Method using a three-dimensional bioprocessor

Patent metadata

Field	Value
Publication number	US-11492580-B2
Application number	US-202016872823-A
Country	US
Kind code	B2
Filing date	May 12, 2020
Priority date	May 12, 2020
Publication date	Nov 8, 2022
Grant date	Nov 8, 2022

Abstract

Described herein is a beads-free bioprocessor as an automated and cost-effective T cell processing and manufacturing platform. T cells are a core component in CAR T cell therapies for cancer treatment, but are difficult to manufacture to scale in clinically relevant quantities. The 3D bioprocessor provides an alternative device that is scalable, beads-free, easy-to-use, and cost-effective for using CAR T cell therapy in cancer immunotherapy. Besides CAR T cell application, this platform technology has potential for many other applications such as cancer cell isolation.

First claim

The invention claimed is:

1. A method, comprising;
supplying a 3D bioprocessor comprising a plurality of spheres and a plurality of rods interconnecting the plurality of spheres, the plurality of rods assisting in providing spacing between the plurality of spheres that are adjacent to each other;
providing a coating on a surface area of the 3D bioprocessor, the surface area including the plurality of spheres and the plurality of rods;
providing a protein attached to the coating;
providing one or more biotinylated antibodies immobilized on the protein;
flowing cells through the 3D bioprocessor, wherein the cells bind to the one or more biotinylated antibodies.

2. The method of claim 1 , wherein the cells comprise T-cells, Wherein receptors of the T-cells bind to the one or more biotinylated antibodies and are activated and further comprising exposing the activated T-cells to a perfusion media containing a signaling molecule to promote T-cell expansion.

3. The method of claim 1 , wherein the coating is a protein coating.

4. The method of claim 1 , wherein the protein comprises a tetrameric protein.

5. The method of claim 4 , wherein the tetrameric protein comprises avidin or streptavidin.

6. The method of claim 1 , wherein the one or more biotinylated antibody is selected from the group consisting of: anti-CD3 antibody, anti-CD28 antibody, and anti-CD2 antibody.

7. The method of claim 2 , wherein the signaling molecule comprises a cytokine signaling molecule.

8. The method of claim 1 , wherein the 3D processor is formed from a material that has a Tensile Modulus of at least 0.01 GPa.

9. The method of claim 1 , wherein the 3D bioprocessor is formed from a material that is biocompatible.

10. The method of claim 1 , wherein the 3D bioprocessor is formed from a material not susceptible to hydrolysis during cell cultivation such that the amount of hydrolysis does not exceed 5.0% by weight of the material present.

11. The method of claim 1 , wherein the diameter of the plurality of spheres is from about 10 μm to about 10 mm.

12. The method of claim 11 wherein the diameter of the plurality of spheres is from about 1 mm to about 3 mm.

13. The method of claim 1 , wherein the length of the plurality of rods is from about 0.1 μm to about 10 mm.

14. The method of claim 13 , wherein the length of the plurality of rods is from about 100 μm to about 3 mm.

15. The method of claim 1 , wherein the plurality of spheres and the plurality of rods are organized in one or more layers.

16. The method of claim 15 , wherein the one or more layers comprises at least two layers, with each layer offset from an adjacent layer.

17. The method of claim 1 , wherein the plurality of spheres and the plurality of rods form a repeatable and non-random mesh structure.

18. The method of claim 1 , wherein the plurality of spheres and the plurality of rods form a fixed structure of substantial uniformity.

19. The method of claim 1 , wherein the 3D bioprocessor comprises a polymeric material selected from polystyrene, polycarbonate, acrylonitrile-butadiene-styrene (ABS), polylactic acid (PLA), and polycaprolactone (PCL).

20. The method of claim 1 , wherein the 3D bioprocessor comprises a photo-polymerization biocompatible material.

21. The method of claim 20 , wherein the photo-polymerization biocompatible material is poly(methyl methacrylate).

22. The method of claim 1 , wherein the 3D bioprocessor comprises a material having a tensile modulus of from about 0.01 to about 10.0 GPa.

23. The method of claim 1 , wherein each of the diameters of the plurality of rods is less than half of each of the diameters of the plurality of spheres.

24. A method, comprising;
supplying a 3D bioprocessor comprising a plurality of spherical beads and a plurality of rods interconnecting the plurality of spherical beads, the plurality of rods assisting in providing spacing between the plurality of spherical beads that are adjacent to each other;
providing a coating on a surface area of the 3D bioprocessor, the surface area including the plurality of spherical beads and the plurality of rods;
providing a protein attached to the coating;
providing one or more biotinylated antibodies immobilized on the protein;
flowing cells through the 3D bioprocessor, wherein the cells bind to the one or more biotinylated antibodies.

25. The method of claim 24 , wherein the plurality of rods is cylindrical.

26. The method of claim 24 , wherein the coating is a protein coating.

27. The method of claim 24 , wherein the diameter of the plurality of spherical beads is from about t mm to about 3 mm.

28. The method of claim 24 , wherein the plurality of spherical beads and the plurality of rods form a repeatable and non-random mesh structure.

29. The method of claim 24 , wherein the plurality of spherical beads and the plurality of rods form a fixed structure of substantial uniformity.

30. The method of claim 24 , wherein each of the diameters of the plurality of rods is less than half of each of the diameters of the plurality of spherical beads.

Assignees

• Southwest Res Inst
• Univ Pennsylvania

Inventors

• Ling Jian
• Zimmern Kreg A
• Milone Michael C

Classifications

• C12M29/10

  Perfusion · CPC title

• C12M25/14

  Scaffolds; Matrices (in general C12N5/0068) · CPC title

• C12M23/02 Primary

  Form or structure of the vessel (large containers B65D88/00) · CPC title

• C12N5/10

  Cells modified by introduction of foreign genetic material · CPC title

• C12N2513/00

  3D culture · CPC title