Cultured thymus tissue transplantation promotes donor-specific tolerance to allogeneic solid organ transplants

1. A method for promoting donor-specific tolerance to an allogeneic heart, kidney, liver, lung, pancreas, small or large bowel, or combination thereof transplant obtained from a donor, in a recipient in need of a heart, kidney, liver, lung, pancreas, small or large bowel, or combination thereof transplant, the method comprising the steps of:

(a) removing the thymus of the recipient;

(b) treating the recipient with an induction immunosuppressive regimen comprising one or more immunosuppressive agent to deplete the recipient's T cells and/or to suppress the recipient's T cells from rejecting the transplanted solid organ;

(c) providing both (i) a heart, kidney, liver, lung, pancreas, small or large bowel, or combination thereof from a donor and (ii) a thymus gland from the donor;

(d) transplanting the heart, kidney, liver, lung, pancreas, small or large bowel, or combination thereof into the recipient;

(e) treating the recipient with a maintenance immunosuppressive regimen;

(f) providing partially T-cell depleted donor thymus tissue slices by subjecting thymus tissue, of the donor thymus gland, to a conditioning regimen for a period up to 21 days,

wherein the conditioning regimen comprises aseptically processing the donor thymus tissue in a thymus organ medium to produce the partially T-cell depleted donor thymus tissue slices,

wherein, upon receipt, the donor thymus tissue slices are assessed by pathology and show >50% of areas positive for keratin in a lacy staining pattern, show Hassall bodies, show cytokeratin 14 (CK14) staining in a lacy pattern, and show >90% intact nuclei, and

wherein, between day 5 and 9 of the conditioning regimen, the donor thymus tissue slices are assessed by pathology and show staining positive for keratin, show CK14 staining scattered throughout, show at least one Hassall body, and show intact nuclei; and

(g) implanting at least a portion of the partially T-cell depleted donor thymus tissue slices into the recipient,

wherein the dosage of the implanted partially T-cell depleted donor thymus tissue slices is about 1,000-20,000 m m of surface area of the partially T-cell depleted donor thymus tissue slices/recipient body surface area in m 2 , and

further wherein the implanted partially T-cell depleted donor thymus tissue slices induce thymopoiesis and tolerance in the recipient.

2. The method of claim 1 , further comprising the step of cryopreserving peripheral blood mononuclear cells from the donor for future use in a mixed lymphocyte reaction to demonstrate cellular tolerance.

3. The method of claim 2 , wherein the mixed lymphocyte reaction is performed using peripheral blood mononuclear cells from the recipient and cryopreserved peripheral blood mononuclear cells from the donor following the implantation of step (g).

4. The method of claim 3 , wherein the mixed lymphocyte reaction is performed with peripheral blood mononuclear cells from the recipient and cryopreserved peripheral blood mononuclear cells from the donor about 6 to 12 months following the implantation of step (g).

5. The method of claim 4 , wherein the mixed lymphocyte reaction is performed with peripheral blood mononuclear cells from the recipient and cryopreserved peripheral blood mononuclear cells from the donor after naïve T cells constitute about 10% of total T cells in the recipient.

6. The method of claim 1 , wherein the implanted partially T-cell depleted donor thymus tissue slices induce thymopoiesis in the subject within 12 months following the implantation of step (g).

7. The method of claim 1 , further comprising determining humoral tolerance by performing at least one assay for the development of humoral immunity and the absence of donor reactive antibodies toward the WIC of the donor.

8. The method of claim 1 , wherein the induction immunosuppressive regimen comprises administration of an immunosuppressive agent selected from the group consisting of glucocorticoid, an anti-thymocyte globulin, and alemtuzimab.

9. The method of claim 1 , wherein the maintenance immunosuppressive regimen comprises administration of a glucocorticoid.

10. The method of claim 9 , wherein the glucocorticoid comprises methylprednisolone sodium succinate.

11. The method of claim 10 , wherein the methylprednisolone sodium succinate is administered intravenously at no greater than 4 mg/kg/day after the implantation of step (g).

12. The method of claim 1 , wherein the maintenance immunosuppressive regimen comprises administration of a glucocorticoid plus tacrolimus or cyclosporine, and further administering mycophenylate mofetil, mycophenolic acid, or azathioprine.

13. The method of claim 1 , wherein the heart, kidney, liver, lung, pancreas, small or large bowel, or combination thereof is a portion of a heart, kidney, liver, lung, pancreas, small or large bowel, or combination thereof.

