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    • 5. 发明申请
    • REPLICONS OF PESTIVIRUSES THAT DO NOT EXPRESS C AND OR E1 PROTEIN AND INFECTIOUS VIRAL PARTICLES CONTAINING SAME, THAT CAN BE USED IN VACCINES
    • 不能表达C和/或蛋白质的蛋白质和含有可能在疫苗中使用的感染性病毒颗粒的替代物
    • WO2004016794A1
    • 2004-02-26
    • PCT/EP2003/009031
    • 2003-08-12
    • AKZO NOBEL N.V.BEER, MartinREIMANN, Ilona
    • BEER, MartinREIMANN, Ilona
    • C12N15/86
    • C12N7/00A61K39/12A61K2039/5254A61K2039/543A61K2039/552C07K14/005C12N15/86C12N2770/24322C12N2770/24343C12N2770/24362
    • The present invention provides new Pestiviral RNA genomes (replicons) that are able to replicate, and can be packaged into infectious viral particles in cells that complement the missing protein(s), but do not produce infectious progeny virus. Such replicons can be useful for vaccine purposes. The replicons encode most, preferably all, envelop proteins of the virus, while, on the other hand, it would not be capable of producing infectious progeny virus. The present invention provides a Pestiviral replicon, preferably from the Bovine Viral Diarrhea Virus (BVDV), which expresses all structural proteins except for a functional C or E1 protein. Preferably at least part of the coding sequence of the E1 or C protein has been deleted from said replicon. The present inventors proved for the first time, that both C and E1 structural proteins are essential for the formation of infectious pestiviruses. Furthermore it was shown that deletion of C and E1 does not impact the ability of RNA self-replication. By using cell lines constitutively expressing pestiviral structural proteins, Capsid- or E1-proteins can be efficiently trans -complemented. The resulting virions are able to infect bovine target cells and to transfer the replicons without generating replication-competent virus progeny. In other words, no infectious progeny virus is produced. The complemented virions are indistinguishable from wild-type Pestivirus.in virus neutralization experiments. Recombinations yielding infectious wild-type virus were not detected in any of the complementation experiments. The complemented viruses may be used for the safe and efficacious immunization against BVDV.
    • 本发明提供能够复制并且可以包装在补充缺失的蛋白质但不产生感染性后代病毒的细胞中的感染性病毒颗粒中的新的瘟病毒RNA基因组(复制子)。 这样的复制子可以用于疫苗目的。 复制子编码病毒的大多数,优选全部包膜蛋白,而另一方面,它将不能产生感染性后代病毒。 本发明提供了一种优选来自牛病毒性腹泻病毒(BVDV)的Pestiviral复制子,其表达除功能性C或E1蛋白以外的所有结构蛋白。 优选地,E1或C蛋白的编码序列的至少一部分已经从所述复制子中删除。 本发明人首次证明了C和E1结构蛋白对于形成感染性瘟病毒是必不可少的。 此外,显示C和E1的缺失不影响RNA自我复制的能力。 通过使用组成型表达瘟病毒结构蛋白的细胞系,衣壳蛋白或E1蛋白可以有效地反式互补。 所得到的病毒粒子能够感染牛靶细胞并转移复制子,而不产生具有复制能力的病毒后代。 换句话说,没有产生感染性后代病毒。 补充的病毒粒子与野生型瘟病毒在病毒中和实验中无法区分。 在任何互补实验中未检测到产生感染性野生型病毒的重组。 补充的病毒可用于对BVDV的安全有效的免疫。