.
The activity of check EMP-NfusMG4 construct in UT7 external biological determination of activity experiment.This experimental result as shown in figure 12.This experiment has only been carried out once, because the changeableness of biological activity determination, so its result should be considered to PRELIMINARY RESULTS.In this experiment, low 227 times of the potency ratio rHuEPO of EMP-NfusMG4 is higher 44 times than EMP-1 peptide.But the EC of rHuEPO in should testing 50Than low 3 times of history average, so the difference between EMP-NfusMG4 and the rHuEPO may be lower significantly.This experiment does not directly contrast between EMP-NfusCG4 and EMP-NfusMG4.Average EC based on EMP-NfusCG4 50, promptly 3.5 * 10 -10, and the contrast between the EC50 of the EMP-NfusMG4 that observes in this experiment, difference is 2 times as can be known.Though The above results is a PRELIMINARY RESULTS, illustrate that human J sequence and the multiple shortage of Gly-Gly-Gly-SeT can not reduce tiring of EMP-NfusMG4 construct basically.Whether still unknown this observed result can be extended to EMP-NfusMG1 construct.
For avoiding undesirable activity, studied at present and expressed other construct that exists single or multiple aminoacid to change.These variations can be expressed separately, and perhaps the form with multiple variation combination is present in the single construct.Usually the cysteine that participates in disulfide bond between HC and the LC will be changed into alanine.Though this cysteine may pass through to form the 3rd disulfide bond, thereby relates to the stabilisation of construct, it also might form disulfide bond singularly with other cysteine in the construct, perhaps forms disulfide bond between two constructs.By removing this cysteine, can strengthen correct folding and assembling, and reduce from the probability that connects.
There are some researches show, in the low hinge region of IgG1, from two lysines (L) residue, L234﹠amp; L235 to the sudden change of alanine (A) will eliminate the cytotoxicity (ADCC) of immunoglobulin-mediated complement dependent form cytotoxicity (CDC) and antibody dependent type cell ability (Hezerehetal., 2001, J.Virol., vol.75 (24), 12161-68).Preliminary study shows that EMP-NfusCG1 can not mediate the complement cracking of the cell that can express the EPO receptor.This may be found on the erythroid progenitor cell acceptor quantity of existence low due to.In addition, may not have functional dependency by amplification explanation immunological effect subfunction in the body that significantly improves the erythroid progenitor cell that is confirmed of hematocrit.But, though do not observe the influence relevant with effector function, still can be with the step of said mutation introduction for preventing.
Can cause the another kind modification of the mediation reduction of immunological effect subfunction is to remove the glycosylation attachment site.This modification can be achieved by making agedoite (N297) on 297 sport glutamine (Q).Other variation can randomly comprise threonine (T) is substituted by optional amino acid, to reduce or modification O-glycosylation, for example, the T34 of the glycosylation version of the known IgG1 of having subclass or T47 are weak mediator (the Jefferis et al. of immunological effect subfunction, 1998, Immol.Rev., vol.163,50-76).
Figure A0382011000911
It more than is the suggestion sudden change in EMP-NfusCG1 hinge and the CH2 coded sequence.(SEQ ID NO：47)
It is the same area that IgG1CH2 and CH3 district with EMP-NfusCG1 are substituted by I gG4 hypotype that the another kind that just is being studied is at present modified, and keeps the G1 hinge region simultaneously.As previously mentioned, the ability of IgG4 subclass mediation immunological effect subfunction is far below the G1 subclass.Also as previously mentioned, it is theorized that the difference in two IgG subclass between the hinge region flexibility may be the reason that has activity difference between EMP-NfusCG1 and the EMP-NfusCG4.Therefore this construct may keep the activity of EMP-NfusCG1, and does not relate to potential immunological effect subfunction.
Other modification of being looked forward to is to reduce the potential immunogenic modification of this construct.The peptide that an immunogenic important determiner is a protein source by the MHC molecule effectively in conjunction with and present cell to T, and excite based on the immunne response of cell or help the ability of the T cell of antibody response.Utilize general in the MHC combination, and based on algorithm (SYFPETHI, the Ramensee et al. of network, 1999, Immunogenetics, vol.50,213-19 and BIMAS), analyze potential MHC in the N-terminal zone that contains EMP-1 among the EMP-NfusCG1 in conjunction with epi-position.The immunogenic sudden change of prediction that can reduce one or more peptides can be evaluated as influencing in the immunogenic body.
(SEQ ID NO:48): the predicted and bonded peptide of MHC I quasi-molecule
QIQGGTYSCHFGPLTWVCKPQGGGSGGGSGTLVTVSSEPKSCDKTHTC
PPC
IQGGTYSCHF
GTYSCHFGPL
TYSCHFGPL
YSCHFGPLTW
CHFGPLTWV
GSGGGSGTL
SGGGSGTLV
GGGSGTLVTV
GGSGTLVTV
TLVTVSSEPK
LVTVSSEPK
SSEPKSCDKT
Predicted and the bonded peptide of MHC II quasi-molecule (SEQ ID NO:49) of SSEPKSCDK
QIQGGTYSCHFGPLTWVCKPQGGGSGGGSGTLVTVSSEPKSCDKTHTC
PPC
FGPLTWVCKPQGGGS
LTWVCKPQGGGSGGG
SGTLVTVSSEPKSCD
SCHFGPLTWVCKPQG
GGGSGTLVTVSSEPK
GTLVTVSSEPKSCDK
VTVSSEPKS
The limiting examples of other sequence comprises:
No QIQ:
(SEQ ID NO：62)
GGTYSCHPGPLTWVCKPQGGGSGGGSGTLVTVSSEPKSCDKTHTCPPCPAPELL
The sudden change of two cysteine among the EMP1:
(SEQ ID NO：63)
(QVQ)GGTYSSHFGPLTWVSKPQGGGSGGGSGTLVTVSSEPKSADKTHTCPPCPAPEAA
The sudden change of Leu56 and Leu57:
(SEQ ID NO：64)
(QIQ)GGTYSCHFGPLTWVCKPQGGGSGGGSGTLVTVSSEPKSADKTHTCPPCPAPEAA
(SEQ ID NO：65)
(QVQ)GGTYSCHFGPLTWVCKPQGGGSGGGSGTLVTVSSEPKSADKTHTCPPCPAPEAA
(SEQ ID NO：66)
(QVQ)GGTYSCHFGPLTWVCKPQGGGSGGGSGTLVTVSSEPKSADKTHTCPPCPAPELA
(SEQ ID NO：67)
(QVQ)GGTYSCHFGPLTWVCKPQGGGSGGGSGTLVTVSSEPKSADKTHTCPPCPAPEAL
(SEQID NO：68)
(QVQ)GGTYSCHPGPLTWVCKPQGGGSGGGSGTLVTVSSEPKSADKTHTCPPCPAPELE
IgG4 with IgG1 hinge region:
(SEQ ID NO：69)
(QIQ)GGTYSCHFGPLTWVCKPQGGGSGGGSGTLVTVSSEPKSSDKTHTCPPCPAPEFL
G1-G4IgG4 with different joint length:
(SEQ ID NO：70)
(QIQ)GGTYSCHFGPLTWVCKPQGGGSGTLVTVSSESKYGPPCPSCPAPEFL
(SEQ ID NO：71)
(QIQ)GGTYSCHFGPLTWVCKPQGGGSGGGSGTLVTVSSESKYGPPCPSCPAPEFL
(SEQ ID NO：72)
(QIQ)GGTYSCHFGPLTWVCKPQGGGSGGGSGGGTLVTVSSESKYGPPCPSCPAPEFL
(SEQ ID NO：73)
(QIQ)GGTYSCHFGPLTWVCKPQGGGSGGGSGGGSGTLVTVSSESKYGPPCPSCPAPEFL
IgG1 with truncate hinge region:
(SEQ ID NO：74)
(QVQ)GGTYSCHFGPLTWVCKPQGGGSGGGSGTLVTVSSCPPCPAPELL
(SEQ ID NO：75)
(QVQ)GGTYSCHFGPLTWVCKPQGGGSGGGSGTLVTVSSTHTCPPCPAPELL
(SEQ ID NO：76)
(QVQ)GGTYSCHFGPLTWVCKPQGGGSGGGSGTLVTVSSAKDTHTCPPCPAPELL
The IgG1 that the O-glycosylation site is knocked out:
(SEQ ID NO：77)
(QIQ)GGTYSCHFGPLTWVCKPQGGGSGGGSGTLVTVSSEPKSADKTHACPPCPAPELL
(SEQ ID NO：78)
(QIQ)GGTYSCHFGPLTWVCKPQGGGSGGGSGTLVAVSSEPKSADKTHTCPPCPAPELL
(SEQ ID NO：79)
(QIQ)GGTYSCHFGPLTWVCKPQGGGSGGGSGTLVTVSSEPKSADKAHTCPPCPAPELL
(SEQ ID NO：80)
(QIQ)GGTYSCHFGPLTWVCKPQGGGSGGGSGTLVAVSSEPKSADKTHACPPCPAPELL
Advantage:Novel constructs, promptly above-mentioned EMP-NfusCG1 provide an optional approach of showing the peptide EMP-1 of biologically active.The activity of this construct is in the scope of rHuEPO, and the half-life of interior half-life of its body and IgG is approximate.In addition, suggestion is modified and is considered to can be combined in together, and the new feature of EMP-NfusCG1, thereby strengthens the effectiveness of EMP-NfusCG1 construct.
Should be understood that and to adopt other method different to put into practice the present invention with specific descriptions method in above-mentioned explanation and the example.
According to foregoing, many improvement of the present invention and variation all are possible, thereby also in category of the present invention.
Sequence table
<110>Heaver，George A.；Knight，David M.；Ghrayeb，John；Scallon，Bernard J.；Nesspor，Thomas C.；Centocor，Inc.
＜120〉mammalian EPO mimetic CH 1 deleted mimetibodies, compositions, method and purposes
<130>CEN0311
<140>10/609,783
<141>2003-06-30
<150>60/412,144
<151>2002/09/12
<160>80
<170>PatentIn version 3.3
<210>1
<211>14
<212>PRT
＜213〉artificial
<220>
＜223〉peptide
<220>
＜221〉comprehensive characteristics
<222>(2)..(2)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(4)..(5)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(8)..(8)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(11)..(11)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(13)..(13)
＜223〉Xaa can be any naturally occurring aminoacid
<400>1
Tyr Xaa Cys Xaa Xaa Gly Pro Xaa Thr Trp Xaa Cys Xaa Pro
1 5 10
<210>2
<211>28
<212>PRT
＜213〉artificial
<220>
＜223〉peptide
<220>
＜221〉comprehensive characteristics
<222>(2)..(2)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(4)..(5)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(8)..(8)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(11)..(11)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(13)..(13)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(16)..(16)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(18)..(19)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(22)..(22)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(25)..(25)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(27)..(27)
＜223〉Xaa can be any naturally occurring aminoacid
<400>2
Tyr Xaa Cys Xaa Xaa Gly Pro Xaa Thr Trp Xaa Cys Xaa Pro Tyr Xaa
1 5 10 15
Cys Xaa Xaa Gly Pro Xaa Thr Trp Xaa Cys Xaa Pro
20 25
<210>3
<211>28
<212>PRT
＜213〉artificial
<220>
＜223〉peptide
<220>
＜221〉comprehensive characteristics
