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    Method of preparing a eukaryotic viral vector
    1.
    发明申请
    Method of preparing a eukaryotic viral vector 有权
    制备真核病毒载体的方法

    公开(公告)号：US20030203480A1

    公开(公告)日：2003-10-30

    申请号：US10427759

    申请日：2003-04-30

    Applicant: GenVec, Inc.

    Inventor： Imre Kovesdi ,  Duncan L. McVey

    IPC: C12N015/861 ,  C12N007/00

    CPC classification number: C12N15/86 ,  C12N2710/10043 ,  C12N2795/10043 ,  C12N2810/80

    Abstract: The inventive method of producing a eukaryotic viral vector comprises contacting a eukaryotic cell, which comprises a unique enzyme that nicks or cleaves a DNA molecule, with a recombinant phage vector, or contacting a eukaryotic cell, which does not comprise a unique enzyme that nicks or cleaves a DNA molecule, simultaneously or sequentially, in either order, with (i) a unique enzyme that nicks or cleaves a DNA molecule, and (ii) a recombinant phage vector. The recombinant phage vector comprises the DNA molecule comprising (a) a eukaryotic viral vector genome comprising a coding sequence, (b) a phage packaging site that is not contained within the eukaryotic viral vector genome, and (c) a promoter that is operably linked to the coding sequence. Alternatively, the DNA molecule is not present within the recombinant phage vector. The eukaryotic cell is contacted with the first DNA molecule and a recombinant phage vector. The first DNA molecule comprises a replication deficient eukaryotic viral vector genome comprising at least one adenoviral inverted terminal repeat and a packaging signal. The recombinant phage vector comprises a second DNA molecule and a phage packaging site, wherein the second DNA molecule complements in trans the replication deficient eukaryotic viral vector genome. The DNA molecule(s) enter the eukaryotic cell, and the unique enzyme nicks or cleaves the DNA molecule in the eukaryotic cell in at least one region not contained within the eukaryotic viral vector genome, thereby inducing the production of and ultimately producing a eukaryotic viral vector.

    Abstract translation: 产生真核病毒载体的本发明方法包括将包含切割或切割DNA分子的独特酶与重组噬菌体载体接触的真核细胞或接触真核细胞，所述真核细胞不包含缺陷或 以（i）切割或切割DNA分子的独特的酶，和（ii）重组噬菌体载体以任一顺序同时或顺序地切割DNA分子。 重组噬菌体载体包含DNA分子，其包含（a）真核病毒载体基因组，其包含编码序列，（b）不包含在真核病毒载体基因组内的噬菌体包装位点，和（c）可操作地连接的启动子 到编码序列。 或者，DNA分子不存在于重组噬菌体载体内。 将真核细胞与第一DNA分子和重组噬菌体载体接触。 第一DNA分子包含复制缺陷型真核病毒载体基因组，其包含至少一个腺病毒反向末端重复和包装信号。 重组噬菌体载体包含第二DNA分子和噬菌体包装位点，其中第二DNA分子与复制缺陷型真核病毒载体基因组相反。 DNA分子进入真核细胞，独特的酶在至少一个不包含在真核病毒载体基因组内的区域中切割或切割真核细胞中的DNA分子，由此诱导产生并最终产生真核病毒 向量。

    Plasmids for construction of eukaryotic viral vectors
    2.
    发明申请
    Plasmids for construction of eukaryotic viral vectors 有权
    用于构建真核病毒载体的质粒

    公开(公告)号：US20020004242A1

    公开(公告)日：2002-01-10

    申请号：US09905758

    申请日：2001-07-13

    Applicant: GenVec, Inc.

    Inventor： Duncan L. McVey ,  Douglas E. Brough ,  Imre Kovesdi

    IPC: C12N015/86 ,  C07H021/04

    CPC classification number: C12N15/86 ,  C12N15/65 ,  C12N2710/10343 ,  C12N2800/30 ,  C12N2830/001 ,  C12N2830/15 ,  C12N2830/55

    Abstract: The present invention provides a dual selection cassette (DSC) comprising first and second DNA segments having homology to a eukaryotic viral vector, positive and negative selection genes, each operably linked to their own promoter, and one or more unique restriction enzyme sites (URES) or sitey-directed homologous recombination sites. The present invention also provides a plasmid, pN/P, comprising an independent positive selection marker gene, an origin of replication, and a dual selection cassette. The dual selection cassette and pN/P plasmid can be used to produce eukaryotic gene transfer vectors without requiring temporally-linked double recombination events or the use of specialized bacterial strains that allow the replication of plasmids comprising defective origins of replication. This method usefully increases the ratio of desired to undesired plasmid and vector constructs. Additionally, this invention provides a method for the creation of eukaryotic viral vector libraries.

