Early evaluation of immune reconstitution following allogeneic CD3/CD19-depleted grafts from alternative donors in childhood acute leukemia

Abstract

Graft engineering procedures for hematopoietic SCT (HSCT) may improve the chance of success in matched unrelated donor (MUD) and haploidentical donor transplantations. Successful donor immune reconstitution is important to mediate GVL effects in reduced-intensity conditioning (RIC) HSCT. We prospectively investigated early immune reconstitution and clinical outcome in 30 CD3/CD19-depleted MUD (n ¼ 15) or HP (n ¼ 15) HSCTs for high-risk childhood leukemia using a fludarabine-based RIC without serotherapy. The graft consisted of a mean of 10.5 Â 10 6 /kg CD34 þ , 77 Â 10 3 /kg CD3 þ and 39 Â 10 6 /kg CD56 þ cells. After transplantation, 86% of the patients engrafted. In all, 13% of patients had 4grade 3 acute GVHD. Natural killer (NK) cell, DC and T-cell recovery achieved normal values within the first 60 days after transplantation. DC recovery was dominated by the DC2 À subset. NK-cell phenotype was altered and cytotoxicity was lower compared with their donors. EFS was 50 ± 9% (73 ± 11% for those in CR1 and 26 ± 11% for those with advanced disease). Faster DC2 À recovery was associated with better outcome, especially in the MUD setting. In summary, CD3/CD19-depleted HSCT with fludarabine-based RIC without serotherapy resulted in favorable patient survival, and rapid NK, DC and T-cell recovery. T-cell-depleted grafts have been proposed as a strategy to avoid GVHD. For example, CD34-selected HSCT followed by DLI has been proposed to maximize the effects of GVL with low rates of GVHD in HLA-matched HSCT. 3 However, secondary graft rejection is an important problem, especially in the MUD transplant setting. To facilitate engraftment, CD3/CD19-depleted grafts containing different pools of cells have been proposed to be preferable to T-cell-depleted grafts in MUD and HP HSCT in our unit. 4 In addition to T-cell depletion, B-cell depletion was used to prevent EBV-related lymphoproliferative disease. To reduce toxicity, we developed a reduced intensity conditioning (RIC) with i.v. BU, fludarabine and thiotepa. In addition, a high-dose steroid was used to further suppress recipient T-cell function to prevent graft rejection. We did not use antithymocyte globulin because of its long half-life and its potential to suppress T cells and natural killer (NK) cells, resulting in a high risk of infection and reduced GVL. 5 Clinical GVL effect has been associated with a fast recovery of alloreactive effector cells, such as NK cells in HP settings and T cells in MUD settings. 6,7 A recent report has described a delay in T-cell reconstitution using CD3/CD19-depleted grafts and RIC in the HP setting. 8 Furthermore, Dulphy et al. 9 have described an immature NK-cell population during the early post-transplant period following unmanipulated HLA-matched HSCTs. Apart from effector cells, the reconstitution of other immune cells such as DCs has not been systematically studied after T-cell-depleted HSCTs.

A phase I/II trial of interleukin-15–stimulated natural killer cell infusion after haplo-identical stem cell transplantation for pediatric refractory solid tumors

“…The NK cell isolation procedure was performed as described previously [25]. The following fluorochrome-labeled monoclonal antibodies (mAbs) against human antigens were obtained from Becton Dickinson: CD3 phycoerythrin (PE)-Cy7, CD158b1 fluorescein isothiocyanate (FITC), CD20-PE, CD45-FITC, CD69-FITC, NKG2D-alllophycocyanin (APC), CD45RA-PE-Cy5, CD127-APC, CD27-APC and CCR7-PE.…”

Prognostic factors and outcomes for pediatric patients receiving an haploidentical relative allogeneic transplant using CD3/CD19-depleted grafts

“…Phenotyping of natural killer (NK) cells, T lymphocytes, T lymphocyte subsets, B lymphocytes, NKT lymphocytes and dendritic cells (DCs) was performed on fresh samples of whole blood by multiparametric flow cytometry as previously described. 7 The following fluorochrome-labeled monoclonal Abs against human Ags were obtained from Becton Dickinson (San Jose, CA, USA): CD3 PE-Cy7, CD20-PE, CD45-FITC, lineage-FITC, HLADR-APC-Cy7, CD11c-PECy5, CD45RA-PE-Cy5 and CCR7-PE. Fluorochrome-labeled monoclonal Abs against CD19-PE, CD56-APC and CD25-PE were obtained from Beckman Coulter (Fullerton, CA, USA).…”

Refinement of treatment strategies in ex vivo T-cell-depleted haploidentical SCT for pediatric patients

“…[10][11][12][13][14] Although all of our patients received GVHD prophylaxis with calcineurin inhibitors and mycophenolate mofetil, the incidences of severe acute and chronic GVHD were acceptable for our partial depletion of T cells.…”

“…There have been several reports on HHCT using CD3-depleted grafts in pediatric patients. 2,[10][11][12][13][14] The Tübingen group reported their experience with CD3/19-depleted HHCT using reduced-intensity conditioning in 38 pediatric patients. 2 The authors reported a primary engraftment rate of 83%, with GR occurring in 17% of patients.…”