Immune Reconstitution Kinetics And Its Impact On Outcome After Umbilical Cord Blood Transplantation With Myeloablative Regimen Without Antithymocyte Globulin

Background Unrelated umbilical cord blood (UCB) is an alternative donor stem cell source with the advantages of faster procurement and harmless to the donor. Powerful graft-versus-leukemia (GVL) effect and a low risk of severe graft-versus-host disease (GVHD) enhance the appeal of UCB transplantation (UCBT) for patients with high-risk hematopoietic malignances. The long-term survivors who underwent UCBT had a higher quality of life because of low-relapse rate and low risk of chronic GVHD. Compared to recipients of bone marrow transplants, patients receiving UCBT had significantly slow neutrophil and lymphocyte recovery because UCBT recipients received one log less cells. While rate of neutrophil recovery is the most important determinant of risk for early infections, reconstitution of the innate lymphoid and the adaptive immune systems play a major role in the subsequent phase of transplant recovery, long-term resistance to opportunistic infections, development of GVHD, and GVL effects. Transplant protocols (GVHD prophylaxis and conditioning regimen) play an important role in immune reconstitution (IR) besides host factors (such as age, disease, and initial immune status), the degree of genetic differences between donor and recipient, and source of HSC. Following the transplants performed after myeloablative conditioning regimens, IR will depend upon the ability of the haematopoietic graft to generate de novo lymphoid and myeloid lineage cells and on the function of mature cells contained in the graft. ATG is a polyclonal IgG that has been used in conditioning regimens for UCBT both to promote engraftment and reduce GvHD.As a long half-life of the drug, ATG may led to delayed T-cell reconstitution in UCBT. On the other hand, omitting ATG prior to UCBT may be an increased incidence of GVHD which may have a negative impact on survival. So far, only one report of IR after UCBT with no use of ATG (Chiesa et al,2012), the major of cases were non-malignant disease and using reduced intensity regimen. Data on IR after UCBT with myeloablative regimens without ATG are scare.Objective The severity of complications of transplantation is governed mainly by the status of immune reconstitution. Compared to other stem cell sources, umbilical cord blood transplantation (UCBT) is characterized by an impairment of early immune reconstitution because of a lower T-cell dose, the naivety of cord blood T-cells infused and the use of in vivo T-cell depletion. Given that the omission of in vivo T-cell depletion may promote early immune reconstitution and reduce infection related morbidity and mortality in patients undergoing unrelated UCBT, we investigated the kinetics of immune reconstitution after UCBT with myeloablative regimens without antithymocyte globulin (ATG).Methods (1) All patients who received an unrelated UCBT at Anhui provincial hospital from July 2011 to Dec.2013 were included in this study. The patients received myeloablative conditioning without ATG and cyclosporin A (CSA) and mycophenolate mofetile as GVHD prophylaxis in UCBT protocols. (2) Blood cells and lymphocyte recovery after UCBT were monitored dynamically. (3)Lymphocyte subsets were assessed by flow-cytometry after fluorochrome-conjugated monoclonal antibodies for T-cells (CD3, CD4, and CD8), B-cells (CD19), NK-cells (CD56 and CD16), naive and memory T-cells (CD45RA and CD62L). Immunoglobulin levels were measured by immunoturbidimetry assay. These analyses were done approximately at 1-6,9,12,18, 24 months post-transplantation. (3) The recovery of thymopoiesis using a polymerase chain reaction (PCR)-based assessment of T-cell receptor excision circles (TRECs) in UCBT recipients was analyzed. (4) Phenotypic analysis of NK receptors, intracellular cytokine staining for IFN-y and TNF-a, and CD 107a degranulation activities of NK cells after UCBT. (5)The relationship between immune reconstitution and clinical outcome was analyzed.Results (1) Between July 2011 and Dec.2013,122 patients included in this study. Engraftment rate was 93.4%. Twenty-one cases died of severe infection and the median time of death from infection was 91 days after transplantation (range,31-413 days). Sixteen patients relapsed; the 2-year cumulative incidences of relapse were 16.95%. Up to September 30,2014, the median duration of follow-up for the surviving UCBT recipients was 559 days (range,265-1159 days).The 2-year probabilities of overall survival (OS) and disease-free survival (DFS) were 59%, respectively. (2) From 1 month after UCBT, the absolute monocyte count (AMC) and absolute lymphocyte count (ALC) sustained to increase until month 18 post-transplantation. (3) CD56+and or CD 16* natural killer (NK) cells recovered more rapidly than other lymphocyte subsets. The median absolute NK cell count recovered to normal at 1 month after UCBT. Frequency of the CD56bright population was increased and maintained one year after transplant, while CD56dim population was reduced from 1 month. On the other hand, a marked increase in CD16+CD56- NK subpopulation in the peripheral blood (PB) was observed. The majority of CD16+ CD56- NK subsets express CDllb and CD57, hardly express CD27, suggesting their mature characteristics. CD56bnght NK subset expresses a variety of receptors, showed an increased production of IFN-γ and TNF-a, contained much more perform and granzymes, and showed a stronger CD 107a mobilization, sugesting the NK subsets are fully functional.(4) A rapid increase of CD3+and CD8+T-cell counts after UCBT were observed. The median absolute CD3+and CD8+T-cell counts reached normal at month 3 and month 2, respectively. CD4+T cell recovery delayed at early phase after UCBT, the median CD4+T cell reached normal at month 5 after UCBT. The median ratio of naive CD4+T cells reached normal at month 9, and in the same time, detectable post-UCBT thymopoiesis were increased. (5) B-cell recovery was delayed with gradual increase in the number of B-cells, starting around 3 months after the transplantation. The median CD19+-cell count at days 30,60 and 90 was 1/μl,2/μl, and 7/μl, respectively. The median CD19+-cell count reached normal at month 5, the counts continue to increase beyond the normal range at month 18 post-UCBT. The median serum IgM, IgG and IgA levels reached the normal range at month 3, month 4, and month 6 after UCBT, respectively. (6) The patients who experienced early life-threatening infections after UCBT showed slow recovery monocyte, lymphocyte and T-lymphocyte subsets during the first three months. Delayed B-cell recovery after month 3 is found in patients with systemic GVHD and its treatment.Conclusions (1)Omission of ATG in myeloablative conditioning promotes rapid expansion of total lymphocyte, total CD3+T cells,and CD8+T cell subsets in the recipients of UCBT.CD4+T cell subsets and CD19+B cells recovery delayed in the first 3 months after UCBT, which may be reflected the impact of the myeloablative conditioning regimen. (2) The patients with delayed reconstitution of total lymphocyte and T cell subsets were specifically associated with increased risk of infection-related mortality, but the increased risk is limited to the early post-transplant period after UCBT. Delayed B-cell reconstitution after three months was associated with systemic acute GVHD and its treatment, early B-cell recovery delayed before month 3 after UCBT may be associated with full-intensity conditioning therapy.