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
Priority claimed from US06/677,257external-prioritypatent/US4650674A/en
Application filed by Genentech, Inc.filedCriticalGenentech, Inc.
Priority claimed from YU113285Aexternal-prioritypatent/YU47968B/en
Publication of SI8511132ApublicationCriticalpatent/SI8511132A/en
Publication of SI8511132BpublicationCriticalpatent/SI8511132B/en
C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
C—CHEMISTRY; METALLURGY
C07—ORGANIC CHEMISTRY
C07K—PEPTIDES
C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
C07K14/52—Cytokines; Lymphokines; Interferons
C07K14/525—Tumour necrosis factor [TNF]
C07K14/5255—Lymphotoxin [LT]
C—CHEMISTRY; METALLURGY
C07—ORGANIC CHEMISTRY
C07K—PEPTIDES
C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
C07K14/52—Cytokines; Lymphokines; Interferons
C07K14/525—Tumour necrosis factor [TNF]
G—PHYSICS
G01—MEASURING; TESTING
G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
G01N33/6863—Cytokines, i.e. immune system proteins modifying a biological response such as cell growth proliferation or differentiation, e.g. TNF, CNF, GM-CSF, lymphotoxin, MIF or their receptors
A—HUMAN NECESSITIES
A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
C07K2319/01—Fusion polypeptide containing a localisation/targetting motif
C07K2319/02—Fusion polypeptide containing a localisation/targetting motif containing a signal sequence
Landscapes
Health & Medical Sciences
(AREA)
Life Sciences & Earth Sciences
(AREA)
Chemical & Material Sciences
(AREA)
Molecular Biology
(AREA)
Engineering & Computer Science
(AREA)
Organic Chemistry
(AREA)
Genetics & Genomics
(AREA)
Zoology
(AREA)
Biomedical Technology
(AREA)
Biochemistry
(AREA)
General Health & Medical Sciences
(AREA)
Biotechnology
(AREA)
Biophysics
(AREA)
Medicinal Chemistry
(AREA)
Proteomics, Peptides & Aminoacids
(AREA)
Immunology
(AREA)
Wood Science & Technology
(AREA)
Bioinformatics & Cheminformatics
(AREA)
Toxicology
(AREA)
Hematology
(AREA)
Microbiology
(AREA)
Urology & Nephrology
(AREA)
Cell Biology
(AREA)
Physics & Mathematics
(AREA)
Gastroenterology & Hepatology
(AREA)
General Engineering & Computer Science
(AREA)
Pathology
(AREA)
General Physics & Mathematics
(AREA)
Plant Pathology
(AREA)
Analytical Chemistry
(AREA)
Food Science & Technology
(AREA)
Medicines That Contain Protein Lipid Enzymes And Other Medicines
(AREA)
Micro-Organisms Or Cultivation Processes Thereof
(AREA)
Peptides Or Proteins
(AREA)
Preparation Of Compounds By Using Micro-Organisms
(AREA)
Medicines Containing Material From Animals Or Micro-Organisms
(AREA)
Claims (33)
57 57 / PATENTNI ZAHTEVKI 1. Postopek za pripravo faktorja nekroze tumorja iz zmesi z drugimi proteini, označen s tem, da obsega adsorbiranje faktorja nekroze tumorja iz take zmesi na substanco, izbrano iz skupine, ki jo sestavljajo silikat, steklo s kontroliranimi porami, alkil sefaroza in delci anionske izmenjalne smole, ki imajo v glavnem enakomerno velikost delcev, in nato eluiranje faktorja nekroze tumorja.A process for the preparation of a tumor necrosis factor from a mixture with other proteins, characterized in that it comprises adsorbing the tumor necrosis factor from such a mixture onto a substance selected from the group consisting of silicate, controlled pore glass, alkyl sepharose and anion exchange resin particles having substantially uniform particle size, and then elution of tumor necrosis factor.2. Postopek po zahtevku 1, označen s tem, da faktor nekroze tumorja eluiramo iz stekla s kontroliranimi porami z etilen glikolom.Process according to Claim 1, characterized in that the tumor necrosis factor is eluted from the controlled pore glass with ethylene glycol.3. Postopek po zahtevku 1, označen s tem, da kot anionsko izmenjalno smolo uporabimo premrežen polistiren, substituiran s kvaternim aminom.Process according to Claim 1, characterized in that crosslinked polystyrene substituted with a quaternary amine is used as the anion exchange resin.4. Faktor nekroze humanega tumorja, označen s tem, daje: a) neglikoziliran; b) ima molekulsko maso približno 17000, kot je določeno z