Method for production of high titer virus and high...

C - Chemistry – Metallurgy – 12 – N

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C12N 15/86 (2006.01) C07K 14/725 (2006.01) C07K 14/73 (2006.01) C07K 16/10 (2006.01) C12N 5/10 (2006.01) C12N 7/04 (2006.01) C12N 15/867 (2006.01) C12N 5/08 (2006.01)

Patent

CA 2164835

The invention provides a novel retroviral packaging system, in which retroviral packaging constructs, and packaging vector transcripts are produced from high expression plasmids by transfection in human cells. High titers of recombinant retroviruses are produced in infected cells. The methods of the invention include the use of the novel retroviral constructs to transduce primary human cells, including T cells and human hematopoietic stem cells, with foreign genes by cocultivation at high efficiencies. The invention is useful for the rapid production of high titer viral supernatants, and to transduce with high efficiency cells that are refractory to transduction by conventional means.

L'invention concerne un nouveau système d'encapsidation rétrovirale dans lequel des structures d'encapsidation rétrovirales illustrées par la figure 1 et des transcripts de vecteurs d'encapsidation sont produits à partir de plasmides à fort pouvoir d'expression par transfection dans des cellules humaines. Des titres élevés de rétrovirus recombinés sont produits dans les cellules infectées. Les procédés de l'invention consistent à utiliser les nouvelles structures rétrovirales afin de transduire des cellules humaines primaires, y compris des lymphocytes B et des cellules souche hématopoïétiques humaines à l'aide de gènes étrangers mis en culture conjointement à rendements élevés. L'invention est utile dans la production rapide de surnageants viraux à titre élevé et pour transduire à l'aide de cellules à rendement élevé, lesquelles sont réfractaires à la transduction par des moyens classiques.

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