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1. 发明公开

EP1685245A4 A PROCESS OF MAKING TRANSGENIC MAMMALS THAT PRODUCE EXOGENOUS PROTEINS IN MILK AND TRANSGENIC MAMMALS PRODUCED THEREBY 审中-公开
标题翻译：法生产转基因哺乳动物中，外源蛋白质在牛奶的生产，而且生产的转基因哺乳动物
公开(公告)号：EP1685245A4
公开(公告)日：2007-01-03
申请号：EP04789272
申请日：2004-09-29
申请人： STERRENBELD BIOTECHNOLOGIE NORTH AMERICA INC
发明人： MELO CARLOS A , BARANAO LINO
IPC分类号： C12N15/09 , A01K67/027 , C07K1/00 , C07K14/00 , C07K14/61 , C07K16/00 , C07K17/00 , C12N20060101 , C12N15/00 , C12N15/63 , C12N15/79 , C12N15/85 , C12N15/877 , C12P21/00
CPC分类号： A01K67/0275 , A01K2217/05 , A01K2227/101 , A01K2267/01 , C07K14/61 , C12N15/8509 , C12N15/8771 , C12N2830/008
摘要： The invention relates to a method of producing a protein of interest, comprising making a non-human transgenic mammal that produces said protein in its milk, obtaining said mild from the non-human transgenic mammal and purifying said protein of interest from the milk. Transgenic bovine animals were generated, which are able to produce human growth hormone in mammary glands. The method involves cloning of a genetic construct encoding hGH gene and beta casein promoter conveniently in an expression vector. It also includes transfection procedures into fetal bovine somatic cells, generally fibroblasts, and the nuclear transfer into enucleated bovine oocytes, generating thus transgenic embryos. The method also includes other procedures to generate transgenic embryos for the further expansion of the transgenic herd, such as the subcloning of transgenic female bovines, the superovulation of transgenic cows and their insemination with semen from a non-transgenic or a transgenic male bovine, and the superovulation of non-transgenic cows and their insemination with semen from a transgenic male bovine. Afterwards, transgenic embryos give rise to transgenic cattle that produce human growth hormone in huge amounts in their milk, from which the hormone is completely purified and analysed to fulfill all the requirements for the manufacture of a pure biopharmaceutical product.

2. 发明公开

EP1441682A1 METHOD TO INDUCE NEOVASCULAR FORMATION AND TISSUE REGENERATION 有权
标题翻译： VEGF-A诱导的心肌再生
公开(公告)号：EP1441682A1
公开(公告)日：2004-08-04
申请号：EP02729151.7
申请日：2002-05-13
申请人： Sterrenbeld Biotechnologie North America, Inc. , Fundacion Universitaria DR Rene G. Favaloro
发明人： MELO, Carlos Alberto, , JANAVEL, Gustavo Vera, , LAGUENS, Rubén, , CROTTOGINI, José Alberto, , ARGUELLES, Maracelo Luis, , PICHEL, Ricardo Horacia, , CRISCUOLO, Marcelo Eduardo,
IPC分类号： A61K6/00
CPC分类号： A61K38/1866 , A61K9/0019 , A61K9/127 , A61K48/0058 , A61K48/0075
摘要： The present invention refers to a method for inducing neovascular formation and/or tissue regeneration in a subject. The method comprises the administration of a plasmid vector comprising a nucleotide sequence encoding for the active site of the vascular endothelial growth factor (VEGF) or a polypeptide including the active site of VEGF. The method induces cell mitosis, myocardiogenesis, arteriogenesis, vasculogenesis and lymphangiogenesis in mammalian tissues.

3. 发明公开

EP2061896A1 PROCESS FOR PRODUCING EXOGENOUS PROTEIN IN THE MILK OF TRANSGENIC MAMMALS 审中-公开
标题翻译：工艺进行生产外源蛋白在转基因哺乳动物乳汁
公开(公告)号：EP2061896A1
公开(公告)日：2009-05-27
申请号：EP08768440.3
申请日：2008-06-13
申请人： Sterrenbeld Biotechnologie North America, Inc.
发明人： BERCOVICH, Andrés , PRYNC, Aida , MELO, Carlos , FERNANDEZ, Nahuel , JUDEWICZ, Norberto , CRISCUOLO, Marcelo
IPC分类号： C12P21/02 , B01D15/04
CPC分类号： A01K67/0278 , A01K2207/15 , A01K2217/00 , A01K2227/101 , A01K2267/01 , C07K14/62 , C12N15/8509 , C12N15/8771 , C12N2830/008
摘要： The invention relates to a non-human transgenic mammal that is useful for the production of a protein of interest that may be toxic to the mammal. The mammal is characterized by the fact that it is transgenic for the production in its milk of an inactive form of the protein of interest, preferably recombinant human insulin. It is not possible to produce recombinant human insulin in transgenic mammals since this molecule has a certain degree of biological activity in the mammals and could be toxic to the mammal. Thus, the invention involves cloning a genetic construct comprising a sequence encoding a modified human insulin precursor under the control of a beta casein promoter in an expression vector. It also involves transfecting the expression plasmid into fetal bovine somatic cells, such as fibroblasts, and enucleating bovine oocytes by nuclear transfer to generate transgenic embryos. The invention gives rise to transgenic bovine that will be able to produce a modified human insulin precursor in their mammary glands. Afterwards, the milk of these transgenic mammals can be collected, the modified human insulin precursor can be converted in vitro into recombinant human insulin, and the recombinant human insulin can be purified to homogeneity as a pure biopharmaceutical product.

