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21. 发明公开

US20240043831A1 EXPRESSION AND SECRETION SYSTEM 审中-公开
公开(公告)号：US20240043831A1
公开(公告)日：2024-02-08
申请号：US18120020
申请日：2023-03-10
申请人： Genentech, Inc.
发明人： Devin TESAR , Xiaocheng CHEN , Mark DENNIS , Isidro HOTZEL
IPC分类号： C12N15/10 , C12N15/70 , C12N15/85 , C12P21/02 , C07K16/00 , C12N15/74
CPC分类号： C12N15/1037 , C12N15/70 , C12N15/85 , C12P21/02 , C07K16/00 , C12N15/74 , C07K2319/02 , C07K2319/40 , C07K2319/735 , C12N2820/10 , C12N2820/55 , C12N2820/85 , C12N2830/42 , C12N2830/55 , C12N2830/85
摘要： The invention provides an expression and secretion system, and methods of using the same, for the expression and secretion of one fusion protein in prokaryotic cells and a second fusion protein in eukaryotic cells. Also provided herein are nucleic acid molecules, vectors and host cells comprising such vectors and nucleic acid molecules.

22. 发明授权

US08137963B2 Method for highly amplifying target gene in mammalian cell and vector therefor 失效
标题翻译：用于高度扩增哺乳动物细胞中靶基因的方法及其载体
公开(公告)号：US08137963B2
公开(公告)日：2012-03-20
申请号：US12438545
申请日：2007-08-20
申请人： Noriaki Shimizu , Toshihiko Hashizume , Masashi Shimizu
发明人： Noriaki Shimizu , Toshihiko Hashizume , Masashi Shimizu
IPC分类号： C12N15/63 , C12N15/85 , C12P19/34
CPC分类号： C12N15/67 , C12N2510/02 , C12N2800/108 , C12N2820/85 , C12N2830/46
摘要： A vector of the present invention is a vector for amplifying a target gene in a mammalian cell, the vector including an amplification-activating fragment, which is a partial fragment of a mammalian replication initiation region and has a gene amplification activity site, and a mammalian nuclear matrix attachment region. In the case where the mammalian replication initiation region as described above derives from a c-myc locus, for example, the above-described partial fragment at least contains a duplex unwinding element and a topoisomerase II-binding domain. The vector as described above improves gene transfer efficiency and gene amplification efficiency compared with the existing high gene amplification systems. Thus, a method whereby a “high gene amplification system” developed by the inventors can amplify a target gene with better gene transfer efficiency and a vector to be used in this method are provided.
摘要翻译：本发明的载体是用于扩增哺乳动物细胞中的靶基因的载体，所述载体包含作为哺乳动物复制起始区域的部分片段的扩增活化片段，并具有基因扩增活性位点和哺乳动物核基质附着区。在如上所述的哺乳动物复制起始区源自c-myc基因座的情况下，例如，上述部分片段至少含有双链体展开元件和拓扑异构酶II结合结构域。如上所述的载体与现有的高基因扩增系统相比提高了基因转移效率和基因扩增效率。因此，提供了本发明人开发的“高基因扩增系统”可以扩增具有更好的基因转移效率的靶基因和用于该方法的载体的方法。

23. 发明申请

US20080235817A1 Artificial mammalian chromosome 审中-公开
标题翻译：人工哺乳动物染色体
公开(公告)号：US20080235817A1
公开(公告)日：2008-09-25
申请号：US11987206
申请日：2007-11-28
申请人： Tsuneko Okazaki , Masashi Ikeno , Toshihide Itou , Nobutaka Suzuki
发明人： Tsuneko Okazaki , Masashi Ikeno , Toshihide Itou , Nobutaka Suzuki
IPC分类号： A01K67/027 , C07K14/00 , C12N5/06
CPC分类号： C12N15/85 , A01K2217/05 , A61K48/00 , C12N2800/204 , C12N2800/206 , C12N2800/208 , C12N2820/00 , C12N2820/85
摘要： It is intended to provide an artificial mammalian chromosome which is stably held in mammalian cells and allows efficient expression of a target gene carried thereby. Namely, a first cyclic vector containing a mammalian centromere sequence and a selection marker gene and a second cyclic vector containing a functional sequence are transferred into mammalian host cells. Then transformed cells are selected by using the above-described selection marker gene and cells holding an artificial mammalian chromosome are selected from among the transformed cells thus selected. Thus, it is possible to construct an artificial mammalian chromosome which has a mammalian replication origin, the mammalian centromere sequence and the functional sequence, is in a cyclic form, can be replicated in mammalian cells, extrachromosomally held in the host cells and transferred to daughter cells in cell division.
摘要翻译：旨在提供稳定保持在哺乳动物细胞中的人造哺乳动物染色体，并且能够有效地表达由其携带的靶基因。即，将含有哺乳动物着丝粒序列和选择标记基因的第一个环状载体和含有功能序列的第二个环状载体转移到哺乳动物宿主细胞中。然后通过使用上述选择标记基因选择转化的细胞，并且从这样选择的转化细胞中选择携带人造哺乳动物染色体的细胞。因此，可以构建具有哺乳动物复制起点，哺乳动物着丝粒序列和功能序列为环状的人造哺乳动物染色体，可以在哺乳动物细胞中复制，染色体外保留在宿主细胞中并转移到女儿细胞分裂细胞。

