EXPANSION OF ALLOANTIGEN-REACTIVE REGULATORY T CELLS

• US 20150110761A1
• Filed: 03/01/2013
• Published: 04/23/2015
• Est. Priority Date: 03/02/2012
• Status: Active Grant

• Current Assignee: Regents of the University of California (University of California)
• Original Assignee: Regents of the University of California (University of California)
• Inventors: Tang, Qizhi, Bluestone, Jeffrey A.
• Granted Patent: US 9,801,911 B2
• Time in Patent Office: Days
• US Class Current: 424/93.71
• CPC Class Codes: A61K 2239/26 Universal/off- the- shelf cellular immunotherapy; Allogenic cells or means to avoid rejection
• CPC Class Codes: A61K 2239/38 characterised by the dose, timing or administration schedule
• CPC Class Codes: A61K 2300/00 Mixtures or combinations of active ingredients, wherein at least one active ingredient is fully defined in groups A61K31/00 - A61K41/00
• CPC Class Codes: A61K 31/436 the heterocyclic ring system containing a six-membered ring having oxygen as a ring hetero atom, e.g. rapamycin
• CPC Class Codes: A61K 31/5377 not condensed and containing further heterocyclic rings, e.g. timolol
• CPC Class Codes: A61K 31/573 substituted in position 21, e.g. cortisone, dexamethasone, prednisone or aldosterone
• CPC Class Codes: A61K 35/17 Lymphocytes; B-cells; T-cells; Natural killer cells; Interferon-activated or cytokine-activated lymphocytes when activated by a specific antigen A61K39/00
• CPC Class Codes: A61K 39/001 Preparations to induce tolerance to non-self, e.g. prior to transplantation
• CPC Class Codes: A61K 39/461 characterised by the cell type used
• 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 39/4621 immunosuppressive or immunotolerising
• CPC Class Codes: A61K 39/46434 Antigens related to induction of tolerance to non-self
• CPC Class Codes: A61P 37/00 Drugs for immunological or allergic disorders
• CPC Class Codes: A61P 37/06 Immunosuppressants, e.g. drugs for graft rejection
• CPC Class Codes: C12N 2501/04 Immunosuppressors, e.g. cyclosporin, tacrolimus
• CPC Class Codes: C12N 2501/2304 Interleukin-4 (IL-4)
• CPC Class Codes: C12N 2501/515 CD3, T-cell receptor complex
• CPC Class Codes: C12N 2502/1107 B cells
• CPC Class Codes: C12N 2502/30 tumour cells
• CPC Class Codes: C12N 5/0635 B lymphocytes
• CPC Class Codes: C12N 5/0637 : Immunosuppressive T lymphocytes, e.g. regulatory T cells or Treg

Abstract

The present disclosure relates generally to the manufacture of regulatory T cells (Tregs) for use in immunotherapy. In particular, the present disclosure relates to robust approaches for the expansion of alloantigen-reactive Tregs ex vivo. Alloantigen-reactive Tregs produced in this way are suitable for the induction and/or maintenance of immunologic tolerance in recipients of allogeneic transplants.

Claims

1. A method for the production of human, donor-reactive regulatory T cells (Tregs), comprising:

a) co-culturing CD19+ B cells of a human donor with irradiated CD40L+ human leukemia feeder cells under conditions effective in producing stimulated B cells (sBc); and

b) co-culturing CD4+, CD25+, CD127−

/lo T cells of a human recipient with said sBc under conditions effective in selectively expanding human donor-reactive regulatory T cells (Tregs).

2. The method of claim 1, further comprising step c) re-stimulating said donor-reactive Tregs by cross-linking CD3 and CD28 of said donor-reactive Tregs under conditions effective in producing re-stimulated donor-reactive Tregs.

3. The method of claim 2, wherein step c) commences 9-12 days after step b) commences.

4. The method of claim 1, where said donor-reactive Tregs are CD4+, Helios+ and Foxp3+.

5. The method of claim 1, further comprising a step before a) of isolating CD4+, CD25+, CD127−

/lo T cells from cryopreserved peripheral blood mononuclear cells (PBMC) obtained from said human recipient.

6. The method of claim 1, wherein step a) comprises co-culturing said B cells and said feeder cells in medium comprising insulin, transferrin, interleukin-4 and cyclosporine A.

7. The method of claim 1, wherein step b) comprises co-culturing said sBc and said CD4+, CD25+, CD127−

/lo T cells in medium comprising interleukin-2 after said sBc have been irradiated.

8. The method of claim 1, wherein said re-stimulated donor-reactive Tregs comprise 200 fold to 2000 fold more cells than said CD4+, CD25+, CD127−

/lo T cells at the onset of step b).

9. A composition comprising the restimulated donor-reactive Tregs produced using the method of claim 2, and a physiologically acceptable buffer.

10. A method for treating or preventing rejection of a solid organ allograft, said method comprising:

administering from 10⁷ to 10¹¹ of the restimulated donor-reactive Tregs produced using the method of claim 2, to a human recipient of the solid organ allograft.

11. The method of claim 10, wherein said solid organ allograft is selected from the group consisting of cardiac, lung, cardiac/lung, kidney, pancreas, kidney/pancreas, liver, intestine, and skin allografts.

12. The method of claim 10, wherein said administering is effective in reducing the likelihood of acute and/or chronic rejection.

13. The method of claim 10, wherein said administering is effective in achieving one or more of the group consisting of increasing Treg percentages over baseline, increasing donor-reactive Treg frequency, increasing donor-reactive Treg activity, and induction of tolerance gene expression profiles in PBMC and/or transplant tissue.

14. The method of claim 10, further comprising subjecting the human recipient to a Treg-supportive immunosuppression regimen comprising administering rabbit anti-thymocyte globulin in an amount effective to achieve lymphocyte depletion to the human recipient before administering the restimulated donor-reactive Tregs.

15. The method of claim 10, wherein the restimulated donor-reactive Tregs are administered to the human recipient concurrently with prednisone, mycophenolate mofetile and tacrolimus at doses below standard of care.

16. The method of claim 10, wherein the restimulated donor-reactive Tregs are administered to the human recipient concurrently with sirolimus.