    Abstract translation: 本发明提供了一种双重选择盒（DSC），其包含与真核病毒载体具有同源性的第一和第二DNA片段，每个与其本身的启动子可操作地连接的阳性和阴性选择基因，以及一个或多个独特的限制酶位点（URES） 或定点同源重组位点。 本发明还提供了包含独立的阳性选择标记基因，复制起点和双选择盒的质粒pN / P。 双选择盒和pN / P质粒可用于产生真核基因转移载体，而不需要时间连接的双重重组事件或使用允许复制包含有缺陷复制起点的质粒的专门细菌菌株。 该方法有效地增加了期望的和不需要的质粒和载体构建体的比例。 此外，本发明提供了一种用于产生真核病毒载体文库的方法。

    Materials and methods for treating ocular-related disorders
    3.
    发明申请
    Materials and methods for treating ocular-related disorders 审中-公开
    用于治疗眼部相关疾病的材料和方法

    公开(公告)号：US20030045498A1

    公开(公告)日：2003-03-06

    申请号：US10211701

    申请日：2002-08-02

    Applicant: GenVec, Inc.

    Inventor： Imre Kovesdi ,  Douglas E. Brough ,  Lisa Wei ,  Duncan L. McVey

    IPC: A61K048/00 ,  C12N015/85

    CPC classification number: C12N15/86 ,  A61K48/00 ,  A61K48/005 ,  A61K48/0075 ,  C07K14/71 ,  C07K14/811 ,  C12N2710/10343 ,  C12N2799/022

    Abstract: The present invention is directed to a method of prophylactically or therapeutically treating an animal for at least one ocular-related disorder, e.g., ocular neovascularization or age-related macular degeneration. The method comprises contacting an ocular cell with an expression vector comprising a nucleic acid sequence encoding an inhibitor of angiogenesis and the same or different nucleic acid sequence encoding a neurotrophic agent. The method also can comprise contacting an ocular cell with different expression vectors, each comprising a nucleic acid sequence encoding an inhibitor of angiogenesis and/or a nucleic acid sequence encoding a neurotrophic agent. In addition, the present invention provides a viral vector comprising a nucleic acid sequence encoding pigment epithelium-derived factor (PEDF) or a therapeutic fragment thereof.

    Abstract translation: 本发明涉及一种预防性或治疗性地治疗动物至少一种与眼部相关的疾病，例如眼睛新生血管形成或年龄相关性黄斑变性的方法。 该方法包括使眼细胞与包含编码血管生成抑制剂的核酸序列和编码神经营养剂的相同或不同核酸序列的表达载体接触。 该方法还可以包括使眼细胞与不同的表达载体接触，每种表达载体包含编码血管生成抑制剂的核酸序列和/或编码神经营养剂的核酸序列。 此外，本发明提供了包含编码色素上皮衍生因子（PEDF）的核酸序列或其治疗片段的病毒载体。

    Methods of preparing and using a viral vector library
    8.
    发明申请
    Methods of preparing and using a viral vector library 有权
    制备和使用病毒载体文库的方法

    公开(公告)号：US20010026794A1

    公开(公告)日：2001-10-04

    申请号：US09780526

    申请日：2001-02-09

    Applicant: GenVec, Inc.

    Inventor： Imre Kovesdi ,  Duncan L. McVey ,  Thomas J. Wickham ,  Joseph T. Bruder ,  Douglas E. Brough

    IPC: A61K048/00 ,  C12Q001/70 ,  C12N007/01

    CPC classification number: C12N15/1093 ,  A61K48/00 ,  C12N15/86 ,  C12N2710/10343 ,  C12N2710/10345 ,  C12N2810/40

    Abstract: The present invention provides a library of viral vectors, wherein each member comprises a first heterologous DNA encoding a first gene product and a second heterologous DNA encoding a second gene product. The first heterologous DNA is common to each member of the library, while the second heterologous DNA varies between members of the library. The present invention additionally provides a method of constructing a library of viral vectors. The method comprises carrying out homologous recombination between a first DNA molecule and a second DNA molecule to form a pool of intermediate viral vector genomes. One or more linear third DNA molecules are ligated into the pool of intermediate viral genomes to produce a library of viral vector genomes. Alternatively, homologous recombination between linear DNA molecules and recipient DNA molecules produces a library of viral vector genomes. The library of viral vector genomes is converted into a library of viral vectors.

    Abstract translation: 本发明提供病毒载体文库，其中每个成员包含编码第一基因产物的第一异源DNA和编码第二基因产物的第二异源DNA。 第一个异源DNA对于文库的每个成员是共同的，而第二异源DNA在文库的成员之间是不同的。 本发明另外提供构建病毒载体文库的方法。 该方法包括在第一DNA分子和第二DNA分子之间进行同源重组以形成中间病毒载体基因组池。 将一个或多个线性第三DNA分子连接到中间病毒基因组池中以产生病毒载体基因组文库。 或者，线性DNA分子和受体DNA分子之间的同源重组产生病毒载体基因组文库。 将病毒载体基因组文库转化为病毒载体文库。

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