SDS-PAGE; c) sposoben za preferenčno destrukcijo ali inhibicijo rasti tumorskih celic v primerjavi z normalnimi celicami pri istih pogojih; d) v bistvu homogen, kot je določeno z SDS-PAGE; ima specifično aktivnost, večjo od približno 10 milijonov enot/ mg proteina; in je očiščen do stopnje, ki zadostuje za sekvenciranje intaktnega faktorja nekroze tumorja ali njegovega fragmenta, hidroliziranega s tripsinom, s sekvencialno Edmanovo razgradnjo na sekvenatorju z vrtljivo skodelico, opremljenim s hladnimi pastmi ter ob uporabi PolybrenaR kot nosilca v skodelici sekvenatorja; in e) obsega: (i) amino kislinsko sekvenco 1 do 157, navedeno na sliki 10; ali (ii) amino kislinsko sekvenco Glu Thr Pro Glu Gly Ala Glu Ala Lys Pro Trp Tyr Glu Pro; ali (iii) amino kislinsko sekvenco ostankov 35-66, prikazano na sliki 10; ali (iv) amino kislinsko sekvenco ostankov 110-133, prikazano na sliki 10; ali (v) amino kislinsko sekvenco ostankov 1-6, 7-44, 45-82, 83-90 ali 99-157, prikazano na sliki 10.4. Human tumor necrosis factor, characterized in that: a) non-glycosylated; b) has a molecular weight of about 17000, as determined by SDS-PAGE; c) capable of preferentially destroying or inhibiting the growth of tumor cells compared to normal cells under the same conditions; d) substantially homogeneous as determined by SDS-PAGE; has a specific activity greater than about 10 million units / mg protein; and purified to a degree sufficient to sequence the intact tumor necrosis factor or its trypsin-hydrolyzed fragment by sequential Edman degradation on a rotary cup sequencer equipped with cold traps and using PolybrenaR as a carrier in the sequencer cup; and e) comprising: (i) the amino acid sequence 1 to 157 shown in Figure 10; or (ii) the amino acid sequence of Glu Thr Pro Glu Gly Ala Glu Ala Lys Pro Trp Tyr Glu Pro; or (iii) the amino acid sequence of residues 35-66 shown in Figure 10; or (iv) the amino acid sequence of residues 110-133 shown in Figure 10; or (v) the amino acid sequence of residues 1-6, 7-44, 45-82, 83-90 or 99-157 shown in Figure 10.5. Faktor nekroze tumorja po zahtevku 4, označen s tem, da ne vsebuje drugih citotoksičnih proteinov. / 58Tumor necrosis factor according to claim 4, characterized in that it does not contain other cytotoxic proteins. / 586. Faktor nekroze tumorja po zahtevku 5, označen s tem, da ne vsebuje proteina, izbranega iz skupine, ki jo sestavljajo limfotoksin, alfa-globulini, serin proteaze in in-terferoni.Tumor necrosis factor according to claim 5, characterized in that it does not contain a protein selected from the group consisting of lymphotoxin, alpha-globulins, serine proteases and interferons.7. Faktor nekroze tumorja, označen s tem, da je sposoben za preferenčno destrukcijo ali inhibicijo rasti celic tumoija v primerjavi z normalnimi celicami pri istih pogojih in je sestavljen v glavnem iz amino kislinske sekvence 1-157, prikazane na sl. 10; v t danem primeru z N-terminalnim metionilnim ostankom; ali iz variante te sekvence (različne od zajčjega TNF) v kateri sta ohranjena ostanka Val1 in/ali Arg2.7. Tumor necrosis factor, characterized in that it is capable of preferentially destroying or inhibiting the growth of tumor cells compared to normal cells under the same conditions and consists mainly of amino acid sequence 1-157 shown in FIG. 10; optionally with an N-terminal methionyl residue; or from a variant of this sequence (other than rabbit TNF) in which Val1 and / or Arg2 residues are preserved.8. Faktor nekroze tumorja po zahtevku 7, označen s tem, da amino kislinski ostanek substituiramo, izpustimo ali vključimo v amino kislinski sekvenci 1 do 157, prikazani na sl. 10.Tumor necrosis factor according to claim 7, characterized in that the amino acid residue is substituted, omitted or included in amino acid sequences 1 to 157 shown in FIG. 10.9. Faktora nekroze tumorja, označen s tem, da je sposoben za preferenčno destrukcijo ali inhibicijo rasti celic tumorja v primerjavi z normalnimi celicami pri istih pogojih in obsega varianto amino kislinske sekvence 1-157, prikazano na sl. 10 (različno od zajčjega TNF) v