24. 发明申请

US20080131871A1 Compositions and methods for identifying antigens which elicit an immune response 失效
标题翻译：用于鉴定引发免疫应答的抗原的组合物和方法
公开(公告)号：US20080131871A1
公开(公告)日：2008-06-05
申请号：US11896337
申请日：2007-08-31
申请人： Si-Yi Chen , Zhaoyang You
发明人： Si-Yi Chen , Zhaoyang You
IPC分类号： C12Q1/70 , C12Q1/68
CPC分类号： C12N15/1086 , A61K38/00 , A61K39/00 , A61K39/0011 , A61K39/12 , A61K39/292 , A61K47/646 , A61K48/00 , A61K2039/515 , A61K2039/5154 , A61K2039/5156 , A61K2039/53 , A61K2039/55533 , A61K2039/57 , A61K2039/6056 , C07K14/005 , C07K2319/02 , C07K2319/30 , C07K2319/74 , C12N15/86 , C12N2730/10122 , C12N2730/10134 , C12N2740/10043 , C12N2740/10051 , C12N2740/13043 , C12N2740/13051 , C12N2740/15043 , C12N2740/15051 , C12N2820/85 , C12N2840/20 , C12N2840/203 , C12Q1/6883 , C12Q1/6886 , C12Q1/70
摘要： This invention relates to an expression vector wherein said expression vector comprises a polynucleotide promoter sequence, a polynucleotide encoding a signal sequence, a polynucleotide encoding an antigen protein or peptide, a polynucleotide encoding a cell binding element, and a polynucleotide polyadenylation sequence all operatively linked. More particularly, it relates to the method of eliciting an immune response directed against an antigen in a mammal comprising the steps of introducing the expression vector into a cell, expressing the vector to produce an antigen under conditions wherein the antigen is secreted from the cell, endocytosing the secreted antigen into the cell, processing the antigen, and presenting fragments to a receptor to elicit a T-cell response. In addition, this invention relates to a vaccine and a method of use. The invention also relates to the method of identifying MHC-II restricted epitopes.
摘要翻译：本发明涉及表达载体，其中所述表达载体包含多核苷酸启动子序列，编码信号序列的多核苷酸，编码抗原蛋白或肽的多核苷酸，编码细胞结合元件的多核苷酸和全部可操作地连接的多核苷酸聚腺苷酸化序列。更具体地说，本发明涉及在哺乳动物中引发针对抗原的免疫应答的方法，包括以下步骤：将表达载体导入细胞，在其中抗原从细胞分泌的条件下表达载体产生抗原，将分泌的抗原内吞到细胞中，处理抗原，并将片段呈递给受体以引发T细胞应答。此外，本发明涉及疫苗和使用方法。本发明还涉及鉴定MHC-II限制性表位的方法。

25. 发明申请

US20080032341A1 Method and kit for expressing protein under regulation of the expression from repeated sequence formed by gene amplification, and transformant 有权
标题翻译：用于通过基因扩增形成的重复序列表达调控表达蛋白质的方法和试剂盒，以及转化体
公开(公告)号：US20080032341A1
公开(公告)日：2008-02-07
申请号：US11889512
申请日：2007-08-14
申请人： Noriaki Shimizu
发明人： Noriaki Shimizu
IPC分类号： C12P21/00 , C12N5/10
CPC分类号： C12N15/67 , C12N15/85 , C12N2810/85 , C12N2820/85 , C12P21/02
摘要： A method is disclosed for releasing the transcriptional regulation caused by a repeated sequence in a gene, a kit therefor and so on to thereby establish a system capable of producing a protein in a large amount. At least one embodiment of the method can be achieved by any one or more of the following methods: (a) in the amplification of a gene encoding a target protein, co-amplifying a polynucleotide of 10 kbp or more such as a α-phage DNA or an insulator sequence; (b) selecting by culturing cells having undergone gene amplification in media containing a drug with a gradual increase in concentration; (c) elevating the promoter activity of inducing the expression of a gene encoding a target protein; (d) excising an amplified gene region from a chromosome with the use of Cre-LoxP System; (e) treating cells having undergone gene amplification with 5-aza-2′-deoxycytidine to thereby lower the methylation degree of DNA; and (f) selecting the mammalian cells having undergone gene amplification on double minute chromosomes.
摘要翻译：公开了一种用于释放由基因中的重复序列引起的转录调控的方法，其用于其的试剂盒等，从而建立能够大量产生蛋白质的系统。该方法的至少一个实施方案可以通过以下任何一种或多种方法来实现：（a）扩增编码靶蛋白的基因，共扩增10kbp或更多的多核苷酸，例如α-噬菌体 DNA或绝缘子序列; （b）通过培养在具有逐渐增加浓度的药物的培养基中培养经历基因扩增的细胞进行选择; （c）提高诱导编码靶蛋白的基因表达的启动子活性; （d）使用Cre-LoxP系统从染色体切除扩增的基因区域; （e）处理已经用5-氮杂-2'-脱氧胞苷进行基因扩增的细胞，从而降低DNA的甲基化程度; 和（f）选择在双分子染色体上经历基因扩增的哺乳动物细胞。

