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1. 发明申请

US20130330340A1 PRODUCTION OF N- AND O-SIALYLATED TNFRII-FC FUSION PROTEIN IN YEAST 审中-公开
标题翻译：在YEAST中生产N-和O-齐聚的TNFRII-FC融合蛋白
公开(公告)号：US20130330340A1
公开(公告)日：2013-12-12
申请号：US13985130
申请日：2012-02-20
申请人： Stephen R. Hamilton , W. James Cook , Sujatha Gomathinayagam
发明人： Stephen R. Hamilton , W. James Cook , Sujatha Gomathinayagam
IPC分类号： C07K14/705 , C07K16/46
CPC分类号： C07K14/70578 , C07K14/7151 , C07K16/46 , C07K2319/30 , C12N9/1051 , C12N9/2402 , C12Y204/01101 , C12Y302/01113
摘要： Production of recombinant Tumor Necrosis Factor Receptor fused to the Fc region of an antibody (TNFRII-Fc fragment fusion protein) in a glycoengineered yeast strain that is capable of producing sialylated N-glycans and O-glycans is described. Compositions of TNFRII-Fc fragment fusion protein comprising dystroglycan type O-glycans and sialylated N- and O-glycans with only terminal N-acetylneuraminic acid (NANA) residues in an α2,6-linkage are provided. In particular aspects, methods are provided for modulating the in vivo pharmacokinetics of the TNFRII-Fc fragment fusion protein by altering the O-glycan structure on the molecule.
摘要翻译：描述了能够产生唾液酸化N-聚糖和O-聚糖的糖工程化酵母菌株中与抗体Fc区（TNFRII-Fc片段融合蛋白）融合的重组肿瘤坏死因子受体的产生。提供包含赖氨酸聚糖型O-聚糖和唾液酸化N-和O-聚糖的TNFRII-Fc片段融合蛋白的组合物，其仅具有α2,6键中的末端N-乙酰神经氨酸（NANA）残基。在具体方面，提供了通过改变分子上的O-聚糖结构来调节TNFRII-Fc片段融合蛋白的体内药代动力学的方法。

2. 发明授权

US08067551B2 Combinatorial DNA library for producing modified N-glycans in lower eukaryotes 有权
公开(公告)号：US08067551B2
公开(公告)日：2011-11-29
申请号：US12291373
申请日：2008-11-07
申请人： Tillman U. Gerngross , Stefan Wildt , Byung-Kwon Choi , Juergen Hermann Nett , Piotr Bobrowicz , Stephen R Hamilton , Robert C. Davidson
发明人： Tillman U. Gerngross , Stefan Wildt , Byung-Kwon Choi , Juergen Hermann Nett , Piotr Bobrowicz , Stephen R Hamilton , Robert C. Davidson
IPC分类号： C07K14/00
CPC分类号： C12P21/005 , C07K14/47 , C12N9/1051 , C12N15/1082 , C12N15/79
摘要： The present invention relates to eukaryotic host cells having modified oligosaccharides which may be modified further by heterologous expression of a set of glycosyltransferases, sugar transporters and mannosidases to become host-strains for the production of mammalian, e.g., human therapeutic glycoproteins. The invention provides nucleic acid molecules and combinatorial libraries which can be used to successfully target and express mammalian enzymatic activities such as those involved in glycosylation to intracellular compartments in a eukaryotic host cell. The process provides an engineered host cell which can be used to express and target any desirable gene(s) involved in glycosylation. Host cells with modified oligosaccharides are created or selected. N-glycans made in the engineered host cells have a Man5GlcNAc2 core structure which may then be modified further by heterologous expression of one or more enzymes, e.g., glycosyltransferases, sugar transporters and mannosidases, to yield human-like glycoproteins. For the production of therapeutic proteins, this method may be adapted to engineer cell lines in which any desired glycosylation structure may be obtained.