4. 发明公开

EP1687421A2 A PROCESS FOR PRODUCING EXOGENOUS PROTEIN IN THE MILK OF TRANGENIC MAMMALS AND A PROCESS FOR PURIFYING PROTEINS THEREFROM 有权
标题翻译： PROCESS外源蛋白的生产转基因哺乳动物和方法牛奶提纯蛋白THEREOF
公开(公告)号：EP1687421A2
公开(公告)日：2006-08-09
申请号：EP04789169.2
申请日：2004-09-29
申请人： Sterrenbeld Biotechnologie North America, Inc.
发明人： MELO, Carlos, Alberto , BARANAO, Lino , CARBONNETTO, Cesar
IPC分类号： C12N15/00 , C12P21/00 , A01K67/027 , C07K1/00 , C07K14/00 , C07K16/00 , C07K17/00
CPC分类号： A01K67/0275 , A01K2217/05 , A01K2227/101 , A01K2267/01 , C07K14/61 , C12N15/8509 , C12N15/8771 , C12N2830/008
摘要： The invention relates to a method of producing a protein of interest, comprising making a non-human transgenic mammal that produces said protein in its milk, obtaining said milk from the non-human transgenic mammal and purifying said protein of interest from the milk. Transgenic bovine animals were generated, which are able to produce human growth hormone in mammary glands. The method involves cloning of a genetic construct encoding hGH gene and beta casein promoter conveniently in an expression vector. It also includes transfection procedures into fetal bovine somatic cells, generally fibroblasts, and the nuclear transfer into enucleated bovine oocytes, generating thus transgenic embryos. The method also includes other procedures to generate transgenic embryos for the further expansion of the transgenic herd, such as the subcloning of transgenic female bovines, the superovulation of transgenic cows and their insemination with semen from a non-transgenic or a transgenic male bovine, and the superovulation of non-transgenic cows and their insemination with semen from a transgenic male bovine Afterwards, transgenic embryos give rise to transgenic cattle that produce human growth hormone in huge amounts in their milk, from which the hormone is completely purified and analysed to fulfill all the requirements for the manufacture of a pure biopharmaceutical product.

5. 发明公开

EP1127104A1 METHOD FOR THE MASSIVE CULTURE OF CELLS PRODUCING RECOMBINANT HUMAN ERYTHROPOIETIN 有权
标题翻译：方法对细胞进行大量培养重组人红细胞生成素PRODUCE
公开(公告)号：EP1127104A1
公开(公告)日：2001-08-29
申请号：EP99958810.6
申请日：1999-11-08
申请人： Sterrenbeld Biotechnologie North America, Inc.
发明人： CARCAGNO, Carlos, Miguel , CRISCUOLO, Marcelo , MELO, Carlos , VIDAL, Juan, Alejandro
IPC分类号： C12N5/00 , C12N5/02 , A01N63/00 , A01N1/02
CPC分类号： C07K14/505
摘要： The present invention relates, in general, to a method for the massive culture of recombinant mammalian cells for the production of recombinant human erythropoietin (EPO) in culture medium containing insulin. The present invention also refers to a method of producing EPO and to the EPO thus produced.