14. The method of claim 1 , wherein the heart is a pediatric heart.

15. The method of claim 1 , wherein the heart is an adult heart.

16. The method of claim 1 , further comprising evaluating the recipient for HLA class I and HLA class II panel reactive antibodies (“PRA”) score prior to transplanting the heart, kidney, liver, lung, pancreas, small or large bowel, or combination thereof.

17. The method of claim 1 , wherein recipients with HLA antibodies are cross-matched with potential donors.

18. The method of claim 16 , wherein, if a PRA score of >20% virtual cross-match is recorded, the method will further comprises performing plasmapheresis in the operating room at the time of heart, kidney, liver, lung, pancreas, small or large bowel, or combination thereof transplant in the recipient.

19. The method of claim 1 , wherein the heart, kidney, liver, lung, pancreas, small or large bowel, or combination thereof is HLA-mismatched.

20. The method of claim 19 , wherein HLA-mismatched is determined by typing HLA alleles HLA-A, HLA-B, HLA-C, HLA-DRB1, HLA-DQB1, HLA-DRB3, HLA-DRB4, HLA-DRB5, HLA-DQA1, HLA-DPB1, and HLA-DPA1 in the donor and the recipient.

21. The method of claim 20 , wherein HLA-mismatched comprises at least one of HLA alleles HLA-A, HLA-B, HLA-C, HLA-DRB1, HLA-DQB1, HLA-DRB3, HLA-DRB4, HLA-DRB5, HLA-DQA1, HLA-DPB1, and HLA-DPA1 is different between the donor and the recipient.

22. The method of claim 1 , wherein the implantation of step (g) comprises surgically implanting the at least a portion of the partially T-cell depleted donor thymus tissue slices into the quadriceps thigh muscle of the recipient.

23. The method of claim 22 , wherein the remaining non-implanted portion of the partially T-cell depleted donor thymus tissue slices is cryopreserved in liquid nitrogen for future transplantation into the recipient.

24. The method of claim 1 , wherein the conditioning regimen is for a period of about 12 days to about 21 days.

25. The method of claim 24 , wherein the conditioning regimen is for a period of about 12 days.

26. The method of claim 24 , wherein the conditioning regimen is for a period of about 21 days.

27. The method of claim 8 , wherein the induction immunosuppressive regimen comprises administration of at least one of rabbit anti-thymocyte globulin or equine anti-thymocyte globulin.

28. The method of claim 1 , wherein the induction immunosuppressive regimen comprises administration of alemtuzumab.

29. The method of claim 1 , wherein the maintenance immunosuppressive regimen comprises administration of one or more immunosuppressive agent selected from the group consisting of a calcineurin inhibitor, an inosine monophosphate dehydrogenase inhibitor, and azathioprine.

30. The method of claim 29 , wherein the immunosuppressive agent is a calcineurin inhibitor.

31. The method of claim 29 , wherein the immunosuppressive agent is an inosine monophosphate dehydrogenase inhibitor.

32. The method of claim 31 , wherein the inosine monophosphate dehydrogenase inhibitor is mycophenolate mofetil.

33. The method of claim 32 , wherein the mycophenolate mofetil is administered orally or intravenously.

34. The method of claim 31 , wherein the inosine monophosphate dehydrogenase inhibitor is mycophenolic acid.

35. The method of claim 1 , wherein the maintenance immunosuppressive regimen comprises a glucocorticoid selected from the group consisting of methylprednisolone, prednisone and prednisolone.

36. The method of claim 29 , wherein the calcineurin inhibitor is tacrolimus.

37. The method of claim 29 , wherein the calcineurin inhibitor is cyclosporine A.

38. The method of claim 1 , wherein the maintenance immunosuppressant regimen is weaned after naïve T cells reach 10% of total T cells.

39. The method of claim 1 , wherein the implanted partially T-cell depleted donor thymus tissue slices are biopsied at 2 to 12 months post-implantation to assess for evidence of thymopoiesis by immunochemistry.

40. The method of claim 1 , wherein the allogeneic heart, kidney, liver, lung, pancreas, small or large bowel, or combination thereof transplant is obtained from a donor.

41. The method of claim 1 , wherein the kidney, liver, lung, pancreas, small or large bowel, or combination thereof transplant is obtained from a living donor.

42. The method of claim 1 , wherein the thymus gland is obtained from a donor.

43. The method of claim 1 , wherein the thymus gland is obtained from a living donor.