3. 发明授权

US07935513B2 Combinatorial DNA library for producing modified N-glycans in lower eukaryotes 有权
标题翻译：用于在低等真核生物中生产修饰的N-聚糖的组合DNA文库
公开(公告)号：US07935513B2
公开(公告)日：2011-05-03
申请号：US12156936
申请日：2008-06-05
申请人： Tillman U. Gerngross , Stefan Wildt , Byung-Kwon Choi , Juergen Hermann Nett , Piotr Bobrowicz , Stephen R. Hamilton , Robert C. Davidson
发明人： Tillman U. Gerngross , Stefan Wildt , Byung-Kwon Choi , Juergen Hermann Nett , Piotr Bobrowicz , Stephen R. Hamilton , Robert C. Davidson
IPC分类号： C12N1/15 , C12P21/00
CPC分类号： C12P21/005 , C07K14/47 , C12N9/1051 , C12N15/1082 , C12N15/79
摘要： The present invention relates to eukaryotic host cells having modified oligosaccharides which may be modified further by heterologous expression of a set of glycosyltransferases, sugar transporters and mannosidases to become host-strains for the production of mammalian, e.g., human therapeutic glycoproteins. The invention provides nucleic acid molecules and combinatorial libraries which can be used to successfully target and express mammalian enzymatic activities such as those involved in glycosylation to intracellular compartments in a eukaryotic host cell. The process provides an engineered host cell which can be used to express and target any desirable gene(s) involved in glycosylation. Host cells with modified oligosaccharides are created or selected. N-glycans made in the engineered host cells have a Man5GlcNAc2 core structure which may then be modified further by heterologous expression of one or more enzymes, e.g., glycosyltransferases, sugar transporters and mannosidases, to yield human-like glycoproteins. For the production of therapeutic proteins, this method may be adapted to engineer cell lines in which any desired glycosylation structure may be obtained.
摘要翻译：本发明涉及具有修饰的寡糖的真核宿主细胞，其可以通过异源表达一组糖基转移酶，糖转运体和甘露糖苷酶进一步修饰，以成为用于产生哺乳动物例如人治疗性糖蛋白的宿主菌株。本发明提供核酸分子和组合文库，其可用于成功靶向和表达哺乳动物酶活性，例如参与糖基化的真核宿主细胞中的细胞内区室的那些。该方法提供了可用于表达和靶向参与糖基化的任何所需基因的工程化宿主细胞。建立或选择具有修饰寡糖的宿主细胞。在工程化宿主细胞中制备的N-聚糖具有Man5GlcNAc2核心结构，然后可以通过异源表达一种或多种酶，例如糖基转移酶，糖转运蛋白和甘露糖苷酶来进一步修饰，以产生人样糖蛋白。为了生产治疗性蛋白质，该方法可适用于工程化可能获得任何所需糖基化结构的细胞系。

