Mixed Chimerism, Lymphocyte Recovery, and Evidence for Early Donor-Specific Unresponsiveness in Patients Receiving Combined Kidney and Bone Marrow Transplantation to Induce Tolerance

Background We have previously reported operational tolerance in patients receiving HLA-mismatched combined kidney and bone marrow transplantation (CKBMT). We now report on transient multilineage hematopoietic chimerism and lymphocyte recovery in five patients receiving a modified CKBMT protocol, and evidence for early donor-specific unresponsiveness in one of these patients. Methods Five patients with end-stage renal disease received CKBMT from HLA-mismatched, haploidentical living related donors following modified non-myeloablative conditioning. Polychromatic flow cytometry (FCM) was used to assess multilineage chimerism where evaluable and lymphocyte recovery post-transplant. Limiting dilution analysis was used to assess helper-T-lymphocyte reactivity to donor antigens. Results Transient multilineage mixed chimerism was observed in all patients but chimerism became undetectable by 2 weeks post-CKBMT. A marked decrease in T and B lymphocyte counts immediately following transplant was followed by gradual recovery. Initially recovering T cells were depleted of CD45RA+/CD45RO− “naïve-like” cells, which have shown strong recovery in two patients and CD4/CD8 ratios increased immediately following transplant but then declined markedly. NK cells were enriched in the peripheral blood of all patients following transplant. For Subject 2, a pre-transplant limiting dilution assay revealed T helper cells recognizing both donor and third-party PBMCs. However, the anti-donor response was completely undetectable by Day 24, while third-party reactivity persisted. Conclusion These results characterize the transient multilineage mixed hematopoietic chimerism and recovery of lymphocyte subsets in patients receiving a modified CKBMT protocol. The observations are relevant to the mechanisms of donor-specific tolerance in this patient group

Peritransplant energy changes and their correlation to outcome after human liver transplantation

BACKGROUND: The ongoing shortage of donor livers for transplantation and the increased use of marginal livers necessitate the development of accurate pretransplant tests of viability. Considering the importance energy status during transplantation, we aimed to correlate peritransplant energy cofactors to posttransplant outcome and subsequently model this in an ex vivo setting. METHODS: Sequential biopsies were taken from 19 donor livers postpreservation, as well as 30 min after portal venous (PVR) and hepatic arterial reperfusion (HAR) and analyzed by LC-MS for energetic cofactors (ATP/ADP/AMP, NADH/NAD, NADPH/NADP, FAD, GSSG/GSH). Energy status was correlated to posttransplant outcome. In addition, 4 discarded human DCD livers were subjected to ex vivo reperfusion, modeling reperfusion injury and were similarly analyzed for energetic cofactors. RESULTS: A rapid shift towards higher energy adenine nucleotides was observed following clinical reperfusion, with a 2.45-, 3.17- and 2.12-fold increase in ATP:ADP, ATP:AMP and energy charge (EC) after PVR, respectively. Seven of the 19 grafts developed early allograft dysfunction (EAD). Correlation with peritransplant cofactors revealed a significant difference in EC between EAD and normal functioning grafts (0.09 vs. 0.31, P<0.05). In the simulated reperfusion model, a similar trend in adenine nucleotide changes was observed. CONCLUSION: A preserved energy status appears critical in the peritransplant period. Levels of adenine nucleotides change rapidly following reperfusion and ratios of ATP/ADP/AMP following reperfusion are significantly correlated to graft function. Using these markers as a viability test in combination with ex vivo reperfusion may provide a useful predictor of outcome that incorporates donor, preservation and reperfusion factors

Regulatory cell therapy in kidney transplantation (The ONE Study): a harmonised design and analysis of seven non-randomised, single-arm, phase 1/2A trials

Background Use of cell-based medicinal products (CBMPs) represents a state-of-the-art approach for reducing general immunosuppression in organ transplantation. We tested multiple regulatory CBMPs in kidney transplant trials to establish the safety of regulatory CBMPs when combined with reduced immunosuppressive treatment. Methods The ONE Study consisted of seven investigator-led, single-arm trials done internationally at eight hospitals in France, Germany, Italy, the UK, and the USA (60 week follow-up). Included patients were living-donor kidney transplant recipients aged 18 years and older. The reference group trial (RGT) was a standard-of-care group given basiliximab, tapered steroids, mycophenolate mofetil, and tacrolimus. Six non-randomised phase 1/2A cell therapy group (CTG) trials were pooled and analysed, in which patients received one of six CBMPs containing regulatory T cells, dendritic cells, or macrophages; patient selection and immunosuppression mirrored the RGT, except basiliximab induction was substituted with CBMPs and mycophenolate mofetil tapering was allowed. None of the trials were randomised and none of the individuals involved were masked. The primary endpoint was biopsy-confirmed acute rejection (BCAR) within 60 weeks after transplantation; adverse event coding was centralised. The RTG and CTG trials are registered with ClinicalTrials.gov, NCT01656135, NCT02252055, NCT02085629, NCT02244801, NCT02371434, NCT02129881, and NCT02091232. Findings The seven trials took place between Dec 11, 2012, and Nov 14, 2018. Of 782 patients assessed for eligibility, 130 (17%) patients were enrolled and 104 were treated and included in the analysis. The 66 patients who were treated in the RGT were 73% male and had a median age of 47 years. The 38 patients who were treated across six CTG trials were 71% male and had a median age of 45 years. Standard-of-care immunosuppression in the recipients in the RGT resulted in a 12% BCAR rate (expected range 3·2–18·0). The overall BCAR rate for the six parallel CTG trials was 16%. 15 (40%) patients given CBMPs were successfully weaned from mycophenolate mofetil and maintained on tacrolimus monotherapy. Combined adverse event data and BCAR episodes from all six CTG trials revealed no safety concerns when compared with the RGT. Fewer episodes of infections were registered in CTG trials versus the RGT. Interpretation Regulatory cell therapy is achievable and safe in living-donor kidney transplant recipients, and is associated with fewer infectious complications, but similar rejection rates in the first year. Therefore, immune cell therapy is a potentially useful therapeutic approach in recipients of kidney transplant to minimise the burden of general immunosuppression. Funding The 7th EU Framework Programme