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公开(公告)号：US08021653B2
公开(公告)日：2011-09-20
申请号：US11718680
申请日：2005-11-24
申请人： Yasunobu Kano , Yoshinori Hamaji , Minoru Fujimori , Takayuki Sasaki , Kyoko Kohno , Jun Amano , Shun'ichiro Taniguchi
发明人： Yasunobu Kano , Yoshinori Hamaji , Minoru Fujimori , Takayuki Sasaki , Kyoko Kohno , Jun Amano , Shun'ichiro Taniguchi
CPC分类号： C12N15/70 , A61K38/00 , C12N15/746
摘要： The present invention provides a shuttle vector for a microorganism of the genus Bifidobacterium (BM) and Escherichia coli having a wide host range and a large copy number in BM and capable of highly expressing a desired protein when used as an expression vector; an expression vector capable of expressing a desired gene in BM by use of the shuttle vector; BM transformed with the expression vector; and an antitumor agent comprising the BM as an active ingredient. It comprises a pTB6-derived region portion comprising a replication origin (oriV)-repB region of pTB6 but not comprising MembB, MobA, OrfI, and oriT regions of pTB6 and an Escherichia coli-derived plasmid portion comprising a replication origin (Puc Ori) region of Escherichia coli but having deleted DNA encoding an N-terminal region of an ampicillin resistance gene (ampR) expression product β-lactamase.
摘要翻译： 本发明提供了一种用于双歧杆菌（BM）和微生物的穿梭载体，其具有宽的宿主范围，并且在BM中具有大的拷贝数，并且当用作表达载体时能够高度表达所需的蛋白质。 能够通过穿梭载体在BM中表达所需基因的表达载体; BM用表达载体转化; 和包含BM作为活性成分的抗肿瘤剂。 其包含pTB6衍生的区域部分，其包含pTB6的复制起点（oriV）-repB区，但不包含pBB6的MembB，MobA，OrfI和oriT区以及包含复制起点（Puc Ori）的大肠杆菌来源的质粒部分， 大肠杆菌的区域，但是具有编码氨苄青霉素抗性基因（ampR）表达产物＆amp;内酰胺酶的N末端区域的缺失的DNA。
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公开(公告)号：US20100129323A1
公开(公告)日：2010-05-27
申请号：US11718680
申请日：2005-11-24
申请人： Yasunobu Kano , Yoshinori Hamaji , Minoru Fujimori , Takayuki Sasaki , Kyoko Kohno , Jun Amano , Shun'ichiro Taniguchi
发明人： Yasunobu Kano , Yoshinori Hamaji , Minoru Fujimori , Takayuki Sasaki , Kyoko Kohno , Jun Amano , Shun'ichiro Taniguchi
CPC分类号： C12N15/70 , A61K38/00 , C12N15/746
摘要： The present invention provides a shuttle vector for a microorganism of the genus Bifidobacterium (BM) and Escherichia coli having a wide host range and a large copy number in BM and capable of highly expressing a desired protein when used as an expression vector; an expression vector capable of expressing a desired gene in BM by use of the shuttle vector; BM transformed with the expression vector; and an antitumor agent comprising the BM as an active ingredient. It comprises a pTB6-derived region portion comprising a replication origin (oriV)-repB region of pTB6 but not comprising MembB, MobA, OrfI, and oriT regions of pTB6 and an Escherichia coli-derived plasmid portion comprising a replication origin (oriC) region of Escherichia coli but having deleted DNA encoding an N-terminal region of an ampicillin resistance gene (ampR) expression product β-lactamase, and constitute a shuttle vector for Escherichia coli with a BM having an average copy number of 6 to 30.