4. 发明授权

US07598055B2 N-acetylglucosaminyltransferase III expression in lower eukaryotes 失效
标题翻译： N-乙酰葡糖胺基转移酶III在低等真核生物中的表达
公开(公告)号：US07598055B2
公开(公告)日：2009-10-06
申请号：US10680963
申请日：2003-10-07
申请人： Piotr Bobrowicz , Stephen R. Hamilton , Tillman U. Gerngross , Stefan Wildt , Byung-Kwon Choi , Juergen H. Nett , Robert C. Davidson
发明人： Piotr Bobrowicz , Stephen R. Hamilton , Tillman U. Gerngross , Stefan Wildt , Byung-Kwon Choi , Juergen H. Nett , Robert C. Davidson
IPC分类号： C12P21/06 , A61K38/43
CPC分类号： C07K14/39 , A61K38/00 , C07K2319/05 , C12N9/1051 , C12P21/005 , Y02A50/396
摘要： The present invention relates to eukaryotic host cells having modified oligosaccharides which may be modified further by heterologous expression of a set of glycosyltransferases, sugar transporters and mannosidases to become host-strains for the production of mammalian, e.g., human therapeutic glycoproteins. The process provides an engineered host cell which can be used to express and target any desirable gene(s) involved in glycosylation. Host cells with modified lipid-linked oligosaccharides are created or selected. N-glycans made in the engineered host cells exhibit GnTIII activity, which produce bisected N-glycan structures and may be modified further by heterologous expression of one or more enzymes, e.g., glycosyltransferases, sugar transporters and mannosidases, to yield human-like glycoproteins. For the production of therapeutic proteins, this method may be adapted to engineer cell lines in which any desired glycosylation structure may be obtained.
摘要翻译：本发明涉及具有修饰的寡糖的真核宿主细胞，其可以通过异源表达一组糖基转移酶，糖转运体和甘露糖苷酶进一步修饰，以成为用于产生哺乳动物例如人治疗性糖蛋白的宿主菌株。该方法提供了可用于表达和靶向参与糖基化的任何所需基因的工程化宿主细胞。制备或选择具有修饰的脂质连接寡糖的宿主细胞。在工程化的宿主细胞中制备的N-聚糖表现出GnTIII活性，其产生二等分的N-聚糖结构，并且可以通过异源表达一种或多种酶（例如糖基转移酶，糖转运蛋白和甘露糖苷酶）进一步修饰，以产生人样糖蛋白。为了生产治疗性蛋白质，该方法可适用于工程化可能获得任何所需糖基化结构的细胞系。

5. 发明授权

US08999671B2 Production of sialylated N-glycans in lower eukaryotes 有权
标题翻译：在低等真核生物中生产唾液酸化的N-聚糖
公开(公告)号：US08999671B2
公开(公告)日：2015-04-07
申请号：US13554126
申请日：2012-07-20
申请人： Stephen R. Hamilton
发明人： Stephen R. Hamilton
IPC分类号： C12P19/26 , C12P21/00 , C12N15/00 , C12N9/10 , C12N9/24
CPC分类号： C12P21/005 , C07K2319/036 , C07K2319/05 , C07K2319/055 , C12N9/1081 , C12N9/2402 , C12Y302/01018
摘要： The present invention relates to eukaryotic host cells which have been modified to produce sialylated glycoproteins by the heterologous expression of a set of glycosyltransferases, including sialyltransferase and/or trans-sialidase, to become host-strains for the production of mammalian, e.g., human therapeutic glycoproteins. Novel eukaryotic host cells expressing a CMP-sialic acid biosynthetic pathway for the production of sialylated glycoproteins are also provided. The invention provides nucleic acid molecules and combinatorial libraries which can be used to successfully target and express mammalian enzymatic activities (such as those involved in sialylation) to intracellular compartments in a eukaryotic host cell. The process provides an engineered host cell which can be used to express and target any desirable gene(s) involved in glycosylation.
摘要翻译：本发明涉及通过异源表达一组糖基转移酶（包括唾液酸转移酶和/或反式唾液酸酶）而被修饰以产生唾液酸化糖蛋白的真核宿主细胞，以成为用于产生哺乳动物例如人类治疗的宿主菌株糖蛋白。还提供了表达用于产生唾液酸化糖蛋白的CMP-唾液酸生物合成途径的新型真核宿主细胞。本发明提供核酸分子和组合文库，其可用于成功靶向并表达哺乳动物酶活性（例如涉及唾液酸化的那些）到真核宿主细胞中的细胞内区室。该方法提供了可用于表达和靶向参与糖基化的任何所需基因的工程化宿主细胞。