26. 发明授权

US07273752B2 Compositions and methods for identifying antigens which elicit an immune response 有权
标题翻译：用于鉴定引发免疫应答的抗原的组合物和方法
公开(公告)号：US07273752B2
公开(公告)日：2007-09-25
申请号：US10681410
申请日：2003-10-08
申请人： Si-Yi Chen , Zhaoyang You
发明人： Si-Yi Chen , Zhaoyang You
IPC分类号： C12N15/00
CPC分类号： C12N15/1086 , A61K38/00 , A61K39/00 , A61K39/0011 , A61K39/12 , A61K39/292 , A61K47/646 , A61K48/00 , A61K2039/515 , A61K2039/5154 , A61K2039/5156 , A61K2039/53 , A61K2039/55533 , A61K2039/57 , A61K2039/6056 , C07K14/005 , C07K2319/02 , C07K2319/30 , C07K2319/74 , C12N15/86 , C12N2730/10122 , C12N2730/10134 , C12N2740/10043 , C12N2740/10051 , C12N2740/13043 , C12N2740/13051 , C12N2740/15043 , C12N2740/15051 , C12N2820/85 , C12N2840/20 , C12N2840/203 , C12Q1/6883 , C12Q1/6886 , C12Q1/70
摘要： This invention relates to an expression vector wherein said expression vector comprises a polynucleotide promoter sequence, a polynucleotide encoding a signal sequence, a polynucleotide encoding an antigen protein or peptide, a polynucleotide encoding a cell binding element, and a polynucleotide polyadenylation sequence all operatively linked. More particularly, it relates to the method of eliciting an immune response directed against an antigen in a mammal comprising the steps of introducing the expression vector into a cell, expressing the vector to produce an antigen under conditions wherein the antigen is secreted from the cell, endocytosing the secreted antigen into the cell, processing the antigen, and presenting fragments to a receptor to elicit a T-cell response. In addition, this invention relates to a vaccine and a method of use. The invention also relates to the method of identifying MHC-II restricted epitopes.
摘要翻译：本发明涉及表达载体，其中所述表达载体包含多核苷酸启动子序列，编码信号序列的多核苷酸，编码抗原蛋白或肽的多核苷酸，编码细胞结合元件的多核苷酸和全部可操作地连接的多核苷酸聚腺苷酸化序列。更具体地说，本发明涉及在哺乳动物中引发针对抗原的免疫应答的方法，包括以下步骤：将表达载体导入细胞，在其中抗原从细胞分泌的条件下表达载体产生抗原，将分泌的抗原内吞到细胞中，处理抗原，并将片段呈递给受体以引发T细胞应答。此外，本发明涉及疫苗和使用方法。本发明还涉及鉴定MHC-II限制性表位的方法。

27. 发明申请

US20070004002A1 Artificial mammalian chromosome 审中-公开
标题翻译：人工哺乳动物染色体
公开(公告)号：US20070004002A1
公开(公告)日：2007-01-04
申请号：US10526425
申请日：2003-09-01
申请人： Tsuneko Okazaki
发明人： Tsuneko Okazaki
IPC分类号： C12P21/06 , C12N15/09 , C07K14/80 , C07H21/04
CPC分类号： C12N15/85 , A01K2217/05 , A61K48/00 , C12N2800/204 , C12N2800/206 , C12N2800/208 , C12N2820/00 , C12N2820/85
摘要： It is intended to provide an artificial mammalian chromosome which is stably held in mammalian cells and allows efficient expression of a target gene carried thereby. Namely, a first cyclic vector containing a mammalian centromere sequence and a selection marker gene and a second cyclic vector containing a functional sequence are transferred into mammalian host cells. Then transformed cells are selected by using the above-described selection marker gene and cells holding an artificial mammalian chromosome are selected from among the transformed cells thus selected. Thus, it is possible to construct an artificial mammalian chromosome which has a mammalian replication origin, the mammalian centromere sequence and the functional sequence, is in a cyclic form, can be replicated in mammalian cells, extrachromosomally held in the host cells and transferred to daughter cells in cell division.
摘要翻译：旨在提供稳定保持在哺乳动物细胞中的人造哺乳动物染色体，并且能够有效地表达由其携带的靶基因。即，将含有哺乳动物着丝粒序列和选择标记基因的第一个环状载体和含有功能序列的第二个环状载体转移到哺乳动物宿主细胞中。然后通过使用上述选择标记基因选择转化的细胞，并且从这样选择的转化细胞中选择携带人造哺乳动物染色体的细胞。因此，可以构建具有哺乳动物复制起点，哺乳动物着丝粒序列和功能序列为环状的人造哺乳动物染色体，可以在哺乳动物细胞中复制，染色体外保留在宿主细胞中并转移到女儿细胞分裂细胞。

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