Feasibility, long-term safety, and immune monitoring of regulatory T cell therapy in living donor kidney transplant recipients

Abstract

Short-term outcomes in kidney transplantation are marred by progressive transplant failure and mortality secondary to immunosuppression toxicity. Immune modulation with autologous polyclonal regulatory T cell (Treg) therapy may facilitate immunosuppression reduction promoting better long-term clinical outcomes. In a Phase I clinical trial, 12 kidney transplant recipients received 1-10 × 10⁶ Treg per kg at Day +5 posttransplantation in lieu of induction immunosuppression (Treg Therapy cohort). Nineteen patients received standard immunosuppression (Reference cohort). Primary outcomes were rejection-free and patient survival. Patient and transplant survival was 100%; acute rejection-free survival was 100% in the Treg Therapy versus 78.9% in the reference cohort at 48 months posttransplant. Treg therapy revealed no excess safety concerns. Four patients in the Treg Therapy cohort had mycophenolate mofetil withdrawn successfully and remain on tacrolimus monotherapy. Treg infusion resulted in a long-lasting dose-dependent increase in peripheral blood Tregs together with an increase in marginal zone B cell numbers. We identified a pretransplantation immune phenotype suggesting a high risk of unsuccessful ex-vivo Treg expansion. Autologous Treg therapy is feasible, safe, and is potentially associated with a lower rejection rate than standard immunosuppression. Treg therapy may provide an exciting opportunity to minimize immunosuppression therapy and improve long-term outcomes.

Keywords: clinical research/practice; clinical trial; immune regulation; immunosuppression/immune modulation; immunosuppressive regimens - minimization/withdrawal; kidney transplantation/nephrology; kidney transplantation: living donor; monitoring: immune; translational research/science.

Figure 1

ONE Study immunosuppression protocol for Reference and Treg Therapy kidney transplant recipients. MMF, mycophenolate mofetil; nTreg, naturally occurring Treg; Tx, transplantation; BW, body weight [Color figure can be viewed at wileyonlinelibrary.com]

Figure 2. Trial cohorts

Figure 3. Clinical outcomes.

(A) Rejection-free graft survival at 4 years posttransplant. (B) CMV- and BK-free survival over 4 years posttransplant. BK and CMV infection defined by blood PCR titers meeting criteria for immunosuppression adjustment or evidence of tissue invasive disease. (C) Adverse events per 100 patient study years over first 60 weeks posttransplant (censored for events occurring at >20 per 100 PSY). (D) Serious adverse events per 100 patient study years over first 60 weeks posttransplant. CMV, cytomegalovirus; BKV, BK virus

Figure 4

Representative H&E section from a renal transplant biopsy in cell therapy-treated patients at 8 months posttransplant. Focal infiltrates as demonstrated, that were not typical of rejection, were seen in the first five patients treated with regulatory T cells with otherwise stable graft function [Color figure can be viewed at wileyonlinelibrary.com]

Figure 5

(A) Pre-expansion phenotype predicts feasibility of successful Treg expansion. (B–E) Treg detection post-infusion. (A) Patients for whom Treg expansion failed (red triangles) or was difficult (yellow triangle) showed reduced frequencies of lymphocytes, HLA-DR⁺ or CD57⁺ activated CD4⁺ T cells as well as higher proportions of naïve conventional and regulatory T cells prior to transplantation as compared to patients with successful expansion (n = 11). (B) Dose-dependent increase in absolute numbers of CD4⁺CD25^highCD127^low regulatory T cells upon Treg infusion 2 weeks after transplantation determined by whole blood flow cytometry in samples collected from patients receiving either 1 × 10⁶, 3 × 10⁶, 6 × 10⁶, or 10 × 10⁶ Tregs/kg (n = 3 per dose) prior to transplantation (V01), at 2 (V03), 4 (V04), 8 (V05), 12 (V06), 24 (V07), 36 (V08), 48 (V09), and 60 (V10) weeks posttransplant. Statistical analysis by mixed-effects analysis and Tukey's multiple comparison test.*p < .05. (C) Trend toward dose-dependent increase in absolute numbers of CD4⁺CD25^highCD127^low regulatory T cells at the end of the observation period (60 weeks posttransplant). Statistical analysis by linear regression. (D) Cytometry time of flight (CyTOF) example tSNE plots from one patient pretransplantation (V01), and at 2 (V03) and 60 (V10) weeks posttransplantation, highlighting distinct Treg clusters. (E) CCR7 expression on peripheral Tregs pretransplant (V01) and at 60 weeks posttransplant (V10) in 6 Treg-treated patients by CyTOF. Paired t test, **p < .01 [Color figure can be viewed at wileyonlinelibrary.com]

Figure 6. Effect of Treg therapy on immune cell composition.

(A) Treg infusion of either 1 × 10⁶, 3 × 10⁶, 6 × 10⁶, and 10 × 10⁶ cells/kg. (n = 3 patients/dose) did not lead to a significant increase in HLA-DR expression on CD14⁺ monocytes 1 day (F02) or 7 days later (F03). (B,C) Differences in posttransplant changes in CD14^highCD16⁺ monocytes and marginal zone-like B cells. Scatter plots of absolute numbers in whole blood samples collected pretransplant (V01) and at the end of the observation period (60 weeks posttransplant, V10) from reference (n = 19) and cell therapy trial patients (n = 12). Statistical analysis by Wilcoxon matched-pairs signed rank and Dunn's multiple comparison test. *p < .05. (D) Trend toward dose-dependent increase in absolute numbers of marginal zone-like B cells upon Treg infusion 2 weeks after transplantation determined by whole blood flow cytometry in samples collected from patients receiving either 1 × 10⁶, 3 × 10⁶, 6 × 10⁶, or 10 × 10⁶ cells/kg. (n = 3 patients per dose) prior to transplantation (V01), at 2 (V03), 4 (V04), 8 (V05), 12 (V06), 24 (V07), 36 (V08), 48 (V09), and 60 (V10) weeks posttransplant. (E) Trend toward dose-dependent increase in absolute numbers of marginal zone-like B cells at the end of the observation period (60 weeks posttransplant, V10). Statistical analysis by linear regression [Color figure can be viewed at wileyonlinelibrary.com]