Autologous T Cell Manufacturing Processes
First Claim
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1. A method for stable transduction of primary cells of the hematopoietic system and/or hematopoietic stem cells, comprising contacting, in vitro or ex vivo, the surface of the cells with both a lentiviral vector and at least one molecule which binds the cell surface, and culturing the cells in a ventilated vessel comprising two or more layers under conditions conducive to growth and/or proliferation, wherein the vessel is suitable for culturing at least about 100 million cells.
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Abstract
The invention provides novel processes for manufacturing autologous T cells, transducing T cells and expanding the transduced T cell population.
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Citations
37 Claims
- 1. A method for stable transduction of primary cells of the hematopoietic system and/or hematopoietic stem cells, comprising contacting, in vitro or ex vivo, the surface of the cells with both a lentiviral vector and at least one molecule which binds the cell surface, and culturing the cells in a ventilated vessel comprising two or more layers under conditions conducive to growth and/or proliferation, wherein the vessel is suitable for culturing at least about 100 million cells.
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2. A method for stable transduction of primary cells of the hematopoietic system and/or hematopoietic stem cells, comprising contacting, in vitro or ex vivo, the surface of the cells with both a lentiviral vector and at least one molecule which binds the cell surface, and culturing the cells in a ventilated vessel comprising two or more layers under conditions conducive to growth and/or proliferation, wherein the vessel is suitable for culturing at least about 100 million cells;
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wherein the cells are transduced with the lentiviral lentiviral vector at a multiplicity of infection (MOI) such that the copies of lentiviral lentiviral vector per transduced cell is from about 0.5 to about 10; and wherein contacting the cells with a lentiviral vector is for about 24 hours and is optionally repeated at least once.
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3. A method for stable transduction of primary cells of the hematopoietic system and/or hematopoietic stem cells, comprising contacting, in vitro or ex vivo, the surface of the cells with both a lentiviral vector and at least one molecule which binds the cell surface, and culturing the cells in a ventilated vessel comprising two or more layers under conditions conducive to growth and/or proliferation, wherein the vessel is suitable for culturing at least about 100 million cell;
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wherein at least about 50% of the cells are stably transduced after about seven to ten days, or at about 14 days; and
optionally at least 50% of the cells remain stably transduced after about 14 days;
orwherein at least about 75% of the cells are stably transduced after about seven to ten days, or at about 14 days, and optionally at least 75% of the cells remain stably transduced after about 14 days;
orwherein greater than 80%, 85%, 89%, 90%, 91%, 92%, 93%, 94% or 95% of the cells are stably transduced after about 14 days;
orwherein the cells are transduced with the lentiviral vector at a multiplicity of infection (MOI) of from about 2 to about 50, or from about 10 to about 30, or from at 10, or at about 20, or at about 30, or at about 40, or at about 50, or from about 1 to about 400, or less than 500;
orwherein the cells are transduced with the lentiviral vector at a multiplicity of infection (MOI) such that the copies of lentiviral vector per transduced cell is from about 1 to about 100;
orwherein the cells are transduced with the lentiviral vector at a multiplicity of infection (MOI) such that the copies of lentiviral vector per transduced cell is from about 0.5 to about 10;
orwherein contacting the cells with a lentiviral vector is for about 24 hours and is optionally repeated at least once; and wherein the cell surface molecule does not induce apoptosis and the cell surface binding molecule results in the cell being more receptive to transduction by a viral lentiviral vector.
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- 30. A method for stable transduction of primary cells of the hematopoietic system and/or hematopoietic stem cells, comprising contacting, in vitro or ex vivo, the surface of the cells with both a lentiviral vector and at least one molecule which binds the cell surface, and culturing the cells in a ventilated vessel comprising two or more layers under conditions conducive to growth and/or proliferation, wherein the vessel is suitable for culturing at least about 100 million cells, and wherein the contacting of the primary cells with the cell surface molecule makes the cells more receptive to transduction by the lentiviral vector.
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33. A method for stable transduction of a primary cell of the hematopoietic system and/or a hematopoietic stem cell isolated from an HIV-infected subject, comprising the steps of:
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(a) isolating from the HIV-infected subject primary cells of the hematopoietic system cells or hematopoietic stem cells; (b) optionally, pre-stimulating the primary cells or hematopoietic stem cells with at least one cell surface binding molecule; (c) contacting simultaneously in vitro or ex vivo the hematopoietic system cells or hematopoietic stem cells with a lentiviral vector and at least one cell surface binding molecule; and (d) culturing the cells in a ventilated vessel comprising two or more layers under conditions conducive to growth and/or proliferation, wherein the vessel is suitable for culturing at least about 100 million cells.
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34. A system comprising:
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(a) a ventilated vessel comprising two or more layers; and (b) isolated non-adherent primary cells of the hematopoietic system and/or hematopoietic stem cells. - View Dependent Claims (35, 36, 37)
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Specification