METHOD FOR POLYCLONAL STIMULATION OF T CELLS BY FLEXIBLE NANOMATRICES

• US 20140087462A1
• Filed: 09/24/2013
• Published: 03/27/2014
• Est. Priority Date: 09/25/2012
• Status: Active Grant

• Current Assignee: Miltenyi Biotec B.V. & Co. KG (Miltenyi GmbH)
• Original Assignee: Miltenyi Biotec GmbH
• Inventors: SCHEFFOLD, Alexander, ASSENMACHER, Mario
• Granted Patent: US 10,513,687 B2
• Time in Patent Office: Days
• US Class Current: 435/375
• CPC Class Codes: A61K 39/4611 T-cells, e.g. tumor infiltrating lymphocytes [TIL], lymphokine-activated killer cells [LAK] or regulatory T cells [Treg]
• CPC Class Codes: A61K 39/4632 T-cell receptors [TCR]; antibody T-cell receptor constructs
• CPC Class Codes: A61K 39/464491 Melan-A/MART
• CPC Class Codes: A61P 37/04 Immunostimulants
• CPC Class Codes: C12N 2501/51 B7 molecules, e.g. CD80, CD86, CD28 (ligand), CD152 (ligand)
• CPC Class Codes: C12N 2501/515 CD3, T-cell receptor complex
• CPC Class Codes: C12N 2501/998 Proteins not provided for elsewhere
• CPC Class Codes: C12N 2533/70 Polysaccharides
• CPC Class Codes: C12N 2539/00 Supports and/or coatings for cell culture characterised by properties
• CPC Class Codes: C12N 5/0075 using microcarriers
• CPC Class Codes: C12N 5/0636 T lymphocytes
• CPC Class Codes: C12N 5/0637 Immunosuppressive T lymphocytes, e.g. regulatory T cells or Treg

Abstract

The present invention provides a method polyclonal stimulation of T cells, the method comprising contacting a population of T cells with a nanomatrix, the nanomatrix comprising a) a flexible matrix, wherein said matrix is of polymeric material; and b) attached to said polymeric flexible matrix one or more polyclonal stimulatory agents which provide activation signals to the T cells; thereby activating and inducing the T cells to proliferate; wherein the nanomatrix is 1 to 500 nm in size. At least one first and one second stimulatory agents are attached to the same or to separate flexible matrices. If the stimulatory agents are attached to separate beads, fine-tuning of nanomatrices for the stimulation of the T cells is possible.

Claims

1. An in vitro method for polyclonal stimulation of T cells, the method comprising contacting a population of T cells with a nanomatrix, the nanomatrix comprising

a) a flexible matrix, wherein said matrix is of polymeric material;

and

b) attached to said polymeric flexible matrix one or more polyclonal stimulatory agents which provide activation signals to the T cells;

thereby activating and inducing the T cells to proliferate;

wherein the nanomatrix is 1 to 500 nm in size

2. The method according to claim 1, wherein the nanomatrix is biologically inert with regard to alteration of the cell function.

3. The method according to claim 1, wherein at least one first and one second stimulatory agents are attached to the same flexible matrix.

4. The method according to claim 1, wherein at least one first and one second stimulatory agents are attached to separate flexible matrices.

5. The method according to claim 4, wherein the ratio of nanomatrices to cells is larger than 500:

1 allowing fine-tuning of T cell stimulation.

6. The method according to claim 1, wherein one stimulatory agent is an anti-CD3 antibody.

7. The method according to claim 1, wherein a second stimulatory agent is an anti-CD28 antibody.

8. The method according to claim 1, wherein the flexible matrix consists of a polymer of dextran.

9. The method according to claim 1, wherein the nanomatrix is biodegradable.

10. The method according to claim 1, wherein the nanomatrix carries magnetic, paramagnetic or superparamagnetic nano-crystals, embedded into the flexible matrix.

11. The method according to claim 1, wherein the nanomatrix is 10 to 200 nm in size.

12. The method according to claim 1, wherein the stimulatory agent is attached at high density with more than 25 μ

g per mg nanomatrix.

13. The method according to claim 1, wherein the stimulated T cells are Treg cells.

14. The method according to claim 1, wherein the method is used within a closed cell culture system.

15. A pharmaceutical composition comprising a population of stimulated T cells produced according to the method of claim 14.

16. The use of a nanomatrix for in-vitro stimulation of T cells, the nanomatrix comprising

a) a flexible matrix, wherein said matrix is of polymeric material;

and

b) attached to said polymeric flexible matrix one or more polyclonal stimulatory agents which provide activation signals to the T cells;

wherein the nanomatrix is 1 to 500 nm in size.