6. 发明授权

EP1687421B1 A PROCESS FOR PRODUCING EXOGENOUS PROTEIN IN THE MILK OF TRANSGENIC MAMMALS AND A PROCESS FOR PURIFYING PROTEINS THEREFROM 有权
公开(公告)号：EP1687421B1
公开(公告)日：2018-06-27
申请号：EP04789169.2
申请日：2004-09-29
申请人： Sterrenbeld Biotechnologie North America, Inc.
发明人： MELO, Carlos, Alberto , BARANAO, Lino , CARBONNETTO, Cesar
IPC分类号： C12N15/09
CPC分类号： A01K67/0275 , A01K2217/05 , A01K2227/101 , A01K2267/01 , C07K14/61 , C12N15/8509 , C12N15/8771 , C12N2830/008
摘要： The invention relates to a method of producing a protein of interest, comprising making a non-human transgenic mammal that produces said protein in its milk, obtaining said milk from the non-human transgenic mammal and purifying said protein of interest from the milk. Transgenic bovine animals were generated, which are able to produce human growth hormone in mammary glands. The method involves cloning of a genetic construct comprising the hGH gene and beta casein promoter sequences conveniently in an expression vector. It also includes transfection procedures into fetal bovine somatic cells, generally fibroblasts, and the nuclear transfer into enucleated bovine oocytes, generating thus transgenic embryos. The method also includes other procedures to generate transgenic embryos for further expansion of the transgenic herd, such as the subcloning of transgenic female bovines, the superovulation of transgenic cows and their insemination with semen from a non-transgenic or a transgenic male bovine, and the superovulation of non-transgenic cows and their insemination with semen from a transgenic male bovine. Afterwards, transgenic embryos give rise to transgenic cattle that produce human growth hormone in huge amounts in their milk, from which the hormone is completely purified and analysed to fulfill all the requirements for the manufacture of a pure biopharmaceutical product.

7. 发明授权

EP1124984B1 HOST CELLS EXPRESSING RECOMBINANT HUMAN ERYTHROPOIETIN 有权
标题翻译：宿主细胞中表达的重组人红细胞生成素
公开(公告)号：EP1124984B1
公开(公告)日：2009-01-07
申请号：EP99971863.8
申请日：1999-11-08
申请人： Sterrenbeld Biotechnologie North America, Inc.
发明人： CARCAGNO, Carlos, Miguel , CRISCUOLO, Marcelo , MELO, Carlos , VIDAL, Juan, Alejandro
IPC分类号： C12P21/02 , C12N15/19 , C07K14/505 , C12N5/06 , C12N15/63
CPC分类号： C07K14/505
摘要： The gene coding for human erythropoietin (EPO) was obtained from human genomic DNA. The gene used does not include sequences from regions at i 5' of the first translated ATG and ii 3' of the stop codon of the EPO gene. The gene was cloned into an expression plasmid for eukaryotic cells that have as sole expression control elements the early promoter of the SV40 virus and its polyadenylation signal. Recombinant cells resulting from transfection with genetic constructs used provide an unexpectedly high level of protein expression of 50 mg of recombinant EPO per liter of culture medium per day.

8. 发明授权

EP1127063B1 METHODS OF PURIFYING RECOMBINANT HUMAN ERYTHROPOIETIN FROM CELL CULTURE SUPERNATANTS 有权
标题翻译：重组人红细胞生成素纯化方法从细胞培养制品
公开(公告)号：EP1127063B1
公开(公告)日：2008-08-06
申请号：EP99958811.4
申请日：1999-11-08
申请人： Sterrenbeld Biotechnologie North America, Inc.
发明人： Carcagno, Carlos Miguel , Criscuolo, Marcelo , Melo, Carlos , Vidal, Juan Alejandro
IPC分类号： C07K1/14 , C07K1/18 , C07K1/20 , C07K1/30 , C07K1/34 , C07K1/36 , C07K14/505
CPC分类号： C07K14/505
摘要： The present invention relates, in general, to a method of purifying recombinant human erythropoietin (EPO). The present invention also relates to a substantially pure EPO. The method comprises a differential precipitation, an hydrophobic interaction chromatography, various concentration and diafiltration steps, tandem anionic and cationic exchange chromatographies and molecular exclusion chromatography for the obtaining of pure EPO. The method does not comprise high performance liquid chromatography steps. The invention also comprises the EPO obtained according to the claimed procedure.