<222>(2)..(2)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(4)..(5)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(8)..(8)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(11)..(11)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(13)..(13)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(16)..(16)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(18)..(19)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(22)..(22)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(25)..(25)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(27)..(27)
＜223〉Xaa can be any naturally occurring aminoacid
<400>3
Tyr Xaa Cys Xaa Xaa Gly Pro Xaa Thr Trp Xaa Cys Xaa Pro Tyr Xaa
1 5 10 15
Cys Xaa Xaa Gly Pro Xaa Thr Trp Xaa Cys Xaa Pro
20 25
<210>4
<211>28
<212>PRT
＜213〉artificial
<220>
＜223〉peptide
<220>
＜221〉comprehensive characteristics
<222>(2)..(2)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(4)..(5)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(8)..(8)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(11)..(11)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(13)..(13)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(16)..(16)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(18)..(19)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(22)..(22)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(25)..(25)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(27)..(27)
＜223〉Xaa can be any naturally occurring aminoacid
<400>4
Tyr Xaa Cys Xaa Xaa Gly Pro Xaa Thr Trp Xaa Cys Xaa Pro Tyr Xaa
1 5 10 15
Cys Xaa Xaa Gly Pro Xaa Thr Trp Xaa Cys Xaa Pro
20 25
<210>5
<211>20
<212>PRT
＜213〉people
<400>5
Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp Val Cys Lys
1 5 10 15
Pro Gln Gly Gly
20
<210>6
<211>20
<212>PRT
＜213〉people
<400>6
Gly Gly Asp Tyr His Cys Arg Met Gly Pro Leu Thr Trp Val Cys Lys
1 5 10 15
Pro Leu Gly Gly
20
<210>7
<211>20
<212>PRT
＜213〉people
<400>7
Gly Gly Val Tyr Ala Cys Arg Met Gly Pro Ile Thr Trp Val Cys Ser
1 5 10 15
Pro Leu Gly Gly
20
<210>8
<211>20
<212>PRT
＜213〉people
<400>8
Val Gly Asn Tyr Met Cys His Phe Gly Pro Ile Thr Trp Val Cys Arg
1 5 10 15
Pro Gly Gly Gly
20
<210>9
<211>20
<212>PRT
＜213〉people
<400>9
Gly Gly Leu Tyr Leu Cys Arg Phe Gly Pro Val Thr Trp Asp Cys Gly
1 5 10 15
Tyr Lys Gly Gly
20
<210>10
<211>20
<212>PRT
＜213〉people
<400>10
Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp Val Cys Lys
1 5 10 15
Pro Gln Gly Gly
20
<210>11
<211>40
<212>PRT
＜213〉people
<400>l1
Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp Val Cys Lys
1 5 10 15
Pro Gln Gly Gly Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr
20 25 30
Trp Val Cys Lys Pro Gln Gly Gly
35 40
<210>12
<211>23
<212>PRT
＜213〉people
<400>12
Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp Val Cys Lys
1 5 10 15
Pro Gln Gly Gly Ser Ser Lys
20
<210>13
<211>23
<212>PRT
＜213〉people
<400>13
Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp Val Cys Lys
1 5 10 15
Pro Gln Gly Gly Ser Ser Lys
20
<210>14
<211>23
<212>PRT
＜213〉people
<400>14
Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp Val Cys Lys
1 5 10 15
Pro Gln Gly Gly Ser Ser Lys
20
<210>15
<211>46
<212>PRT
＜213〉people
<400>15
Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp Val Cys Lys
1 5 10 15
Pro Gln Gly Gly Ser Ser Lys Gly Gly Thr Tyr Ser Cys His Phe Gly
20 25 30
Pro Leu Thr Trp Val Cys Lys Pro Gln Gly Gly Ser Ser Lys
35 40 45
<210>16
<211>23
<212>PRT
＜213〉people
<400>16
Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp Val Cys Lys
1 5 10 15
Pro Gln Gly Gly Ser Ser Lys
20
<210>17
<211>10
<212>PRT
＜213〉artificial
<220>
＜223〉peptide
<220>
＜221〉comprehensive characteristics
<222>(2)..(3)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(6)..(6)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(9)..(9)
＜223〉Xaa can be any naturally occurring aminoacid
<400>17
Cys Xaa Xaa Gly Pro Xaa Thr Trp Xaa Cys
1 5 10
<210>18
<211>19
<212>PRT
＜213〉people
<400>18
Gly Gly Thr Tyr Ser Cys His Gly Pro Leu Thr Trp Val Cys Lys Pro
1 5 10 15
Gln Gly Gly
<210>19
<211>19
<212>PRT
＜213〉people
<400>19
Val Gly Asn Tyr Met Ala His Met Gly Pro Ile Thr Trp Val Cys Arg
1 5 10 15
Pro Gly Gly
<210>20
<211>18
<212>PRT
＜213〉people
<400>20
Gly Gly Pro His His Val Tyr Ala Cys Arg Met Gly Pro Leu Thr Trp
1 5 10 15
Ile Cys
<210>21
<211>18
<212>PRT
＜213〉people
<400>21
Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp Val Cys Lys
1 5 10 15
Pro Gln
<210>22
<211>20
<212>PRT
＜213〉people
<400>22
Gly Gly Leu Tyr Ala Cys His Met Gly Pro Met Thr Trp Val Cys Gln
1 5 10 15
Pro Leu Arg Gly
20
<210>23
<211>22
<212>PRT
＜213〉people
<400>23
Thr Ile Ala Gln Tyr Ile Cys Tyr Met Gly Pro Glu Thr Trp Glu Cys
1 5 10 15
Arg Pro Ser Pro Lys Ala
20
<210>24
<211>13
<212>PRT
＜213〉people
<400>24
Tyr Ser Cys His Phe Gly Pro Leu Thr Trp Val Cys Lys
1 5 10
<210>25
<211>11
<212>PRT
＜213〉people
<400>25
Tyr Cys His Phe Gly Pro Leu Thr Trp Val Cys
1 5 10
<210>26
<211>10
<212>PRT
＜213〉artificial
<220>
＜223〉peptide
<220>
＜221〉comprehensive characteristics
<222>(1)..(3)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(6)..(6)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(9)..(10)
＜223〉Xaa can be any naturally occurring aminoacid
<400>26
Xaa Xaa Xaa Gly Pro Xaa Thr Trp Xaa Xaa
1 5 10
<210>27
<211>12
<212>PRT
＜213〉artificial
<220>
＜223〉peptide
<220>
＜221〉comprehensive characteristics
<222>(2)..(5)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(8)..(8)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(11)..(12)
＜223〉Xaa can be any naturally occurring aminoacid
<400>27
Tyr Xaa Xaa Xaa Xaa Gly Pro Xaa Thr Trp Xaa Xaa
1 5 10
<210>28
<211>14
<212>PRT
＜213〉artificial
<220>
＜223〉peptide
<220>
＜221〉comprehensive characteristics
<222>(1)..(1)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(3)..(6)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(9)..(14)
＜223〉Xaa can be any naturally occurring aminoacid
<400>28
Xaa Tyr Xaa Xaa Xaa Xaa Gly Pro Xaa Xaa Xaa Xaa Xaa Xaa
1 5 10
<210>29
<211>16
<212>PRT
＜213〉artificial
<220>
＜223〉peptide
<220>
＜221〉comprehensive characteristics
<222>(1)..(1)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(3)..(3)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(5)..(6)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(9)..(9)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(12)..(12)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(14)..(16)
＜223〉Xaa can be any naturally occurring aminoacid
<400>29
Xaa Tyr Xaa Cys Xaa Xaa Gly Pro Xaa Thr Trp Xaa Cys Xaa Xaa Xaa
1 5 10 15
<210>30
<211>20
<212>PRT
＜213〉people
<400>30
Gly Gly Leu Tyr Leu Cys Arg Phe Gly Pro Val Thr Trp Asp Cys Gly
1 5 10 15
Tyr Lys Gly Gly
20
<210>31
<211>20
<212>PRT
＜213〉people
<400>31
Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp Val Cys Lys
1 5 10 15
Pro Gln Gly Gly
20
<210>32
<211>20
<212>PRT
＜213〉people
<400>32
Gly Gly Asp Tyr His Cys Arg Met Gly Pro Leu Thr Trp Val Cys Lys
1 5 10 15
Pro Leu Gly Gly
20
<210>33
<211>20
<212>PRT
＜213〉people
<400>33
Val Gly Asn Tyr Met Cys His Phe Gly Pro Ile Thr Trp Val Cys Arg
1 5 10 15
Pro Gly Gly Gly
20
<210>34
<211>20
<212>PRT
＜213〉people
<400>34
Gly Gly Val Tyr Ala Cys Arg Met Gly Pro Ile Thr Trp Val Cys Ser
1 5 10 15
Pro Leu Gly Gly
20
<210>35
<211>19
<212>PRT
＜213〉people
<400>35
Val Gly Asn Tyr Met Ala His Met Gly Pro Ile Thr Trp Val Cys Arg
1 5 10 15
Pro Gly Gly
<210>36
<211>18
<212>PRT
＜213〉people
<400>36
Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp Val Cys Lys
1 5 10 15
Pro Gln
<210>37
<211>21
<212>PRT
＜213〉people
<400>37
Gly Gly Leu Tyr Ala Cys His Met Gly Pro Met Thr Ala Ile Val Cys
1 5 10 15
Gln Pro Leu Arg Gly
20
<210>38
<211>22
<212>PRT
＜213〉people
<400>38
Thr Ile Ala Gln Tyr Ile Cys Tyr Met Gly Pro Glu Thr Trp Glu Cys
1 5 10 15
Arg Pro Ser Pro Lys Ala
20
<210>39
<211>13
<212>PRT
＜213〉people
<400>39
Tyr Ser Cys His Phe Gly Pro Leu Thr Trp Val Cys Lys
1 5 10
<210>40
<211>11
<212>PRT
＜213〉people
<400>40
Tyr Cys His Phe Gly Pro Leu Thr Trp Val Cys
1 5 10
<210>41
<211>12
<212>PRT
＜213〉people
<400>41
Ser Cys His Phe Gly Pro Leu Thr Trp Val Cys Lys
1 5 10
<210>42
<211>13
<212>PRT
＜213〉artificial
<220>
＜223〉peptide
<220>
＜221〉comprehensive characteristics
<222>(2)..(2)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(4)..(6)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(9)..(9)
＜223〉Xaa can be any naturally occurring aminoacid
<220>
＜221〉comprehensive characteristics
<222>(12)..(13)
＜223〉Xaa can be any naturally occurring aminoacid
<400>42
Ala Xaa Asn Xaa Xaa Xaa Gly Pro Xaa Thr Trp Xaa Xaa
1 5 10
<210>43
<211>100
<212>DNA
＜213〉people
<400>43
tacaggccca gatccagggc ggtacctaca gctgccactt cgggcccctc acgtgggtgt 60
gcaagcccca gggcggcgga agcgggggag gctccggtac 100
<210>44
<211>96
<212>DNA
＜213〉people
<400>44
cggagcctcc cccgcttccg ccgccctggg gcttgcacac ccacgtgagg ggcccgaagt 60
ggcagctgta ggtaccgccc tggatctggg cctgta 96
<210>45
<211>85
<212>DNA
＜213〉people
<400>45
tacaggccca gggcggtacc tacagctgcc acttcgggcc cctcacgtgg gtgtgcaagc 60
cccagggcgg cggatcaggt aagtt 85
<210>46
<211>89
<212>DNA
＜213〉people
<400>46
ctagaactta cctgatccgc egccctgggg cttgcacacc cacgtgaggg gcccgaagtg 60
gcagctgtag gtaccgccct gggcctgta 89
<210>47
<211>35
<212>PRT
＜213〉people
<400>47
Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala
1 5 10 15
Pro Glu Leu Leu Gly Gly Pro Ser Glu Glu Gln Tyr Asn Ser Thr Tyr
20 25 30
Arg Val Val
35
<210>48
<211>51
<212>PRT
＜213〉people
<400>48
Gln Ile Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Thr Val Ser Ser Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys
35 40 45
Pro Pro Gys
50
<210>49
<211>51
<212>PRT
＜213〉people
<400>49
Gln Ile Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Thr Val Ser Ser Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys
35 40 45
Pro Pro Cys
50
<210>50
<211>269
<212>PRT
＜213〉people
<400>50
Gln Ile Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Thr Val Ser Ser Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys
35 40 45
Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu
50 55 60
Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu
65 70 75 80
Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys
85 90 95
Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys
100 105 110
Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu
115 120 125
Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys
130 135 140
Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys
145 150 155 160
Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser
165 170 175
Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys
180 185 190
Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln
195 200 205
Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly
210 215 220
Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln
225 230 235 240
Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn
245 250 255
His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys
260 265
<210>51
<211>266
<212>PRT
＜213〉people
<400>51
Gln Ile Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Thr Val Ser Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Ser Cys
35 40 45
Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro
50 55 60
Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys
65 70 75 80
Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp
85 90 95
Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu
100 105 110
Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu
115 120 125
His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn
130 135 140
Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly
145 150 155 160
Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Gln Glu Glu
165 170 175
Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr
180 185 190
Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn
195 200 205
Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe
210 215 220
Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn
225 230 235 240
Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr
245 250 255
Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys
260 265
<210>52
<211>37
<212>PRT
＜213〉people
<400>52
Gln Ile Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Thr Val Ser Ser
35
<210>53
<211>25
<212>PRT
＜213〉people
<400>53
Gln Ile Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser
20 25
<210>54
<211>102
<212>PRT
＜213〉people
<400>54
Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser
1 5 10 15
Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp
20 25 30
Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn
35 40 45
Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val
50 55 60
Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu
65 70 75 80
Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys
85 90 95
Thr Ile Ser Lys Ala Lys
100
<210>55
<211>107
<212>PRT
＜213〉people
<400>55
Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Asp
l 5 10 15
Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe
20 25 30
Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu
35 40 45
Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe
50 55 60
Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly
65 70 75 80
Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr
85 90 95
Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys
100 105
<210>56
<211>20
<212>PRT
＜213〉people
<400>56
Glu Ser Lys Tyr Gly Pro Pro Cys Pro Ser Cys Pro Ala Pro Glu Phe
1 5 10 15
Leu Gly Gly Pro
20
<210>57
<211>102
<212>PRT
＜213〉people
<400>57
Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser
1 5 10 15
Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp
20 25 30
Pro Glu Val Gln Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn
35 40 45
Ala Lys Thr Lys Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val
50 55 60
Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu
65 70 75 80
Tyr Lys Cys Lys Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys
85 90 95
Thr Ile Ser Lys Ala Lys
100
<210>58
<211>107
<212>PRT
＜213〉people
<400>58
Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Gln Glu
1 5 10 15
Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe
20 25 30
Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu
35 40 45
Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe
50 55 60
Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly
65 70 75 80
Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr
85 90 95
Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys
100 105
<210>59
<2ll>102
<212>PRT
＜213〉people
<400>59
Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser
1 5 10 15
Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser Gln Glu Asp
20 25 30
Pro Glu Val Gln Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn
35 40 45
Ala Lys Thr Lys Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val
50 55 60
Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu
65 70 75 80
Tyr Lys Cys Lys Val Ser Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys
85 90 95
Thr Ile Ser Lys Ala Lys
100
<210>60
<2ll>107
<212>PRT
＜213〉people
<400>60
Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Gln Glu
1 5 10 15
Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe
20 25 30
Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu
35 40 45
Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe
50 55 60
Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly
65 70 75 80
Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr
85 90 95
Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys
100 105
<210>61
<211>4
<212>PRT
＜213〉people
<400>61
Gly Gly Gly Ser
1
<210>62
<211>54
<212>PRT
＜213〉people
<400>62
Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp Val Cys Lys
1 5 10 15
Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu Val Thr Val
20 25 30
Ser Ser Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys
35 40 45
Pro Ala Pro Glu Leu Leu
50
<210>63
<211>57
<212>PRT
＜213〉people
<400>63
Gln Val Gln Gly Gly Thr Tyr Ser Ser His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Ser Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Thr Val Ser Ser Glu Pro Lys Ser Ala Asp Lys Thr His Thr Cys
35 40 45
Pro Pro Cys Pro Ala Pro Glu Ala Ala
50 55
<210>64
<211>57
<212>PRT
＜213〉people
<400>64
Gln Ile Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Thr Val Ser Ser Glu Pro Lys Ser Ala Asp Lys Thr His Thr Cys
35 40 45
Pro Pro Cys Pro Ala Pro Glu Ala Ala
50 55
<210>65
<211>57
<212>PRT
＜213〉people
<400>65
Gln Val Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Thr Val Ser Ser Glu Pro Lys Ser Ala Asp Lys Thr His Thr Cys
35 40 45
Pro Pro Cys Pro Ala Pro Glu Ala Ala
50 55
<210>66
<211>57
<212>PRT
＜213〉people
<400>66
Gln Val Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Thr Val Ser Ser Glu Pro Lys Ser Ala Asp Lys Thr His Thr Cys
35 40 45
Pro Pro Cys Pro Ala Pro Glu Leu Ala
50 55
<210>67
<211>57
<212>PRT
＜213〉people
<400>67
Gln Val Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Thr Val Ser Ser Glu Pro Lys Ser Ala Asp Lys Thr His Thr Cys
35 40 45
Pro Pro Cys Pro Ala Pro Glu Ala Leu
50 55
<210>68
<211>57
<212>PRT
＜213〉people
<400>68
Gln Val Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Thr Val Ser Ser Glu Pro Lys Ser Ala Asp Lys Thr His Thr Cys
35 40 45
Pro Pro Cys Pro Ala Pro Glu Leu Glu
50 55
<210>69
<211>57
<212>PRT
＜213〉people
<400>69
Gln Ile Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Thr Val Ser Ser Glu Pro Lys Ser Ser Asp Lys Thr His Thr Cys
35 40 45
Pro Pro Cys Pro Ala Pro Glu Phe Leu
50 55
<210>70
<211>50
<212>PRT
＜213〉people
<400>70
Gln Ile Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Thr Leu Val Thr Val Ser
20 25 30
Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Ser Cys Pro Ala Pro Glu
35 40 45
Phe Leu
50
<210>71
<211>54
<212>PRT
＜2l3〉people
<400>71
Gln Ile Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Thr Val Ser Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Ser Cys
35 40 45
Pro Ala Pro Glu Phe Leu
50
<210>72
<211>56
<212>PRT
＜213〉people
<400>72
Gln Ile Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly
20 25 30
Thr Leu Val Thr Val Ser Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro
35 40 45
Ser Cys Pro Ala Pro Glu Phe Leu
50 55
<210>73
<211>58
<212>PRT
＜213〉people
<400>73
Gln Ile Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly
20 25 30
Ser Gly Thr Leu Val Thr Val Ser Ser Glu Ser Lys Tyr Gly Pro Pro
35 40 45
Cys Pro Ser Cys Pro Ala Pro Glu Phe Leu
50 55
<210>74
<211>47
<212>PRT
＜213〉people
<400>74
Gln Val Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Thr Val Ser Ser Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu
35 40 45
<210>75
<211>50
<212>PRT
＜213〉people
<400>75
Gln Val Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Thr Val Ser Ser Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu
35 40 45
Leu Leu
50
<210>76
<211>53
<212>PRT
＜213〉people
<400>76
Gln Val Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Thr Val Ser Ser Ala Asp Lys Thr His Thr Cys Pro Pro Cys Pro
35 40 45
Ala Pro Glu Leu Leu
50
<210>77
<211>57
<212>PRT
＜213〉people
<400>77
Gln Ile Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Thr Val Ser Ser Glu Pro Lys Ser Ala Asp Lys Thr His Ala Cys
35 40 45
Pro Pro Cys Pro Ala Pro Glu Leu Leu
50 55
<210>78
<211>57
<212>PRT
＜213〉people
<400>78
Gln Ile Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Ala Val Ser Ser Glu Pro Lys Ser Ala Asp Lys Thr His Thr Cys
35 40 45
Pro Pro Cys Pro Ala Pro Glu Leu Leu
50 55
<210>79
<211>57
<212>PRT
＜213〉people
<400>79
Gln Ile Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Thr Val Ser Ser Glu Pro Lys Ser Ala Asp Lys Ala His Thr Cys
35 40 45
Pro Pro Cys Pro Ala Pro Glu Leu Leu
50 55
<210>80
<211>57
<212>PRT
＜213〉people
<400>80
Gln Ile Gln Gly Gly Thr Tyr Ser Cys His Phe Gly Pro Leu Thr Trp
1 5 10 15
Val Cys Lys Pro Gln Gly Gly Gly Ser Gly Gly Gly Ser Gly Thr Leu
20 25 30
Val Ala Val Ser Ser Glu Pro Lys Ser Ala Asp Lys Thr His Ala Cys
35 40 45
Pro Pro Cys Pro Ala Pro Glu Leu Leu
50 55
1
40