摘要翻译： 本发明提供了一种用于双歧杆菌（BM）和微生物的穿梭载体，其具有宽的宿主范围，并且在BM中具有大的拷贝数，并且当用作表达载体时能够高度表达所需的蛋白质。 能够通过穿梭载体在BM中表达所需基因的表达载体; BM用表达载体转化; 以及含有BM作为活性成分的抗肿瘤剂。 其包含pTB6衍生的区域部分，其包含pTB6的复制起点（oriV）-repB区，但不包含pBB6的MembB，MobA，OrfI和oriT区以及包含复制起点（oriC）区的大肠杆菌来源的质粒部分 的大肠杆菌，但是具有编码氨苄青霉素抗性基因（ampR）表达产物和β-内酰胺酶的N末端区域的缺失的DNA，并且构建具有平均拷贝数为6至30的BM的大肠杆菌的穿梭载体。
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公开(公告)号：US08911721B2
公开(公告)日：2014-12-16
申请号：US12301670
申请日：2007-05-24
申请人： Yuko Shimatani , Yoshinori Hamaji , Hitomi Matsuhashi , Jun Amano , Shun'ichiro Taniguchi , Minoru Fujimori
发明人： Yuko Shimatani , Yoshinori Hamaji , Hitomi Matsuhashi , Jun Amano , Shun'ichiro Taniguchi , Minoru Fujimori
CPC分类号： A61K48/0008 , A61K31/513 , A61K35/74 , A61K35/745 , A61K48/00 , C12N9/78 , Y02A50/473
摘要： An object of the present invention is to provide a method of efficiently constructing a gene delivery carrier having a favorable activity and expression efficiency of a protein expressed by a gene introduced by transformation. Moreover, an object of the present invention is to provide a pharmaceutical composition comprising a gene delivery carrier constructed by the construction method and a therapeutic agent for solid tumor comprising the resistant bacterium. The objects of the present invention have been attained by a gene delivery carrier consisting of an anaerobic microorganism capable of growing in a tumor tissue in an anaerobic environment and capable of expressing (A) a protein having an antitumor activity or (B) a protein having an activity of converting an antitumor substance precursor to an antitumor substance, wherein DNA encoding the protein of interest is allowed to further comprise, on the 5′-terminal side thereof, a DNA fragment comprising a nucleotide sequence encoding a peptide consisting of an amino acid sequence from the 1st to at least 4th amino acids at the N-terminus of a histone-like DNA-binding protein.
摘要翻译： 本发明的目的是提供一种有效地构建由通过转化引入的基因表达的蛋白质具有良好活性和表达效率的基因递送载体的方法。 此外，本发明的目的是提供一种药物组合物，其包含通过构建方法构建的基因递送载体和包含耐药性细菌的固体肿瘤治疗剂。 本发明的目的是通过一种基因递送载体实现的，所述基因递送载体由厌氧微生物组成，所述厌氧微生物能够在厌氧环境中在肿瘤组织中生长并且能够表达（A）具有抗肿瘤活性的蛋白质，或（B）具有 将抗肿瘤物质前体转化为抗肿瘤物质的活性，其中编码目的蛋白质的DNA在其5'-末端侧进一步包含编码由氨基酸组成的肽的核苷酸序列的DNA片段 序列从组蛋白样DNA结合蛋白的N末端的第1个至第4个氨基酸。
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公开(公告)号：US08734779B2
公开(公告)日：2014-05-27
申请号：US11910880
申请日：2006-04-04
IPC分类号： A61K48/00 , C12Q1/68 , C12P21/06 , C12N1/20 , C12N15/00 , C12N15/74 , C12N9/78 , C12N9/88 , C12N9/14 , C07H21/04
摘要： The present invention provides a method for producing a cytosine deaminase (CD)-expressing, 5-fluorouracil (5-FU)-resistant microorganism which can grow in anaerobic tumor tissues, can express CD, and has a resistance to 5-FU at a concentration that is at least effective for antitumor activity. More specifically, the method is a method (A) comprising performing subculture or acclimation culture of a CD-expressing microorganism which can grow in anaerobic tumor tissues, in the presence of 5-fluorocytosine (5-FC), or a method (B) comprising (1) performing subculture or acclimation culture of a microorganism which can grow in anaerobic tumor tissues and does not express CD, in the presence of 5-FU to produce a 5-FU-resistant microorganism and (2) transforming the 5-FU-resistant microorganism by introducing a CD gene. The present invention also provides the CD-expressing, 5-FU-resistant microorganism and a pharmaceutical composition comprising the microorganism.