9. 发明公开

EP1687421A4 A PROCESS FOR PRODUCING EXOGENOUS PROTEIN IN THE MILK OF TRANGENIC MAMMALS AND A PROCESS FOR PURIFYING PROTEINS THEREFROM 有权
标题翻译： VERFAHREN ZUR在DER MILCH转基因生产VON EXOGENEM蛋白质SÄUGERSOWIE VERFAHREN ZUR AUFREINIGUNG VON PROTEINEN DARAUS
公开(公告)号：EP1687421A4
公开(公告)日：2006-12-20
申请号：EP04789169
申请日：2004-09-29
申请人： STERRENBELD BIOTECHNOLOGIE NORTH AMERICA INC
发明人： MELO CARLOS ALBERTO , BARANAO LINO , CARBONNETTO CESAR
IPC分类号： C12N15/09 , A01K67/027 , C07K1/00 , C07K14/00 , C07K14/61 , C07K16/00 , C07K17/00 , C12N20060101 , C12N15/00 , C12N15/85 , C12N15/877 , C12P21/00
CPC分类号： A01K67/0275 , A01K2217/05 , A01K2227/101 , A01K2267/01 , C07K14/61 , C12N15/8509 , C12N15/8771 , C12N2830/008
摘要： The invention relates to a method of producing a protein of interest, comprising making a non-human transgenic mammal that produces said protein in its milk, obtaining said milk from the non-human transgenic mammal and purifying said protein of interest from the milk. Transgenic bovine animals were generated, which are able to produce human growth hormone in mammary glands. The method involves cloning of a genetic construct comprising the hGH gene and beta casein promoter sequences conveniently in an expression vector. It also includes transfection procedures into fetal bovine somatic cells, generally fibroblasts, and the nuclear transfer into enucleated bovine oocytes, generating thus transgenic embryos. The method also includes other procedures to generate transgenic embryos for further expansion of the transgenic herd, such as the subcloning of transgenic female bovines, the superovulation of transgenic cows and their insemination with semen from a non-transgenic or a transgenic male bovine, and the superovulation of non-transgenic cows and their insemination with semen from a transgenic male bovine. Afterwards, transgenic embryos give rise to transgenic cattle that produce human growth hormone in huge amounts in their milk, from which the hormone is completely purified and analysed to fulfill all the requirements for the manufacture of a pure biopharmaceutical product.
摘要翻译：本发明涉及生产感兴趣的蛋白质的方法，该方法包括制造在其牛奶中产生所述蛋白质的非人转基因哺乳动物，从非人转基因哺乳动物获得所述乳并从乳中纯化所述感兴趣的蛋白质。产生转基因牛动物，其能够在乳腺中产生人生长激素。该方法包括方便地在表达载体中克隆编码hGH基因和β酪蛋白启动子的遗传构建体。它还包括转染胎儿牛体细胞，通常是成纤维细胞的转染过程，以及核转移到去核的牛卵母细胞中，从而产生转基因胚胎。该方法还包括产生用于进一步扩增转基因畜群的转基因胚胎的其他程序，例如转基因雌性牛的亚克隆，转基因母牛的超数排卵以及它们与来自非转基因雄性牛或转基因雄性牛的精液的授精，以及非转基因母牛的超数排卵及其与来自转基因雄性牛的精液的授精之后，转基因胚胎产生了在其牛奶中产生大量人类生长激素的转基因牛，从中激素被完全纯化并分析以实现所有制造纯生物制药产品的要求。

10. 发明公开

EP2061895A1 TRANSGENIC MAMMALS THAT PRODUCE EXOGENOUS PROTEINS IN MILK 审中-公开
标题翻译：转基因动物在牛奶外源蛋白质生产
公开(公告)号：EP2061895A1
公开(公告)日：2009-05-27
申请号：EP08768438.7
申请日：2008-06-13
申请人： Sterrenbeld Biotechnologie North America, Inc.
发明人： BERCOVICH, Andrés , PRYNC, Aida , FERNANDEZ, Nahuel , MELO, Carlos , CRISCUOLO, Marcelo
IPC分类号： C12P21/02 , C12N15/00 , C12Q1/68
CPC分类号： A01K67/0278 , A01K2207/15 , A01K2217/00 , A01K2227/101 , A01K2267/01 , C07K14/62 , C12N15/8509 , C12N15/8771 , C12N2830/008
摘要： The invention relates to a non-human transgenic mammal that is useful for the production of a protein of interest that may be toxic to the mammal. The mammal is characterized by the fact that it is transgenic for the production in its milk of an inactive form of the protein of interest, preferably recombinant human insulin. It is not possible to produce recombinant human insulin in transgenic mammals since this molecule has a certain degree of biological activity in the mammals and could be toxic to the mammal. Thus, the invention involves cloning a genetic construct comprising a sequence encoding a modified human insulin precursor under the control of a beta casein promoter in an expression vector. It also involves transfecting the expression plasmid into fetal bovine somatic cells, such as fibroblasts, and enucleating bovine oocytes by nuclear transfer to generate transgenic embryos. The invention gives rise to transgenic bovine that will be able to produce a modified human insulin precursor in their mammary glands. Afterwards, the milk of these transgenic mammals can be collected, the modified human insulin precursor can be converted in vitro into recombinant human insulin, and the recombinant human insulin can be purified to homogeneity as a pure biopharmaceutical product.

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