Claims (35)

1. at least a EPO mimetic CH 1 deleted mimetibodies nucleic acid comprises at least a polynucleotide of at least a aminoacid sequence among the coding SEQ ID NO:50-51 or polynucleotide complementary with it.
2. at least a EPO mimetic CH 1 deleted mimetibodies nucleic acid comprises at least a polynucleotide of at least a aminoacid sequence among the coding SEQ ID NO:62-80 or polynucleotide complementary with it.
3. at least a EPO mimetic CH 1 deleted mimetibodies nucleic acid comprises at least a polynucleotide of the polypeptide of coding type (I):
(V1 (n)-Pep (n)-Flex (n)-V2 (n)-pHinge (n)-CH2 (n)-CH3 (n)) (m), wherein V1 is at least one part of immune globulin variable region N-terminal, Pep is the EPO simulating peptide of at least one biologically active, Flex is by making analogue body have optional orientation and binding characteristic, thereby provide the polypeptide of structural flexibility, V2 is at least one part of immune globulin variable region C-terminal, pHinge is at least one part of immunoglobulin variable hinge region, CH2 is at least one part of immunoglobulin CH2 constant region, CH3 is at least one part of immunoglobulin CH3 constant region, and n and m can be the integers among the 1-10.
4. at least a EPO mimetic CH 1 deleted mimetibodies polypeptide comprises all continuous amino acids among the SEQ ID NO:50-51.
5. at least a EPO mimetic CH 1 deleted mimetibodies polypeptide comprises all continuous amino acids among the SEQ ID NO:62-80.
6. at least a EPO mimetic CH 1 deleted mimetibodies polypeptide comprises the polypeptide of formula (I):
(V1 (n)-Pep (n)-Flex (n)-V2 (n)-pHinge (n)-CH2 (n)-CH3 (n)) (m), wherein V1 is QIQ, Pep is the peptide that is selected from least one biologically active among the SEQ ID NOS:1-42, Flex comprises GGGS (SEQ ID NO:61), V2 is GTLVTVSS (SEQ ID NO:52), pHinge is EPKSCDKTHTCPPCPAPELLGGP (SEQ ID NO:53), CH2 is SVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREE QYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAK (SEQ ID NO:54), CH3 is GQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPP VLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO:55), and n and m are the integer among the 1-10.
7. at least a EPO mimetic CH 1 deleted mimetibodies polypeptide comprises the polypeptide of formula (I):
(V1 (n)-Pep (n)-Flex (n)-V2 (n)-pHinge (n)-CH2 (n)-CH3 (n)) (m), wherein V1 is QIQ, Pep is the peptide that is selected from least one biologically active among the SEQ ID NOS:1-42, Flex comprises GGGS, V2 is GTLVTVSS (SEQ ID NO:52), pHinge is ESKYGPPCPSCPAPEFLGGP (SEQ ID NO:56), CH2 is SVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREE QFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAK (SEQ ID NO:57), CH3 is GQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPP VLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGK (SEQ IDNO:58), and n and m are the integer among the 1-10.
8. at least a EPO mimetic CH 1 deleted mimetibodies polypeptide comprises the polypeptide of formula (I):
(V1 (n)-Pep (n)-Flex (n)-V2 (n)-pHinge (n)-CH2 (n)-CH3 (n)) (m), wherein V1 is the N-terminal part of human variable region, Pep is the peptide that is selected from least one biologically active among the SEQ ID NOS:1-42, Flex is by making analogue body have optional orientation and binding characteristic, thereby provide the polypeptide of structural flexibility, V2 is at least one part of immune globulin variable region C-terminal, pHinge is at least one part of immunoglobulin variable hinge region, CH2 is SVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREE QYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAK (SEQ ID NO:54), CH3 is GQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPP VLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO:55), and n and m are the integer among the 1-10.
9. at least a EPO mimetic CH 1 deleted mimetibodies polypeptide comprises the polypeptide of formula (I):
(V1 (n)-Pep (n)-Flex (n)-V2 (n)-pHinge (n)-CH2 (n)-CH3 (n)) (m), wherein V1 is the N-terminal part of human variable region, Pep is the peptide that is selected from least one biologically active among the SEQ ID NOS:1-42, Flex is by making analogue body have optional orientation and binding characteristic, thereby provide the polypeptide of structural flexibility, V2 is at least one part of immune globulin variable region C-terminal, pHinge is at least one part of immunoglobulin variable hinge region, CH2 is SVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREE QFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAK (SEQ ID NO:59), CH3 is GQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPP VLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGK (SEQ IDNO:60), and n and m are the integer among the 1-10.
10. at least a EPO mimetic CH 1 deleted mimetibodies polypeptide comprises the polypeptide of formula (I):
(V1 (n)-Pep (n)-Flex (n)-V2 (n)-pHinge (n)-CH2 (n)-CH3 (n)) (m), wherein V1 is QIQ, P8p is the peptide that is selected from least one biologically active among the SEQ ID NOS:1-42, Flex is by making analogue body have optional orientation and binding characteristic, thereby provide the polypeptide of structural flexibility, V2 is at least one part of immune globulin variable region C-terminal, pHinge is at least one part of immunoglobulin variable hinge region, CH2 is at least one part of immunoglobulin CH2 constant region, CH3 is at least one part of immunoglobulin CH3 constant region, and n and m are the integer among the 1-10.
11. at least a EPO mimetic CH 1 deleted mimetibodies polypeptide comprises the polypeptide of formula (I):
(V1 (n)-Pep (n)-Flex (n)-V2 (n)-pHinge (n)-CH2 (n)-CH3 (n)) (m), wherein V1 is at least one part of immune globulin variable region N-terminal, Pep is the EPO simulating peptide of at least one biologically active, Flex comprises GGGS (SEQ ID NO:61), V2 is at least one part of immune globulin variable region C-terminal, pHinge is at least one part of immunoglobulin variable hinge region, CH2 is at least one part of immunoglobulin CH2 constant region, CH3 is at least one part of immunoglobulin CH3 constant region, and n and m are the integer among the 1-10.
12. at least a EPO mimetic CH 1 deleted mimetibodies polypeptide comprises the polypeptide of formula (I):
(V1 (n)-Pep (n)-Flex (n)-V2 (n)-pHinge (n)-CH2 (n)-CH3 (n)) (m), wherein V1 is at least one part of immune globulin variable region N-terminal, Pep is the EPO simulating peptide of at least one biologically active, Flex is by making analogue body have optional orientation and binding characteristic, thereby provide the polypeptide of structural flexibility, V2 is GTLVTVSS (SEQ ID NO:52), pHinge is at least one part of immunoglobulin variable hinge region, CH2 is at least one part of immunoglobulin CH2 constant region, CH3 is at least one part of immunoglobulin CH3 constant region, and n and m can be the integer among the 1-10.
13. at least a EPO mimetic CH 1 deleted mimetibodies polypeptide comprises the polypeptide of formula (I):
(V1 (n)-Pep (n)-Flex (n)-V2 (n)-pHinge (n)-CH2 (n)-CH3 (n)) (m), wherein V 1 is at least one part of immune globulin variable region N-terminal, Pep is the EPO simulating peptide of at least one biologically active, Flex is by making analogue body have optional orientation and binding characteristic, thereby provide the polypeptide of structural flexibility, V2 is at least one part of immune globulin variable region C-terminal, pHinge is EPKSCDKTHTCPPCPAPELLGGP (SEQ ID NO:53), CH2 is at least one part of immunoglobulin CH2 constant region, CH3 is at least one part of immunoglobulin CH3 constant region, and n and m can be the integer among the 1-10.
14. at least a EPO mimetic CH 1 deleted mimetibodies polypeptide comprises the polypeptide of formula (I):
(V1 (n)-Pep (n)-Flex (n)-V2 (n)-pHinge (n)-CH2 (n)-CH3 (n)) (m), wherein V1 is at least one part of immune globulin variable region N-terminal, Pep is the peptide that is selected from least one biologically active of SEQ IDNOS:1-20, Flex is by making analogue body have optional orientation and binding characteristic, thereby provide the polypeptide of structural flexibility, V2 is at least one part of immune globulin variable region C-terminal, pHinge is at least one part of immunoglobulin variable hinge region, CH2 is at least one part of immunoglobulin CH2 constant region, CH3 is at least one part of immunoglobulin CH3 constant region, and n and m can be the integer among the 1-10.
15. at least a EPO mimetic CH 1 deleted mimetibodies polypeptide comprises the polypeptide of formula (I):