6. 发明授权

US08445227B2 N-acetylglucosaminyltransferase III expression in lower eukaryotes 有权
公开(公告)号：US08445227B2
公开(公告)日：2013-05-21
申请号：US12540849
申请日：2009-08-13
申请人： Piotr Bobrowicz , Stephen R. Hamilton , Tillman U. Gerngross , Stefan Wildt , Byung-Kwon Choi , Juergen Hermann Nett , Robert C. Davidson
发明人： Piotr Bobrowicz , Stephen R. Hamilton , Tillman U. Gerngross , Stefan Wildt , Byung-Kwon Choi , Juergen Hermann Nett , Robert C. Davidson
IPC分类号： C12N15/00
CPC分类号： C07K14/39 , A61K38/00 , C07K2319/05 , C12N9/1051 , C12P21/005 , Y02A50/396
摘要： The present invention relates to eukaryotic host cells having modified oligosaccharides which may be modified further by heterologous expression of a set of glycosyltransferases, sugar transporters and mannosidases to become host-strains for the production of mammalian, e.g., human therapeutic glycoproteins. The process provides an engineered host cell which can be used to express and target any desirable gene(s) involved in glycosylation. Host cells with modified lipid-linked oligosaccharides are created or selected. N-glycans made in the engineered host cells exhibit GnTIII activity, which produce bisected N-glycan structures and may be modified further by heterologous expression of one or more enzymes, e.g., glycosyltransferases, sugar transporters and mannosidases, to yield human-like glycoproteins. For the production of therapeutic proteins, this method may be adapted to engineer cell lines in which any desired glycosylation structure may be obtained.

7. 发明申请

US20120231502A1 METHOD FOR PRODUCING THERAPEUTIC PROTEINS IN PICHIA PASTORIS LACKING DIPEPTIDYL AMINOPEPTIDASE ACTIVITY 有权
标题翻译：在PICHIA PASTORIS中生产治疗蛋白的方法缺乏胰蛋白酶活性
公开(公告)号：US20120231502A1
公开(公告)日：2012-09-13
申请号：US13503663
申请日：2010-10-27
申请人： Stephen R. Hamilton , Terrance A. Stadheim
发明人： Stephen R. Hamilton , Terrance A. Stadheim
IPC分类号： C12P21/00 , C12N1/19
CPC分类号： C12N15/81 , C12N9/48 , C12P21/005 , C12P21/02 , Y02P20/52
摘要： The present invention related to methods and compositions for producing therapeutic proteins in yeast cell lines, and in particular Pichia pastoris, lacking dipeptidyl aminopeptidase (DAP) activity. DAP activity has been eliminated by genetically modifying a Pichia pastoris cell line such that STE13 and DAP2 have been deleted.
摘要翻译：本发明涉及用于在酵母细胞系中产生治疗性蛋白质的方法和组合物，特别是缺乏二肽基氨基肽酶（DAP）活性的巴斯德毕赤酵母。通过遗传修饰巴斯德毕赤酵母细胞系来消除DAP活性，使得STE13和DAP2被缺失。

8. 发明申请

US20110027831A1 PRODUCTION OF SIALYLATED N-GLYCANS IN LOWER EUKARYOTES 审中-公开
标题翻译：在较低的核酸中生产广泛的N-糖类
公开(公告)号：US20110027831A1
公开(公告)日：2011-02-03
申请号：US12896441
申请日：2010-10-01
申请人： STEPHEN R. HAMILTON
发明人： STEPHEN R. HAMILTON
IPC分类号： C12P19/26 , C12N1/19
CPC分类号： C12P21/005 , C07K2319/036 , C07K2319/05 , C07K2319/055 , C12N9/1081 , C12N9/2402 , C12Y302/01018
摘要： The present invention relates to eukaryotic host cells which have been modified to produce sialylated glycoproteins by the heterologous expression of a set of glycosyltransferases, including sialyltransferase and/or trans-sialidase, to become host-strains for the production of mammalian, e.g., human therapeutic glycoproteins. Novel eukaryotic host cells expressing a CMP-sialic acid biosynthetic pathway for the production of sialylated glycoproteins are also provided. The invention provides nucleic acid molecules and combinatorial libraries which can be used to successfully target and express mammalian enzymatic activities (such as those involved in sialylation) to intracellular compartments in a eukaryotic host cell. The process provides an engineered host cell which can be used to express and target any desirable gene(s) involved in glycosylation.
摘要翻译：本发明涉及通过异源表达一组糖基转移酶（包括唾液酸转移酶和/或反式唾液酸酶）而被修饰以产生唾液酸化糖蛋白的真核宿主细胞，以成为用于产生哺乳动物例如人类治疗的宿主菌株糖蛋白。还提供了表达用于产生唾液酸化糖蛋白的CMP-唾液酸生物合成途径的新型真核宿主细胞。本发明提供核酸分子和组合文库，其可用于成功靶向并表达哺乳动物酶活性（例如涉及唾液酸化的那些）到真核宿主细胞中的细胞内区室。该方法提供了可用于表达和靶向参与糖基化的任何所需基因的工程化宿主细胞。

