George W. Burke | University of Miami | Miami, FL

University of Miami | Miami, FL

CLINICAL OBJECTIVE: To successfully withdraw immunosuppression in recipients of living-related donor 1 haplotype matched kidney transplants by treatment with donor stem cell infusions and Campath-1H (alemtuzumab) anti-CD52 humanized monoclonal antibody induction to cause a functional state of specific immunologic tolerance, a significant advance in organ transplant therapy.

Campath-1H will be administered intravenously in two doses of 0.3 mg/kg: 1) on the day of surgery before kidney revascularization, and 2) again four days later. This regimen will include maintenance tacrolimus (tacro) and mycophenolate mofetil (MMF) at one-half the usual dosages, but no corticosteroids. Infusion #1 of donor iliac crest marrow, obtained at surgery (as in our previous whole marrow infusion protocols), will then be purified by the Isolex® technology for CD34+ stem cells, cryopreserved and administered on post-operative day #5 when all the Campath-1H targeted lymphoid cell subpopulations will have reached their nadir. At 4 to 6 months post-operatively, as T and B cells begin to reappear, tacro will be totally discontinued after initiation of overlapping therapy with sirolimus (siro) with target through levels of 8-15 ng/ml. Infusion #2 of freshly obtained donor stem cells, mobilized and purified from peripheral blood, will then be administered. The rationale for this is that, aside from the potent early Campath-1H lymphoablative effect on the recipient marrow and PBL by later substituting sirolimus for tacrolimus, the specific regulatory effect of co-stimulatory signal blockade by donor stem cell infusion #2 is more likely. However, fewer early acute rejections have been observed with Campath-1H using (half-dose) tacro and MMF than with siro; therefore, the plan for the early (4-6 month) half-dose tacro course.

After 1 year, siro withdrawal will take place over 6 months if there is no clinical or biopsy evidence of rejection (Banff Grade 1A or higher). At year 2, again provided there is no rejection (biopsy), MMF withdrawal will take place over the next 6 months. Follow-up will be over the ensuing five years.

MECHANISTIC STUDIES:

1. We will use the PCR-Flow assay in peripheral blood, monthly, as well as in iliac crest marrow at 6 months and yearly, to measure donor subset chimerism in the recipients. A significantly higher percent-age of multilineage chimerism (of several donor immunohematologic subsets) is anticipated than in our previously reported series of marrow-infused recipients, putatively a reflection of donor-specific immunologic unresponsiveness.
2. The newly developed micro-CML and enzyme-linked ELISPOT assays will be tested to measure sequential donor and recipient chimeric marrow and peripheral blood immunohematologic cell subset regulatory effects.
3. Yearly kidney biopsies (x4) from the time of surgery, and before, during and after immunosuppression withdrawal, will be performed. Lymphoid cells derived (cultured) from the biopsies will be studied for chimerism and function.
4. To test for clonal deletion, we will use limiting dilution analysis and the immunoscope or TcLandscape program, the latter to phenotype and follow Vbeta family deletion, in collaboration with Dr. Jean-Paul Soulillou (INSERM, Nantes, France) to take advantage of this ITN Core laboratory.