(V1 (n)-Pep (n)-Flex (n)-V2 (n)-pHinge (n)-CH2 (n)-CH3 (n)) (m), wherein V1 is at least one part of immune globulin variable region N-terminal, Pep is the peptide that is selected from least one biologically active among the SEQ IDNOS:21-42, Flex is by making analogue body have optional orientation and binding characteristic, thereby provide the polypeptide of structural flexibility, V2 is at least one part of immune globulin variable region C-terminal, pHinge is at least one part of immunoglobulin variable hinge region, CH2 is at least one part of immunoglobulin CH2 constant region, CH3 is at least one part of immunoglobulin CH3 constant region, and n and m can be the integer among the 1-10.
16. EPO mimetic CH 1 deleted mimetibodies nucleic acid or the EPO mimetic CH 1 deleted mimetibodies polypeptide of claim 1-15 in each, wherein this polypeptide has at least a activity of at least a Pep polypeptide.
17. anti-idiotype monoclonal or polyclonal antibody, fusion rotein, or its fragment can combine with at least a EPO mimetic CH 1 deleted mimetibodies polypeptid specificity of claim 1-15 in each.
18. coding claim 1-17 in each at least a EPO mimetic CH 1 deleted mimetibodies polypeptide or the EPO mimetic CH 1 deleted mimetibodies nucleic acid of EPO mimetic CH 1 deleted mimetibodies antibody.
19. comprise the EPO mimetic CH 1 deleted mimetibodies carrier of at least a isolating nucleic acid of claim 18.
20. comprise the EPO mimetic CH 1 deleted mimetibodies host cell of the isolating nucleic acid of claim 18.
21. the EPO mimetic CH 1 deleted mimetibodies host cell of claim 20, wherein this host cell is for being selected from COS-1, COS-7, HEK293, BHK21, CHO, BSC-1, Hep G2,653, SP2/0,293, NSO, DG44CHO, CHO K1, HeLa, myeloma or lymphoma cell, or at least a cell in the deriving arbitrarily of above-mentioned cell, immortalization or the transformant.
22. method that generates at least a EPO mimetic CH 1 deleted mimetibodies polypeptide or EPO mimetic CH 1 deleted mimetibodies antibody, be included in external, the body or under the original position condition, the nucleic acid of translation claim 18 makes EPO mimetic CH 1 deleted mimetibodies or antibody can detect or callable quantity is expressed.
23. a compositions comprises at least a EPO mimetic CH 1 deleted mimetibodies nucleic acid, the many skins of EPO mimetic CH 1 deleted mimetibodies, or the EPO mimetic CH 1 deleted mimetibodies antibody of claim 1-17 in each.
24. the compositions of claim 23, wherein said composition further comprises at least a materia medica acceptable carrier or diluent.
25. the compositions of claim 23, further comprise at least a compositions, said composition contains at least a chemical compound for the treatment of effective dose, compositions or polypeptide, this chemical compound, compositions or polypeptide are selected from detectable label or reporter molecule, the TNF antagonist, anti-infectives, cardiovascular (CV) system medicine, central nervous system (CNS) medicine, autonomic nervous system (ANS) medicine, respiratory drugs, gastrointestinal (GI) tract drug, hormonal medicaments, the medicine that is used for balance body fluid or electrolyte, blood substance, antitumor drug, immunoregulation medicament, eye, ear or nose medication, local application, nutritional drugs, at least a in cytokine or the cytokine antagonist.
26. the compositions of claim 23, its form are to be selected from least a in liquid, gas or dry thing, solution, mixture, suspension, emulsion or colloid, lyophilized formulations or the powder.
27. diagnose or treat the conditions associated method of EPO part in cell, tissue, organ or the animal, comprise for one kind
(a) make cell, tissue, organ or animal contact or it is used a kind of compositions, said composition contains at least a EPO mimetic CH 1 deleted mimetibodies nucleic acid, polypeptide or the antibody of claim 1-17 in each of effective dose.
28. being the above-mentioned cell of per kilogram, tissue, organ or animal, the method for claim 27, wherein said effective dose use the EPO mimetic CH 1 deleted mimetibodies antibody of 0.001-50mg; 0.000001-500mg above-mentioned EPO mimetic CH 1 deleted mimetibodies; Or the above-mentioned EPO mimetic CH 1 deleted mimetibodies nucleic acid of 0.0001-100 μ g.
29. the method for claim 27, wherein above-mentioned contact or to use be by being selected from parenteral, subcutaneous, intramuscular, intravenous, intraarticular, in the bronchus, in the abdomen, in the capsule, in the cartilage, intracavity, in the body cavity, in the cerebellum, Intraventricular, colonic, in the cervix uteri, gastric, in the liver, in the cardiac muscle, in the bone, in the pelvis, in the pericardium, intraperitoneal, in the pleura, in the prostate, in the lung, internal rectum, in the kidney, in the retina, in the spinal column, in the synovial membrane, intrathoracic, intrauterine, intravesical; in the pathological changes; medicine group; vagina; rectum; cheek; Sublingual; at least a pattern in intranasal or the percutaneous approach realizes.
30. the method for claim 27, further be included in before above-mentioned (a) contact or use, simultaneously or afterwards, use at least a compositions, said composition contains at least a chemical compound or the polypeptide for the treatment of effective dose, and this chemical compound or polypeptide are selected from detectable label or receptor, the TNF antagonist, anti-infectives, cardiovascular (CV) system medicine, central nervous system (CNS) medicine, autonomic nervous system (ANS) medicine, respiratory drugs, gastrointestinal (GI) tract drug, hormonal medicaments, the medicine that is used for balance body fluid or electrolyte, blood substance, antitumor drug, immunoregulation medicament, eye, ear or nose medication, local application, nutritional drugs, at least a in cytokine or the cytokine antagonist.
31. device, contain at least a isolating EPO mimetic CH 1 deleted mimetibodies polypeptide of claim 1-17 in each, antibody or nucleic acid, wherein this device is applicable to by being selected from parenteral, subcutaneous, intramuscular, intravenous, intraarticular, in the bronchus, in the abdomen, in the capsule, in the cartilage, intracavity, in the body cavity, in the cerebellum, Intraventricular, colonic, in the cervix uteri, gastric, in the liver, in the cardiac muscle, in the bone, in the pelvis, in the pericardium, intraperitoneal, in the pleura, in the prostate, in the lung, internal rectum, in the kidney, in the retina, in the spinal column, in the synovial membrane, intrathoracic, intrauterine, intravesical; in the pathological changes; medicine group; vagina; rectum; cheek; Sublingual; at least a pattern in intranasal or the percutaneous approach contacts or uses above-mentioned at least a EPO mimetic CH 1 deleted mimetibodies polypeptide; antibody or nucleic acid.
32. goods that are used for human medicine or diagnostic uses comprise packaging material and container, this container contains at least a isolating EPO mimetic CH 1 deleted mimetibodies polypeptide, antibody or the nucleic acid of claim 1-17 in each.
33. the goods of claim 34, wherein said vesse is a parenteral, subcutaneous, intramuscular, intravenous, intraarticular, in the bronchus, in the abdomen, in the capsule, in the cartilage, intracavity, in the body cavity, in the cerebellum, Intraventricular, colonic, in the cervix uteri, gastric, in the liver, in the cardiac muscle, in the bone, in the pelvis, in the pericardium, intraperitoneal, in the pleura, in the prostate, in the lung, internal rectum, in the kidney, in the retina, in the spinal column, in the synovial membrane, intrathoracic, intrauterine, intravesical; in the pathological changes; medicine group; vagina; rectum; cheek; Sublingual; an ingredient of intranasal or percutaneous delivery device or system.
34. one kind is used for generating each the method for at least a isolating EPO mimetic CH 1 deleted mimetibodies polypeptide, antibody or nucleic acid of claim 1-17, this method comprise provide at least a can be can detect or callable scale reaches host cell, transgenic animal, transgenic plant, the plant cell of aforementioned polypeptides, antibody or nucleic acid.
35. at least a EPO mimetic CH 1 deleted mimetibodies polypeptide, antibody or nucleic acid that method generated by claim 34.
CNA038201100A 2002-06-28 2003-06-30 Mammalian EPO mimetic CH1 deleted mimetibodies, compositions, methods and uses Pending CN1735433A (en)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
US39243102P 2002-06-28 2002-06-28
US60/392,431 2002-06-28
US60/412,144 2002-09-19