9. 发明申请

US20100016561A1 N-Acetylglucosaminyltransferase III Expression in Lower Eukaryotes 有权
公开(公告)号：US20100016561A1
公开(公告)日：2010-01-21
申请号：US12540849
申请日：2009-08-13
申请人： Piotr Bobrowicz , Stephen R. Hamilton , Tillman U. Gerngross , Stefan Wildt , Byung-Kwon Choi , Juergen Hermann Nett , Robert C. Davidson
发明人： Piotr Bobrowicz , Stephen R. Hamilton , Tillman U. Gerngross , Stefan Wildt , Byung-Kwon Choi , Juergen Hermann Nett , Robert C. Davidson
IPC分类号： C07K14/00 , C12P21/06
CPC分类号： C07K14/39 , A61K38/00 , C07K2319/05 , C12N9/1051 , C12P21/005 , Y02A50/396
摘要： The present invention relates to eukaryotic host cells having modified oligosaccharides which may be modified further by heterologous expression of a set of glycosyltransferases, sugar transporters and mannosidases to become host-strains for the production of mammalian, e.g., human therapeutic glycoproteins. The process provides an engineered host cell which can be used to express and target any desirable gene(s) involved in glycosylation. Host cells with modified lipid-linked oligosaccharides are created or selected. N-glycans made in the engineered host cells exhibit GnTIII activity, which produce bisected N-glycan structures and may be modified further by heterologous expression of one or more enzymes, e.g., glycosyltransferases, sugar transporters and mannosidases, to yield human-like glycoproteins. For the production of therapeutic proteins, this method may be adapted to engineer cell lines in which any desired glycosylation structure may be obtained.

10. 发明申请

US20090155847A1 Combinatorial DNA library for producing modified N-glycans in lower eukaryotes 有权
公开(公告)号：US20090155847A1
公开(公告)日：2009-06-18
申请号：US12291373
申请日：2008-11-07
申请人： Tillman U. Gerngross , Stefan Wildt , Byung-Kwon Choi , Juergen Hermann Nett , Piotr Bobrowicz , Stephen R. Hamilton , Robert C. Davidson
发明人： Tillman U. Gerngross , Stefan Wildt , Byung-Kwon Choi , Juergen Hermann Nett , Piotr Bobrowicz , Stephen R. Hamilton , Robert C. Davidson
IPC分类号： C12P21/00
CPC分类号： C12P21/005 , C07K14/47 , C12N9/1051 , C12N15/1082 , C12N15/79
摘要： The present invention relates to eukaryotic host cells having modified oligosaccharides which may be modified further by heterologous expression of a set of glycosyltransferases, sugar transporters and mannosidases to become host-strains for the production of mammalian, e.g., human therapeutic glycoproteins. The invention provides nucleic acid molecules and combinatorial libraries which can be used to successfully target and express mammalian enzymatic activities such as those involved in glycosylation to intracellular compartments in a eukaryotic host cell. The process provides an engineered host cell which can be used to express and target any desirable gene(s) involved in glycosylation. Host cells with modified oligosaccharides are created or selected. N-glycans made in the engineered host cells have a Man5GlcNAc2 core structure which may then be modified further by heterologous expression of one or more enzymes, e.g., glycosyltransferases, sugar transporters and mannosidases, to yield human-like glycoproteins. For the production of therapeutic proteins, this method may be adapted to engineer cell lines in which any desired glycosylation structure may be obtained.

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