kateri substituiramo, izpustimo ali vključimo amino kislinsko sekvenco na mestu, kije v amino kislinskih ostankih 35 do 66, 110 do 133 ali 150 do 157, kot je prikazano na sl. 10.9. Tumor necrosis factor, characterized in that it is capable of preferentially destroying or inhibiting the growth of tumor cells compared to normal cells under the same conditions and comprising the variant amino acid sequence 1-157 shown in FIG. 10 (other than rabbit TNF) in which the amino acid sequence is substituted, omitted or incorporated at a site which in amino acid residues 35 to 66, 110 to 133 or 150 to 157, as shown in FIG. 10.10. Faktora nekroze tumorja, označen s tem, daje sposoben za preferenčno destrukcijo ali inhibicijo rasti celic tumorja v primerjavi z normalimi celicami pri istih pogojih in obsega varianto amino kislinske sekvence 1-157, prikazano na sl. 10 (različno od zajčjega TNF) v kateri substituiramo, izpustimo ali vključimo amino kislinsko sekvenco na mestu, ki je v amino kislinskih ostankih 67 do 109, kot je prikazano na sl. 10.10. Tumor necrosis factor, characterized in that it is capable of preferentially destroying or inhibiting the growth of tumor cells compared to normal cells under the same conditions and comprising the variant amino acid sequence 1-157 shown in FIG. 10 (other than rabbit TNF) in which the amino acid sequence is substituted, omitted or incorporated at a site present in amino acid residues 67 to 109, as shown in FIG. 10.11. Faktor nekroze tumorja po enem od zahtevkov 8, 9 in 10, označen s tem, da izvedemo substitucijo ostankov iz vrste nevtralnih, kislih ali bazičnih amino kislinskih ostankov za ostanek amino kislinske sekvence faktorja nekroze tumorja, ki ni član razreda kateremu pripada substituirana amino kislina.Tumor necrosis factor according to one of Claims 8, 9 and 10, characterized in that the substitution of residues from the type of neutral, acidic or basic amino acid residues is performed for the residue of the amino acid sequence of the tumor necrosis factor which is not a member of the class to which the substituted amino belongs. acid.12. Faktor nekroze tumorja po enem od zahtevkov 8, 9 ali 10, označen s tem, da s substitucijo uvedemo ostanek s prostorsko masivno stransko verigo namesto ostanka, ki ne vsebuje take stranske verige. 59Tumor necrosis factor according to one of Claims 8, 9 or 10, characterized in that the residue is introduced by substitution with a spatially massive side chain instead of a residue which does not contain such a side chain. 5913. Faktor nekroze tumorja po zahtevku 12, označen s tem, da je ostanek, ki ne vsebuje prostorsko masivne stranske verige, glicin ali serin.Tumor necrosis factor according to claim 12, characterized in that the residue does not contain a spatially massive side chain, glycine or serine.14. Faktor nekroze tumorja po zahtevku 12 ali 13, označen s tem, da je ostanek, ki ima takšno stransko verigo, fenil alanin ali triptofan.Tumor necrosis factor according to claim 12 or 13, characterized in that the residue having such a side chain is phenyl alanine or tryptophan.15. Faktor nekroze tumorja po enem od zahtevkov 8, 9 ali 10, označen s tem, da je lizinski ali argininski ostanek izpuščen ali substituiran z drugim amino kislinskim ostankom, ki ni lizin ali arginin.Tumor necrosis factor according to one of Claims 8, 9 or 10, characterized in that the lysine or arginine residue is omitted or substituted by another amino acid residue other than lysine or arginine.16. Faktor nekroze tumorja po enem od zahtevkov 4 do 15, označen s tem, da je v zmesi s fiziološko sprejemljivim pufrom, amino kislinskim ali neionskim surfaktan-tom ali njihovimi zmesmi.Tumor necrosis factor according to one of Claims 4 to 15, characterized in that it is in a mixture with a physiologically acceptable buffer, an amino acid or a non-ionic surfactant or mixtures thereof.17. Faktor nekroze tumorja po zahtevku 16, označen s tem, da je liofiliziran.Tumor necrosis factor according to claim 16, characterized in that it is lyophilized.18. DNA, označena s tem, da kodira za faktor nekroze