Publications (1)

Publication Number Publication Date
CN1735433A true CN1735433A (en) 2006-02-15

Family

ID=30000868

Family Applications (1)

Application Number Title Priority Date Filing Date
CNA038201100A Pending CN1735433A (en) 2002-06-28 2003-06-30 Mammalian EPO mimetic CH1 deleted mimetibodies, compositions, methods and uses

Country Status (6)

Country Link
EP (1) EP1545608A4 (en)
JP (1) JP2006504406A (en)
CN (1) CN1735433A (en)
AU (1) AU2003280130B2 (en)
CA (1) CA2490409A1 (en)
WO (1) WO2004002417A2 (en)

Families Citing this family (113)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
MY126795A (en) 1998-10-23 2006-10-31 Amgen K A Inc Dimeric thrombopoietic peptide mimetics binding to mp1 receptor and having thrombopoietic activity.
US6660843B1 (en) 1998-10-23 2003-12-09 Amgen Inc. Modified peptides as therapeutic agents
EP1572079A4 (en) * 2002-03-29 2006-09-06 Centocor Inc Mammalian cdr mimetibodies, compositions, methods and uses
US7241733B2 (en) * 2002-06-28 2007-07-10 Centocor, Inc. Mammalian EPO mimetic CH1 deleted mimetibodies, compositions, methods and uses
WO2005081687A2 (en) * 2003-09-30 2005-09-09 Centocor, Inc. Human hinge core mimetibodies, compositions, methods and uses
UA89481C2 (en) * 2003-09-30 2010-02-10 Центокор, Инк. Human epo mimetic hinge core mimetibodies, compositions, methods and uses
CA2563379A1 (en) * 2004-03-31 2005-10-20 Centocor, Inc. Human glp-1 mimetibodies, compositions, methods and uses
MX2007003320A (en) 2004-09-24 2007-05-18 Amgen Inc Modified fc molecules.
CA2581505A1 (en) * 2004-09-27 2006-04-06 Centocor, Inc. Srage mimetibody, compositions, methods and uses
AU2005299440B2 (en) * 2004-10-25 2011-12-15 Centocor, Inc. Melanocortin receptor binding mimetibodies, compositions, methods and uses
EP1871811A4 (en) * 2005-03-28 2009-07-22 Centocor Inc Human glp-1 mimetibodies, compositions, methods and uses
US8008453B2 (en) 2005-08-12 2011-08-30 Amgen Inc. Modified Fc molecules
WO2007115148A2 (en) * 2006-03-31 2007-10-11 Centocor, Inc. Human mimetic epo hinge core mimetibodies
US7943310B2 (en) 2006-08-30 2011-05-17 Centocor Ortho Biotech Inc. Methods for assessing response to therapy in subjects having ulcerative colitis
WO2008079973A2 (en) 2006-12-21 2008-07-03 Centocor, Inc. Egfr binding peptides and uses thereof
US8278415B2 (en) 2006-12-21 2012-10-02 Centocor, Inc. Dimeric high affinity EGFR constructs and uses thereof
WO2008079982A2 (en) 2006-12-21 2008-07-03 Centocor, Inc. Liposome composition for targeting egfr receptor
WO2008092209A1 (en) * 2007-02-01 2008-08-07 Arana Therapeutics Limited Protein construct with improved properties
US8383114B2 (en) 2007-09-27 2013-02-26 Amgen Inc. Pharmaceutical formulations
WO2009064838A1 (en) 2007-11-15 2009-05-22 Amgen, Inc. Aqueous formulation of erythropoiesis stimulating protein stablised by antioxidants for parenteral administration
MX2010008096A (en) 2008-01-25 2010-09-22 Amgen Inc Ferroportin antibodies and methods of use.
ES2487846T3 (en) 2008-05-01 2014-08-25 Amgen, Inc. Anti-hepcindin antibodies and methods of use
CN102165315B (en) 2008-08-29 2014-06-18 森托科尔奥索生物科技公司 Markers and methods for assessing and treating ulcerative colitis and related disorders using a 20 gene panel
EP2352998A4 (en) 2008-11-07 2011-09-21 Centocor Ortho Biotech Inc Markers and methods for assessing and treating lupus patients susceptible to photoprovocation
EP3693014A1 (en) 2008-11-13 2020-08-12 The General Hospital Corporation Methods and compositions for regulating iron homeostasis by modulation bmp-6
WO2011050333A1 (en) 2009-10-23 2011-04-28 Amgen Inc. Vial adapter and system
JP5850846B2 (en) 2009-11-17 2016-02-03 ヤンセン バイオテツク，インコーポレーテツド Display of disulfide-bonded dimeric proteins on filamentous phage
CN102770555B (en) 2009-11-17 2015-01-28 詹森生物科技公司 Improved bacterial membrane protein secrection
EP2536435B1 (en) 2010-02-18 2017-11-15 Janssen Biotech, Inc. Monkey homolog of human interferon omega
JP5908888B2 (en) 2010-04-16 2016-04-26 ヤンセン バイオテツク，インコーポレーテツド Modified plant cysteine protease and use thereof
FI2575935T4 (en) 2010-06-07 2023-11-23 Amgen Inc Drug delivery device
JP6126991B2 (en) 2010-09-27 2017-05-10 ヤンセン バイオテツク，インコーポレーテツド Antibody binding to human type II collagen
EP2691065B1 (en) 2011-03-31 2017-03-01 Amgen Inc. Vial adapter and system
JP6038884B2 (en) 2011-04-20 2016-12-07 アムゲン・インコーポレーテッド Automatic injection device
ES2923766T3 (en) 2011-10-14 2022-09-30 Amgen Inc Injector and mounting method
EP3656426B1 (en) 2012-11-21 2023-05-17 Amgen Inc. Drug delivery device
BR112015022123B1 (en) 2013-03-15 2022-08-09 Intrinsic Lifesciences, Llc ANTIBODIES, ANTIGEN-BINDING FRAGMENTS OF SPECIFICALLY BINDING HEPCIDIN OR A HEPCIDIN PEPTIDE, USE, CONTAINING MEDIUM AND KIT
TWI580452B (en) 2013-03-15 2017-05-01 安美基公司 Cassette for an injector, injector, and method of using an apparatus comprising an autoinjector and a cassette
CA2904725C (en) 2013-03-15 2022-04-12 Amgen Inc. Drug cassette, autoinjector, and autoinjector system
WO2014144542A2 (en) * 2013-03-15 2014-09-18 Omeros Corporation Methods of generating bioactive peptide-bearing antibodies and compositions comprising the same
TWI592183B (en) 2013-03-15 2017-07-21 安美基公司 Body contour adaptable autoinjector device
CN113559363B (en) 2013-03-22 2023-10-31 美国安进公司 Syringe and method of assembly
WO2015061386A1 (en) 2013-10-24 2015-04-30 Amgen Inc. Injector and method of assembly
AU2014340174B2 (en) 2013-10-24 2019-09-12 Amgen Inc. Drug delivery system with temperature-sensitive control
WO2015119906A1 (en) 2014-02-05 2015-08-13 Amgen Inc. Drug delivery system with electromagnetic field generator
BR112016025852B1 (en) 2014-05-07 2022-11-01 Amgen Inc INJECTION DEVICE FOR DRUG APPLICATION
JP6817074B2 (en) 2014-06-03 2021-01-20 アムジエン・インコーポレーテツド Controllable drug delivery system and usage
CA2961917A1 (en) 2014-09-22 2016-03-31 Intrinsic Lifesciences Llc Humanized anti-hepcidin antibodies and uses thereof
EP3206739B1 (en) 2014-10-14 2021-12-01 Amgen Inc. Drug injection device with visual and audio indicators
ES2898469T3 (en) 2014-12-19 2022-03-07 Amgen Inc Medication administration device with proximity sensor
EP3689394A1 (en) 2014-12-19 2020-08-05 Amgen Inc. Drug delivery device with live button or user interface field
WO2016133947A1 (en) 2015-02-17 2016-08-25 Amgen Inc. Drug delivery device with vacuum assisted securement and/or feedback
EP3981450A1 (en) 2015-02-27 2022-04-13 Amgen, Inc Drug delivery device having a needle guard mechanism with a tunable threshold of resistance to needle guard movement
WO2017039786A1 (en) 2015-09-02 2017-03-09 Amgen Inc. Syringe assembly adapter for a syringe
ES2755717T3 (en) 2015-12-09 2020-04-23 Amgen Inc Autoinjector with signaling cap
US11154661B2 (en) 2016-01-06 2021-10-26 Amgen Inc. Auto-injector with signaling electronics
DK3429663T3 (en) 2016-03-15 2020-09-28 Amgen Inc REDUCING THE LIKELIHOOD OF GLASS BREAKING IN MEDICINE ADMINISTRATION DEVICES
US11541168B2 (en) 2016-04-29 2023-01-03 Amgen Inc. Drug delivery device with messaging label
WO2017192287A1 (en) 2016-05-02 2017-11-09 Amgen Inc. Syringe adapter and guide for filling an on-body injector
US10988284B2 (en) 2016-05-13 2021-04-27 Amgen Inc. Vial sleeve assembly
EP3458988B1 (en) 2016-05-16 2023-10-18 Amgen Inc. Data encryption in medical devices with limited computational capability
EP3465124A1 (en) 2016-06-03 2019-04-10 Amgen Inc. Impact testing apparatuses and methods for drug delivery devices
EP3478342A1 (en) 2016-07-01 2019-05-08 Amgen Inc. Drug delivery device having minimized risk of component fracture upon impact events
US20190328965A1 (en) 2016-08-17 2019-10-31 Amgen Inc. Drug delivery device with placement detection
EP3532127A1 (en) 2016-10-25 2019-09-04 Amgen Inc. On-body injector
US20190358411A1 (en) 2017-01-17 2019-11-28 Amgen Inc. Injection devices and related methods of use and assembly
CA3052204A1 (en) 2017-02-17 2018-08-23 Amgen Inc. Insertion mechanism for drug delivery device
JP7064501B2 (en) 2017-02-17 2022-05-10 アムジエン・インコーポレーテツド Drug delivery device with sterile fluid flow path and related assembly methods
EP3592403A1 (en) 2017-03-06 2020-01-15 Amgen Inc. Drug delivery device with activation prevention feature
US11571511B2 (en) 2017-03-07 2023-02-07 Amgen Inc. Insertion mechanism and method of inserting a needle of a drug delivery device
KR102619150B1 (en) 2017-03-09 2023-12-27 암겐 인코포레이티드 Insertion mechanism for drug delivery device
CN110446499A (en) 2017-03-20 2019-11-12 豪夫迈·罗氏有限公司 A kind of method of external glycosyl engineering erythropoiesis stimulating protein
CA3052676A1 (en) 2017-03-28 2018-10-04 Amgen Inc. Plunger rod and syringe assembly system and method
EP3634539A1 (en) 2017-06-08 2020-04-15 Amgen Inc. Syringe assembly for a drug delivery device and method of assembly
MX2019014615A (en) 2017-06-08 2020-02-07 Amgen Inc Torque driven drug delivery device.
AU2018288604B2 (en) 2017-06-22 2023-12-21 Amgen Inc. Device activation impact/shock reduction
WO2018237225A1 (en) 2017-06-23 2018-12-27 Amgen Inc. Electronic drug delivery device comprising a cap activated by a switch assembly
IL270784B2 (en) 2017-07-14 2023-11-01 Amgen Inc Needle insertion-retraction system having dual torsion spring system
WO2019018169A1 (en) 2017-07-21 2019-01-24 Amgen Inc. Gas permeable sealing member for drug container and methods of assembly
EP3658203B1 (en) 2017-07-25 2022-08-31 Amgen Inc. Drug delivery device with gear module and related method of assembly
JP7242562B2 (en) 2017-07-25 2023-03-20 アムジエン・インコーポレーテツド Drug delivery device with container access system and associated method of assembly
US20200164155A1 (en) 2017-08-09 2020-05-28 Amgen Inc. Hydraulic-pneumatic pressurized chamber drug delivery system
WO2019036181A1 (en) 2017-08-18 2019-02-21 Amgen Inc. Wearable injector with sterile adhesive patch
US11103636B2 (en) 2017-08-22 2021-08-31 Amgen Inc. Needle insertion mechanism for drug delivery device
EP3691717B1 (en) 2017-10-04 2023-02-08 Amgen Inc. Flow adapter for drug delivery device
IL272636B2 (en) 2017-10-06 2024-10-01 Amgen Inc Drug delivery device with interlock assembly and related method of assembly
MA50348A (en) 2017-10-09 2020-08-19 Amgen Inc DRUG ADMINISTRATION DEVICE INCLUDING A TRAINING ASSEMBLY AND ASSOCIATED ASSEMBLY PROCEDURE
MA50527A (en) 2017-11-03 2020-09-09 Amgen Inc SYSTEM AND APPROACHES TO STERILIZE A DRUG DELIVERY DEVICE
EP3707075A1 (en) 2017-11-06 2020-09-16 Amgen Inc. Fill-finish assemblies and related methods
EP3706830B1 (en) 2017-11-06 2024-08-07 Amgen Inc. Drug delivery device with placement and flow sensing
US11191904B2 (en) 2017-11-10 2021-12-07 Amgen Inc. Plungers for drug delivery devices
MX2020005066A (en) 2017-11-16 2020-08-20 Amgen Inc Autoinjector with stall and end point detection.
JP7370969B2 (en) 2017-11-16 2023-10-30 アムジエン・インコーポレーテツド Door latch mechanism for drug delivery devices
US10835685B2 (en) 2018-05-30 2020-11-17 Amgen Inc. Thermal spring release mechanism for a drug delivery device
US11083840B2 (en) 2018-06-01 2021-08-10 Amgen Inc. Modular fluid path assemblies for drug delivery devices
US12115360B2 (en) 2018-07-24 2024-10-15 Amgen Inc. Hybrid drug delivery devices with grip portion
WO2020023451A1 (en) 2018-07-24 2020-01-30 Amgen Inc. Delivery devices for administering drugs
WO2020023220A1 (en) 2018-07-24 2020-01-30 Amgen Inc. Hybrid drug delivery devices with tacky skin attachment portion and related method of preparation
EP3826701A1 (en) 2018-07-24 2021-06-02 Amgen Inc. Delivery devices for administering drugs
CA3103105A1 (en) 2018-07-31 2020-02-06 Amgen Inc. Fluid path assembly for a drug delivery device
WO2020068623A1 (en) 2018-09-24 2020-04-02 Amgen Inc. Interventional dosing systems and methods
WO2020068476A1 (en) 2018-09-28 2020-04-02 Amgen Inc. Muscle wire escapement activation assembly for a drug delivery device
CN112805048B (en) 2018-10-02 2023-09-22 安进公司 Injection system for drug delivery with internal force transfer
MA53818A (en) 2018-10-05 2022-01-12 Amgen Inc DRUG DELIVERY DEVICE HAVING A DOSE INDICATOR
IL281894B1 (en) 2018-10-15 2024-08-01 Amgen Inc Drug delivery device having damping mechanism
SG11202101824VA (en) 2018-10-15 2021-03-30 Amgen Inc Platform assembly process for drug delivery device
AU2019370159A1 (en) 2018-11-01 2021-04-22 Amgen Inc. Drug delivery devices with partial drug delivery member retraction
US11213620B2 (en) 2018-11-01 2022-01-04 Amgen Inc. Drug delivery devices with partial drug delivery member retraction
TWI831847B (en) 2018-11-01 2024-02-11 美商安進公司 Drug delivery devices with partial needle retraction and methods for operating the same
JP7510952B2 (en) 2019-04-24 2024-07-04 アムジエン・インコーポレーテツド Syringe sterility verification assembly and method
CA3148261A1 (en) 2019-08-23 2021-03-04 Amgen Inc. Drug delivery device with configurable needle shield engagement components and related methods
KR20240011135A (en) 2021-05-21 2024-01-25 암젠 인크 How to Optimize Filling Recipes for Drug Containers
WO2024094457A1 (en) 2022-11-02 2024-05-10 F. Hoffmann-La Roche Ag Method for producing glycoprotein compositions

