Chimerism, Lymphocyte Recovery, and the Absence of Graft-Versus-Host Disease in Recipients of Mismatched Unrelated Combined Kidney and HSC Transplants for Tolerance Induction

It has been known for over 50 years that hematopoietic stem cell (HSC) chimerism induces tolerance to transplanted tissues and cells. However, the widespread application of this approach has been constrained by graft-versus-host disease (GVHD), the need for close genetic matching between donor and recipient, and the toxicity of conditioning the recipient to establish chimerism. We have demonstrated that full donor chimerism can be established with minimal toxicity in highly-mismatched unrelated and related kidney allograft recipients through nonmyeloablative conditioning followed by infusion of a bioengineered CD8⁺/TCR⁻ facilitating cell stem cell graft (FCRx), to avoid the risk of GVHD while achieving chimerism.

Twelve HLA-mismatched living donor renal transplant recipients have been entered into a phase 2 trial (IDE 13947) involving low-intensity conditioning (fludarabine, cyclophosphamide, 200 cGy TBI days −4 to −1). Patients received a living donor kidney transplant on day 0, followed by infusion of G-CSF cryopreserved FCRx on day +1. All subjects were discharged by post-operative day 3 and managed as outpatients. We herein present data regarding the immunologic recovery observed in our first 8 evaluable patients with > 6 months follow up.

All patients experienced an expected nadir period affecting leukocytes (ANC < 500, range 2–14 days) and platelets (< 50K, range 0–20 days). All patients demonstrated peripheral blood macrochimerism at 1 month post-transplant, ranging from 6% to 100%. Chimerism was gradually lost in two patients at 3 and 6 months post-transplant. Patients demonstrated in vitro evidence of donor-specific hyporesponsiveness (DSH) by MLR +/− CML as early as 3 months post-transplant; of interest, DSH also was observed and persisted in the two patients who lost peripheral blood chimerism. Patients at > 1 yr post-transplant are immunocompetent to respond to mitogen (PHA), MHC-disparate third-party alloantigen, and tetanus in in vitro proliferative assays. Immunologic reconstitution in kidney + FCRx recipients was characterized by a blunted return in CD4⁺ T cells, with inversion of the CD4/CD8 ratio. A preferential recovery of memory (CD4⁺/CD45RO⁺/CD62L^+/−) vs. naïve (CD4⁺/CD45RA⁺/CD62L⁺) T cells was observed. Although total number of CD4⁺/CD25⁺/CD127^lo/FoxP3⁺ T_reg was reduced initially, an increase in the CD4⁺ T_reg /CD4⁺Teff (CD4⁺/CD45RA⁺/CD62L⁻) ratio was seen in patients exhibiting durable chimerism. In addition, an expansion of central memory CD8⁺ T cells was observed in durably chimeric recipients. No patient developed donor-specific antibodies as assessed by flow cytometric analysis. The absence of GVHD correlated with in vitro hyporesponsiveness of the fully chimeric recipients against archived pre-treatment recipient APC.

Combined kidney + FCRx recipients demonstrate characteristic immunophenotypic and functional changes associated with reconstitution following transplantation; additional studies are required to determine whether these changes are mechanistically related to the persistence of chimerism and/or prevention of GVHD.

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