Methods for manufacturing T cells
The disclosure relates to methods of manufacturing T cells for adoptive immunotherapy. The disclosure further provides for methods of genetically transducing T cells, methods of using T cells, and T cell populations thereof. In an aspect, the disclosure provides for methods of thawing frozen peripheral blood mononuclear cells (PBMC), resting the thawed PBMC, activating the T cell in the cultured PBMC with an anti-CD3 antibody and an anti-CD28 antibody immobilized on a solid phase, transducing the activated T cell with a viral vector, expanding the transduced T cell, and obtaining expanded T cells.
1. A method of transducing a T cell, comprising
thawing frozen peripheral blood mononuclear cells (PBMC),
resting the thawed PBMC for about 4 to about 6 hours,
activating the T cell in the rested PBMC with an anti-CD3 antibody and an anti-CD28 antibody,
transducing the activated T cell with a viral vector,
expanding the transduced T cell, and
obtaining the expanded T cell.
2. The method of claim 1 , wherein the activating step comprises immobilizing the T cell in the rested PBMC with the anti-CD3 antibody and the anti-CD28 antibody on a solid phase support.
3. The method of claim 1 , wherein the resting is carried out in the presence of at least one cytokine.
4. The method of claim 3 , wherein the at least one cytokine comprises interleukin 7 (IL-7) and/or interleukin 15 (IL-15).
5. The method of claim 1 , wherein the activation is carried out within a period of time from about 1 hour to about 120 hours.
6. The method of claim 2 , wherein the solid phase support comprises a surface of a bead, a plate, a flask, or a bag.
7. The method of claim 1 , wherein the transducing is carried out within a period of time from about 1 hour to about 72 hours.
8. The method of claim 1 , wherein the viral vector comprises a retroviral vector expressing a T cell receptor (TCR).
9. The method of claim 1 , wherein the viral vector comprises a lentiviral vector expressing a TCR.
10. The method of claim 1 , wherein the transducing is carried out in the presence of at least one cytokine.
11. The method of claim 10 , wherein the at least one cytokine comprises IL-7 and/or IL-15.
12. The method of claim 1 , wherein the expanding is carried out in the presence of at least one cytokine selected from the group consisting of IL-2, IL-7, IL-12, IL-15, and IL-21.
13. The method of claim 12 , wherein the at least one cytokine comprises IL-2.
14. The method of claim 12 , wherein the at least one cytokine comprises IL-7 and IL-15.
15. The method of claim 1 , wherein the obtained T cell comprises a CD3 + CD8 + T cell.
16. The method of claim 1 , wherein the thawing, the resting, the activating, the transducing, the expanding, and the obtaining are performed in a closed system.
17. The method of claim 1 , wherein number of obtained T cells with a resting time of about 4 hours to about 6 hours is about 1.5-fold to about 2.0-fold greater than that of the obtained T cells prepared under identical conditions with a resting time of about 16 hours to 20 hours.
18. The method of claim 4 , wherein the at least one cytokine is IL-7.
19. The method of claim 4 , wherein the at least one cytokine is IL-15.
20. The method of claim 4 , wherein the at least one cytokine is IL-7 and IL-15.