Family Cites Families (91)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3773919A (en) 1969-10-23 1973-11-20 Du Pont Polylactide-drug mixtures
US4309989A (en) 1976-02-09 1982-01-12 The Curators Of The University Of Missouri Topical application of medication by ultrasound with coupling agent
FR2374910A1 (en) 1976-10-23 1978-07-21 Choay Sa PREPARATION BASED ON HEPARIN, INCLUDING LIPOSOMES, PROCESS FOR OBTAINING IT AND MEDICINAL PRODUCTS CONTAINING SUCH PREPARATIONS
US5179017A (en) 1980-02-25 1993-01-12 The Trustees Of Columbia University In The City Of New York Processes for inserting DNA into eucaryotic cells and for producing proteinaceous materials
US4399216A (en) 1980-02-25 1983-08-16 The Trustees Of Columbia University Processes for inserting DNA into eucaryotic cells and for producing proteinaceous materials
US4634665A (en) 1980-02-25 1987-01-06 The Trustees Of Columbia University In The City Of New York Processes for inserting DNA into eucaryotic cells and for producing proteinaceous materials
US4656134A (en) 1982-01-11 1987-04-07 Board Of Trustees Of Leland Stanford Jr. University Gene amplification in eukaryotic cells
US5149636A (en) 1982-03-15 1992-09-22 Trustees Of Columbia University In The City Of New York Method for introducing cloned, amplifiable genes into eucaryotic cells and for producing proteinaceous products
US5169762A (en) 1983-03-03 1992-12-08 Genentech, Inc. Human nerve growth factor by recombinant technology
US5168062A (en) 1985-01-30 1992-12-01 University Of Iowa Research Foundation Transfer vectors and microorganisms containing human cytomegalovirus immediate-early promoter-regulatory DNA sequence
SE448277B (en) 1985-04-12 1987-02-09 Draco Ab INDICATOR DEVICE WITH A DOSAGE DEVICE FOR MEDICINAL PRODUCTS
US4740587A (en) 1985-07-18 1988-04-26 The Salk Institute For Biological Studies Inhibin and method of purifying same
US4721672A (en) 1985-08-28 1988-01-26 President And Fellows Of Harvard College CDNA and gene for human angiogenin (angiogenesis factor) and methods of expression
US4870163A (en) 1985-08-29 1989-09-26 New York Blood Center, Inc. Preparation of pure human tumor necrosis factor and hybridomas producing monoclonal antibodies to human tumor necrosis factor
GB8601597D0 (en) 1986-01-23 1986-02-26 Wilson R H Nucleotide sequences
US4737578A (en) 1986-02-10 1988-04-12 The Salk Institute For Biological Studies Human inhibin
SE453566B (en) 1986-03-07 1988-02-15 Draco Ab POWDER INHALATOR DEVICE
US4767402A (en) 1986-07-08 1988-08-30 Massachusetts Institute Of Technology Ultrasound enhancement of transdermal drug delivery
CH671155A5 (en) 1986-08-18 1989-08-15 Clinical Technologies Ass
US4946778A (en) 1987-09-21 1990-08-07 Genex Corporation Single polypeptide chain binding molecules
DE3631229A1 (en) 1986-09-13 1988-03-24 Basf Ag MONOCLONAL ANTIBODIES AGAINST HUMAN TUMORNESCROSE FACTOR (TNF) AND THEIR USE
EP0832981A1 (en) 1987-02-17 1998-04-01 Pharming B.V. DNA sequences to target proteins to the mammary gland for efficient secretion
US5075236A (en) 1987-04-24 1991-12-24 Teijin Limited Method of detecting kawasaki disease using anti-tumor necrosis antibody
US4873316A (en) 1987-06-23 1989-10-10 Biogen, Inc. Isolation of exogenous recombinant proteins from the milk of transgenic mammals
US4956288A (en) 1988-04-22 1990-09-11 Biogen, Inc. Method for producing cells containing stably integrated foreign DNA at a high copy number, the cells produced by this method, and the use of these cells to produce the polypeptides coded for by the foreign DNA
US5231166A (en) 1988-10-25 1993-07-27 Takeda Chemical Industries, Ltd. Endothelin
US5116964A (en) 1989-02-23 1992-05-26 Genentech, Inc. Hybrid immunoglobulins
US5225538A (en) 1989-02-23 1993-07-06 Genentech, Inc. Lymphocyte homing receptor/immunoglobulin fusion proteins
US5266491A (en) 1989-03-14 1993-11-30 Mochida Pharmaceutical Co., Ltd. DNA fragment and expression plasmid containing the DNA fragment
CA2064915C (en) 1989-08-07 2001-05-29 Deborah A. Rathjen Tumour necrosis factor binding ligands
US5580575A (en) 1989-12-22 1996-12-03 Imarx Pharmaceutical Corp. Therapeutic drug delivery systems
US5210026A (en) 1990-08-20 1993-05-11 American Cyanamid Company Human mk gene and method of expression
GB9022648D0 (en) 1990-10-18 1990-11-28 Charing Cross Sunley Research Polypeptide and its use
US5268465A (en) 1991-01-18 1993-12-07 The Johns Hopkins University Purification and molecular cloning of nitric oxide synthase
US5404871A (en) 1991-03-05 1995-04-11 Aradigm Delivery of aerosol medications for inspiration
SG89295A1 (en) 1991-03-15 2002-06-18 Amgen Inc Pegylation of polypeptides
US5834229A (en) 1991-05-24 1998-11-10 Genentech, Inc. Nucleic acids vectors and host cells encoding and expressing heregulin 2-α
IL101943A0 (en) 1991-05-24 1992-12-30 Genentech Inc Structure,production and use of heregulin
US5637481A (en) * 1993-02-01 1997-06-10 Bristol-Myers Squibb Company Expression vectors encoding bispecific fusion proteins and methods of producing biologically active bispecific fusion proteins in a mammalian cell
DK0592540T3 (en) 1991-07-02 2000-06-26 Inhale Inc Method and apparatus for dispensing aerosolized drugs
US5976835A (en) 1992-03-19 1999-11-02 The Rockefeller University Nucleic acids encoding receptor recognition factor Stat1α and Stat1β, and methods of use thereof
US5447851B1 (en) 1992-04-02 1999-07-06 Univ Texas System Board Of Dna encoding a chimeric polypeptide comprising the extracellular domain of tnf receptor fused to igg vectors and host cells
US5315000A (en) 1992-05-14 1994-05-24 Children's Hospital Medical Center Gene encoding for a L5/3 growth factor and its CDNA
AU4829593A (en) 1992-09-23 1994-04-12 Fisons Plc Inhalation device
ES2127837T3 (en) 1992-10-19 1999-05-01 Dura Pharma Inc INHALER FOR DRY POWDER.
US5643252A (en) 1992-10-28 1997-07-01 Venisect, Inc. Laser perforator
US6143289A (en) 1992-12-09 2000-11-07 Indiana University Foundation Compositions of myeloid-cell-suppressive chemokines
US5849695A (en) 1993-01-13 1998-12-15 The Regents Of The University Of California Parathyroid hormone analogues useful for treatment of osteoporosis and disorders of calcium meatabolism in mammals
DK0680451T3 (en) 1993-01-19 1999-07-19 Glaxo Group Ltd Aerosol dispenser as well as process of its manufacture
US6482919B2 (en) * 1993-02-01 2002-11-19 Bristol-Myers Squibb Company Expression vectors encoding bispecific fusion proteins and methods of producing biologically active bispecific fusion proteins in a mammalian cell
ATE453709T1 (en) 1993-02-12 2010-01-15 Univ Leland Stanford Junior REGULATED TRANSCRIPTION OF TARGETED GENES AND OTHER BIOLOGICAL EVENTS
US5514670A (en) 1993-08-13 1996-05-07 Pharmos Corporation Submicron emulsions for delivery of peptides
US5814599A (en) 1995-08-04 1998-09-29 Massachusetts Insitiute Of Technology Transdermal delivery of encapsulated drugs
US5773569A (en) 1993-11-19 1998-06-30 Affymax Technologies N.V. Compounds and peptides that bind to the erythropoietin receptor
US5981478A (en) 1993-11-24 1999-11-09 La Jolla Cancer Research Foundation Integrin-binding peptides
US6190655B1 (en) 1993-12-03 2001-02-20 Immunex Corporation Methods of using Flt-3 ligand for exogenous gene transfer
US5827690A (en) 1993-12-20 1998-10-27 Genzyme Transgenics Corporatiion Transgenic production of antibodies in milk
US5608035A (en) * 1994-02-02 1997-03-04 Affymax Technologies N.V. Peptides and compounds that bind to the IL-1 receptor
US5786331A (en) 1994-02-02 1998-07-28 Affymax Technologies N.V. Peptides and compounds that bind to the IL-1 receptor
US5880096A (en) 1994-02-02 1999-03-09 Affymax Technologies N.V. Peptides and compounds that bind to the IL-1 receptor
US6174995B1 (en) 1994-08-23 2001-01-16 Haodong Li Human chemokines, CKβ4 and CKβ10/MCP-4
US5767247A (en) 1994-11-11 1998-06-16 Noboru Kaneko Anti-annexin-V monoclonal antibodies, and preparation and use thereof
US5730723A (en) 1995-10-10 1998-03-24 Visionary Medical Products Corporation, Inc. Gas pressured needle-less injection device and method
US5767078A (en) 1995-06-07 1998-06-16 Johnson; Dana L. Agonist peptide dimers
US5817750A (en) 1995-08-28 1998-10-06 La Jolla Cancer Research Foundation Structural mimics of RGD-binding sites
US6284535B1 (en) 1995-09-07 2001-09-04 The Trustees Of Columbia University In The City Of New York Splice variants of the heregulin gene, nARIA and uses thereof
GB9526100D0 (en) 1995-12-20 1996-02-21 Intersurgical Ltd Nebulizer
WO1997025086A2 (en) 1996-01-03 1997-07-17 Glaxo Group Limited Inhalation device
US5700774A (en) 1996-03-26 1997-12-23 Genetics Institute, Inc. Compositions comprising bone morphogenic proteins and truncated parathyroid hormone related peptide, and methods of inducing cartilage by administration of same
IL118003A0 (en) 1996-04-23 1996-08-04 Yeda Res & Dev Novel vip fragments and pharmaceutical compositions comprising them
US6271347B1 (en) 1996-04-26 2001-08-07 Merck & Co., Inc. Eosinophil eotaxin receptor
DE69737229T2 (en) 1996-06-07 2008-01-31 Takeda Pharmaceutical Co. Ltd. PEPTIDE WITH CORTISTATIN OR SOMATOSTATIN ACTIVITY, METHOD FOR THE PRODUCTION THEREOF AND ITS USE
ES2182088T3 (en) 1996-06-10 2003-03-01 Sucampo Ag ENDOTHELINE ANTAGONIST.
US6126939A (en) 1996-09-03 2000-10-03 Yeda Research And Development Co. Ltd. Anti-inflammatory dipeptide and pharmaceutical composition thereof
WO1998009940A1 (en) 1996-09-04 1998-03-12 Warner-Lambert Company Biphenyl butyric acids and their derivatives as inhibitors of matrix metalloproteinases
US6030780A (en) 1996-10-15 2000-02-29 The Rockefeller University Purified Stat proteins and methods of purifying thereof
IL129851A0 (en) 1996-11-15 2000-02-29 Genentech Inc Purification of neurotrophins
JP2001504830A (en) 1996-11-19 2001-04-10 ジョージタウン ユニバーシティ Heregulin antagonist and use thereof
US5948892A (en) 1996-12-16 1999-09-07 Amgen Inc. Analogs of macrophage stimulating protein
US5879681A (en) 1997-02-07 1999-03-09 Emisphere Technolgies Inc. Compounds and compositions for delivering active agents
US6136558A (en) 1997-02-10 2000-10-24 Genentech, Inc. Heregulin variants
US5921447A (en) 1997-02-13 1999-07-13 Glaxo Wellcome Inc. Flow-through metered aerosol dispensing apparatus and method of use thereof
IT1292017B1 (en) 1997-05-28 1999-01-25 Dompe Spa PHARMACEUTICAL COMPOSITIONS CONTAINING MACROPHAGE STIMULATING PROTEIN FOR THE TREATMENT OF NERVOUS SYSTEM PATHOLOGIES
IL120943A (en) 1997-05-29 2004-03-28 Univ Ben Gurion Transdermal delivery system
US6265535B1 (en) 1997-05-30 2001-07-24 The Trustees Of The University Of Pennsylvania Peptides and peptide analogues designed from binding sites of tumor necrosis factor receptor superfamily and their uses
AR016551A1 (en) 1997-07-30 2001-07-25 Smithkline Beecham Corp DERIVATIVES OF 2-OXINDOL, PHARMACEUTICAL COMPOSITIONS THAT INCLUDE THEM AND THE USE OF THE SAME FOR THE MANUFACTURE OF MEDICINES
US6180084B1 (en) * 1998-08-25 2001-01-30 The Burnham Institute NGR receptor and methods of identifying tumor homing molecules that home to angiogenic vasculature using same
US6660843B1 (en) * 1998-10-23 2003-12-09 Amgen Inc. Modified peptides as therapeutic agents
DK1642910T3 (en) * 2000-12-05 2012-05-07 Alexion Pharma Inc Rationally designed antibodies
DE60139430D1 (en) * 2000-12-07 2009-09-10 Lilly Co Eli GLP-1 fusion proteins
EP1572079A4 (en) * 2002-03-29 2006-09-06 Centocor Inc Mammalian cdr mimetibodies, compositions, methods and uses

