Adenoviral amplicon and producer cells for the production of...

C - Chemistry – Metallurgy – 12 – N

Patent

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C12N 15/861 (2006.01)

Patent

CA 2585523

The present invention relates to a plasmid that can be used for the development of efficient producer cell lines for the production of helper independent adenovirus vectors carrying multiple deletions of non-structural as well as structural genes. More specifically, the present invention provides producer cells which comprise a novel adenoviral amplicon that can be used to complement a multi-deleted adenoviral vectors and obtain high titer preparations. The amplicon is an episomal plasmid that expresses Ad5 E2 viral genes (i.e.,polymerase, pre-terminal protein and DNA binding protein) and E4 orf6 , the EBV the latent origin of replication (OriP) as well as adenoviral origins of replications in form of a covalent junction of left and right ITRs. This plasmid is capable of self-replication upon induction of Ad5 E2 gene expression. The invention further includes methods for the preparation of the disclosed producer cells and uses of the cells to produce viral vectors on a scale that is sufficient for therapeutic uses.

L'invention porte sur un plasmide pouvant favoriser le développement de lignées de cellules productrices à partir de vecteurs d'adénovirus indépendants des cellules T auxiliaires porteurs de multiples délétions de gènes structuraux ou non structuraux. L'invention porte plus spécifiquement sur des cellules productrices comprenant un nouvel amplicon adénoviral pouvant compléter des vecteurs adénoviraux à plusieurs délétions et permettant d'obtenir des préparations à titre plus élevé. L'amplicon est un plasmide épisomal exprimant: les gènes viraux Ad5 E2 (par exemple la polymérase, les protéines préterminales, et les protéines se fixant sur l'ADN), et E4orf6) et l'EBV origine latente de la réplication, et origine adénovirale de réplications sous forme de liaison covalente des ITRs gauche et droit, ledit plasmide étant capable d'auto-réplication lors de l'induction de l'expression du gène Ad5 E2. L'invention porte également sur les procédés de préparation desdites cellules productrices, et sur leur utilisation pour produire des vecteurs viraux à une échelle suffisante pour des utilisations thérapeutiques.

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