Expansion of alloantigen-reactive regulatory t cells

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.

1 - 8 . (canceled) 
     
     
         9 . A composition comprising from 10 7  to 10 11  restimulated donor-reactive regulatory T cells (Tregs) and a physiologically acceptable buffer, wherein the restimulated donor-reactive Tregs are produced using a method 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);   b) co-culturing CD4+, CD25+, CD127−/lo T cells isolated from peripheral blood mononuclear cells (PBMC) of a human recipient with said sBc under conditions effective in selectively expanding human donor-reactive regulatory T cells (Tregs); and   c) re-stimulating the donor-reactive Tregs by cross-linking CD3 and CD28 of the donor-reactive Tregs using monoclonal antibodies under conditions effective in producing restimulated donor-reactive Tregs that are CD4+, Helios+ and Foxp3+,   wherein the donor is a first human subject and the recipient is a second human subject and the donor is HLA-mismatched in relation to the human recipient.   
     
     
         10 . A method for treating or preventing rejection of a solid organ allograft, said method comprising: administering the composition of  claim 9  to the 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 administration is effective in reducing the likelihood of acute and/or chronic rejection. 
     
     
         13 . The method of  claim 10 , wherein said administration 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 to the human recipient in an amount effective to achieve lymphocyte depletion before administering the composition. 
     
     
         15 . The method of  claim 10 , wherein the composition is 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 composition is administered to the human recipient concurrently with sirolimus. 
     
     
         17 . The composition of  claim 9 , wherein the restimulated donor-reactive Tregs are CD27+, CD62L+. 
     
     
         18 . The composition of  claim 9 , wherein the restimulated donor-reactive Tregs have a Foxp3 promoter with a demethylated Treg-specific demethylation region.