tumorja po enem od zahtevkov 7 do 15.DNA characterized by encoding a tumor necrosis factor according to one of Claims 7 to 15.19. DNA po zahtevku 18, označena s tem, daje sintetična.DNA according to claim 18, characterized in that it is synthetic.20. DNA po zahtevku 18 ali 19, označena s tem, da je prisotna v replikabilnem vektorju.DNA according to claim 18 or 19, characterized in that it is present in a replicable vector.21. DNA po zahtevku 20, označena stem, da je vektor plazmid ali virus.The DNA of claim 20, wherein the vector is a plasmid or virus.22. Celica, označena s tem, daje transformirana z DNA po zahtevku 20 ali 21.A cell characterized in that it is transformed with DNA according to claim 20 or 21.23. Postopek, označen s tem, da obsega kultiviranje celice, transformirane z DNA po zahtevku 18, tako da povzroči ekspresijo te DNA, da omogoči akumuliranje faktorja nekroze tumorja v kulturi in njegovo regeneriranje iz kulture.The method, which comprises culturing the cell transformed with DNA according to claim 18, so as to cause the expression of this DNA to allow the accumulation of tumor necrosis factor in the culture and its regeneration from the culture.24. Pripravek, označen s tem, da obsega faktor nekroze tumorja po enem od zahtevkov 4 do 15 in komponento iz nehumane celice, ki je fiziološko sprejemljiva po dajanju pacientom. * 60 /Composition characterized in that it comprises a tumor necrosis factor according to one of Claims 4 to 15 and a non-human cell component which is physiologically acceptable after administration to patients. * 60 /25. Pripravek po zahtevku 24, označen s tem, da je navedena komponenta prokariotski protein.The composition of claim 24, wherein said component is a prokaryotic protein.26. Pripravek, označen s tem, da obsega celice, ki vsebujejo heterologni faktor nekroze tumorja po enem od zahtevkov 7 do 15.Composition characterized in that it comprises cells containing a heterologous tumor necrosis factor according to one of Claims 7 to 15.27. Pripravek po zahtevku 26, označen s tem, da so celice prokariotske ali nižje ev-kariotske.The composition of claim 26, wherein the cells are prokaryotic or lower eukaryotic.28. Pripravek, primeren za zdravljenje tumorjev, označen s tem, da obsega faktor nekroze tumorja po enem od zahtevkov 4 do 17 ali dobljen s postopkom po enem od zahtevkov 1 do 3 in interferon.Composition suitable for the treatment of tumors, characterized in that it comprises a tumor necrosis factor according to one of Claims 4 to 17 or obtained by a method according to one of Claims 1 to 3 and interferon.29. Pripravek po zahtevku 28, označen s tem, da ne obsega drugih citotoksičnih pep-tidov.Preparation according to Claim 28, characterized in that it does not comprise other cytotoxic peptides.30. Pripravek po zahtevku 28 ali 29, označen s tem, da ne obsega drugih plazemskih proteinov.Preparation according to Claim 28 or 29, characterized in that it does not comprise other plasma proteins.31. Pripravek brez celic, primeren za zdravljenje tumorjev, označen s tem, da vsebuje faktor nekroze tumorja po enem od zahtevkov 4 do 17 ali dobljen s postopkom po enem od zahtevkov 1 do 3 ter interferon, ne vsebuje pa limfotoksina.Cell-free preparation suitable for the treatment of tumors, characterized in that it contains a tumor necrosis factor according to one of claims 4 to 17 or obtained by a method according to one of claims 1 to 3 and interferon, but does not contain lymphotoxin.32. Pripravek po enem od zahtevkov 28 do 31, označen s tem, daje interferon gama interferon.Preparation according to one of Claims 28 to 31, characterized in that the interferon is gamma interferon.33. Pripravek po enem od zahtevkov 28 do 32, označen s tem, daje liofiliziran. Za Genentech, Inc.:Preparation according to one of Claims 28 to 32, characterized in that it is lyophilized. For Genentech, Inc .:LJUBLJANA, ČOPOVA 14LJUBLJANA, ČOPOVA 14
Analogues of Parathyroid Hormone (PTH) and Parathyroid Hormone Associated Peptides (PTHRP), Their Synthesis and Their Uses for the Treatment of Osteoporosis (ANALOGS OF PTH AND PTHRP