摘要翻译： 本发明提供了一种能够在厌氧性肿瘤组织中生长的胞嘧啶脱氨酶（CD）表达的5-氟尿嘧啶（5-FU）抗性微生物的制备方法，其可以表达CD，并且具有对5-FU的抗性 至少对抗肿瘤活性有效的浓度。 更具体地，该方法是包括在5-氟胞嘧啶（5-FC）或方法（B）的存在下进行可在厌氧性肿瘤组织中生长的表达CD的微生物的传代培养或适应培养的方法（A） 包括（1）在5-FU存在下进行在厌氧性肿瘤组织中生长并且不表达CD的微生物的传代培养或适应培养以产生5-FU抗性微生物，和（2）转化5-FU 引入CD基因的抗性微生物。 本发明还提供了表达CD的5-FU抗性微生物和包含微生物的药物组合物。
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公开(公告)号：US20090169516A1
公开(公告)日：2009-07-02
申请号：US12301670
申请日：2007-05-24
申请人： Yuko Shimatani , Yoshinori Hamaji , Hitomi Matsuhashi , Jun Amano , Shun'ichiro Taniguchi , Minoru Fujimori
发明人： Yuko Shimatani , Yoshinori Hamaji , Hitomi Matsuhashi , Jun Amano , Shun'ichiro Taniguchi , Minoru Fujimori
CPC分类号： A61K48/0008 , A61K31/513 , A61K35/74 , A61K35/745 , A61K48/00 , C12N9/78 , Y02A50/473
摘要： An object of the present invention is to provide a method of efficiently constructing a gene delivery carrier having a favorable activity and expression efficiency of a protein expressed by a gene introduced by transformation. Moreover, an object of the present invention is to provide a pharmaceutical composition comprising a gene delivery carrier constructed by the construction method and a therapeutic agent for solid tumor comprising the resistant bacterium. The objects of the present invention have been attained by a gene delivery carrier consisting of an anaerobic microorganism capable of growing in a tumor tissue in an anaerobic environment and capable of expressing (A) a protein having an antitumor activity or (B) a protein having an activity of converting an antitumor substance precursor to an antitumor substance, wherein DNA encoding the protein of interest is allowed to further comprise, on the 5′-terminal side thereof, a DNA fragment comprising a nucleotide sequence encoding a peptide consisting of an amino acid sequence from the 1st to at least 4th amino acids at the N-terminus of a histone-like DNA-binding protein.
摘要翻译： 本发明的目的是提供一种有效地构建由通过转化引入的基因表达的蛋白质具有良好活性和表达效率的基因递送载体的方法。 此外，本发明的目的是提供一种药物组合物，其包含通过构建方法构建的基因递送载体和包含耐药性细菌的固体肿瘤治疗剂。 本发明的目的是通过一种基因递送载体实现的，所述基因递送载体由厌氧微生物组成，所述厌氧微生物能够在厌氧环境中在肿瘤组织中生长并且能够表达（A）具有抗肿瘤活性的蛋白质，或（B）具有 将抗肿瘤物质前体转化为抗肿瘤物质的活性，其中编码目的蛋白质的DNA在其5'-末端侧进一步包含编码由氨基酸组成的肽的核苷酸序列的DNA片段 序列从组蛋白样DNA结合蛋白的N末端的第1个至第4个氨基酸。
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公开(公告)号：US20090280091A1
公开(公告)日：2009-11-12
申请号：US11910880
申请日：2006-04-04
摘要： The present invention provides a method for producing a cytosine deaminase (CD)-expressing, 5-fluorouracil (5-FU)-resistant microorganism which can grow in anaerobic tumor tissues, can express CD, and has a resistance to 5-FU at a concentration that is at least effective for antitumor activity. More specifically, the method is a method (A) comprising performing subculture or acclimation culture of a CD-expressing microorganism which can grow in anaerobic tumor tissues, in the presence of 5-fluorocytosine (5-FC), or a method (B) comprising (1) performing subculture or acclimation culture of a microorganism which can grow in anaerobic tumor tissues and does not express CD, in the presence of 5-FU to produce a 5-FU-resistant microorganism and (2) transforming the 5-FU-resistant microorganism by introducing a CD gene. The present invention also provides the CD-expressing, 5-FU-resistant microorganism and a pharmaceutical composition comprising the microorganism.
摘要翻译： 本发明提供了一种能够在厌氧性肿瘤组织中生长的胞嘧啶脱氨酶（CD）表达的5-氟尿嘧啶（5-FU）抗性微生物的制备方法，其可以表达CD，并且具有对5-FU的抗性 至少对抗肿瘤活性有效的浓度。 更具体地，该方法是包括在5-氟胞嘧啶（5-FC）或方法（B）的存在下进行可在厌氧性肿瘤组织中生长的表达CD的微生物的传代培养或适应培养的方法（A） 包括（1）在5-FU存在下进行在厌氧性肿瘤组织中生长并且不表达CD的微生物的传代培养或适应培养以产生5-FU抗性微生物，和（2）转化5-FU 引入CD基因的抗性微生物。 本发明还提供了表达CD的5-FU抗性微生物和包含微生物的药物组合物。
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