Also Published As

Publication number Publication date
JP2006504406A (en) 2006-02-09
EP1545608A2 (en) 2005-06-29
AU2003280130B2 (en) 2009-06-11
WO2004002417A3 (en) 2004-11-04
AU2003280130A1 (en) 2004-01-19
EP1545608A4 (en) 2006-09-13
CA2490409A1 (en) 2004-01-08
WO2004002417A2 (en) 2004-01-08

Similar Documents

Publication Publication Date Title
CN1735433A (en) Mammalian EPO mimetic CH1 deleted mimetibodies, compositions, methods and uses
CN1894275A (en) Human EPO mimetic hinge core mimetibodies, compositions, methods and uses
CN1890383A (en) Hinge core mimetibodies, compositions, methods and uses
US7241733B2 (en) Mammalian EPO mimetic CH1 deleted mimetibodies, compositions, methods and uses
US7393662B2 (en) Human EPO mimetic hinge core mimetibodies, compositions, methods and uses
DK2356269T3 (en) FIBRONECTIN TYPE III DOMAIN-BASED SCAFFOLD COMPOSITIONS, PROCEDURES AND APPLICATIONS
CN101415439A (en) Glp-1 agonists, compositions, methods and uses
JP2007508011A (en) Human hinge core mimetibody, compositions, methods and uses
CN1612750A (en) TACI-immunoglobulin fusion proteins
CN1426467A (en) Thrombopoietin receptor modulating peptide
CN1795207A (en) Anti-amyloid antibodies, compositions, methods and uses
CN101389346A (en) Human glp-1 mimetibodies, compositions, methods and uses
CN101426519A (en) Human GLP-1 mimetibodies, compositions, methods and uses
MXPA06003677A (en) Human epo mimetic hinge core mimetibodies, compositions, methods and uses
AU2011202563A1 (en) Human EPO mimetic hinge core mimetibodies, compositions, methods and uses

Legal Events

Date Code Title Description
C06 Publication
PB01 Publication
C10 Entry into substantive examination
SE01 Entry into force of request for substantive examination
C02 Deemed withdrawal of patent application after publication (patent law 2001)
WD01 Invention patent application deemed withdrawn after publication