Retroviral transduction using poloxamers
The present invention relates to a method for transducing a target cell, the method comprising the step of contacting a target cell with a retroviral vector and a poloxamer having a molecular weight of 12.8 kDa to about 15 kDa. Further, the invention relates to the use of a poloxamer as defined herein, optionally in combination with a polycationic substance as defined herein, for transducing a target cell with a retroviral vector and a kit comprising a retroviral vector, a poloxamer as defined herein and, optionally, instructions for use.
1. A method of gene therapy comprising:
(i) transducing a target cell, said transducing comprising contacting the target cell in vitro or ex vivo with a retroviral vector and a poloxamer, said poloxamer having an average molecular weight of 12.8 kDa to about 15 kDa, wherein “about” refers to—10% to +20% of said average molecular weight, and said poloxamer having the formula HO(C 2 H 4 O) x (C 3 H 6 O) z (C 2 H 4 O) y H, wherein the average value of z is at least 43, and the average value of x+y is at least 230, and wherein said poloxamer is in a fluid state during the contacting step, wherein after said contacting step said retroviral vector enters said cell, and
(ii) using the transduced cell for a gene therapy application.
2. The method of claim 1 , selected from one of:
(i) wherein said retroviral vector comprises a target sequence encoding a therapeutic protein, and wherein said transduced cell expresses said therapeutic protein, and
(ii) wherein said retroviral vector comprises a target sequence comprising a regulatory sequence for RNA interference, and wherein said transduced cell expresses said regulatory sequence for RNA interference.
3. The method of claim 1 , wherein said poloxamer is provided at a concentration of about 500 to 1000 μg/ml.
4. The method of claim 1 , further comprising the step of spinoculating said retroviral vector with said target cell prior to, concomitant with or after contacting said target cell with said poloxamer.
5. The method of claim 1 , wherein z is in the range of 44 to 50 and x+y is in the range of 235 to 266.
6. The method of claim 1 , wherein said retroviral vector and said poloxamer are added simultaneously or sequentially to said target cell.
7. The method of claim 1 , wherein said target cell is brought into contact with one or more polycationic substances selected from the group of polycationic polymers and polycationic peptides.
8. A method of gene therapy comprising:
(i) transducing a target cell, said transducing comprising contacting the target cell in vitro or ex vivo with a retroviral vector and a poloxamer, said poloxamer having an average molecular weight of 12.8 kDa to about 15 kDa, wherein “about” refers to—10% to +20% of said average molecular weight, and said poloxamer having the formula HO(C 2 H 4 O) x (C 3 H 6 O) z (C 2 H 4 O) y H, wherein the average value of z is at least 43, and the average value of x+y is at least 230, and wherein said poloxamer is in a fluid state during the contacting step, wherein after said contacting step said retroviral vector enters said cell, wherein said target cell is not brought into contact with one or more polycationic substances selected from the group of polycationic polymers of and polycationic peptides, and
(ii) using the transduced cell for a gene therapy application.
9. The method of claim 1 , wherein said target cell contacted with said retroviral vector and said poloxamer exhibits a higher transduction rate without noticeable toxicity compared with target cells contacted with said retroviral vector without said poloxamer under the same conditions.