Non-Traditional CD4+ CD25–CD69+ Regulatory T Cells Is Correlated To Leukemia Relapse After Allogeneic Hematopoietic Stem Cell Transplantation
Authors: Zhao Xiaosu , Wang Xuhua , Chang Yingjun , Zhao Xiangyu , Zhang Xiaohui , Liu Daihong , Xu Lanping , Liu Kaiyan , Huang Xiaojun 
Published: October 2019

Abstract

Introduction
Leukemia relapse is the most common cause of treatment failure after allogeneic hematopoietic stem cell transplantation (allo-HSCT). The biological mechanism of relapse is still not completely clear. Allo-HSCT can be considered as a kind of immunotherapy directed toward malignant hematologic disease. Confirmation the correlation between clinical relapse and specific subset of T cells and further elucidating the immune mechanism behind this phenomenon would be beneficial to explore new methods of adoptive immunotherapy. Accumulative evidence showed that regulatory T cells (Tregs) might be involved in immune escape mechanism and associated with the failure of host to trigger efficient immunological antitumor response. Recently, a kind of non-traditional CD4+ CD25–CD69+ T cells was found to be involved in disease progression in tumor-bearing mouse models and cancer patients. In this study, we attempted to define whether this subset of T cells was related to leukemia relapse after allo-HSCT and also demonstrate the potential immune regulatory mechanism behind this phenomenon.

Methods
Twenty-nine patients with malignant hematological disease treated with non-T-cell-depleted allo-HSCT at the Peking University Institute of Hematology from November 2009 to April 2011 including patients undergoing hematological relapse (n=22) and with detectable minimal residual disease (MRD, n=7) were selected as the initial subjects in this study. The other fifty-six patients who received allo-HSCT from October 2009 to July 2010 were also enrolled for prospective study. The bone marrow of the initial 29 subjects was collected at the time of hematological relapse or detecting MRD. The MRD status of those 56 patients was examined at regular time points: +30 day (d), +60d, +90d, +180d, +270d, +360d. Bone marrow samples from patients at above time points were collected for the MRD examination and counting of CD4+CD25-CD69+ cells by flow cytometry (FCM).

Results
The frequency of CD4+CD25-CD69+ T cells in healthy donors’ bone marrow was 2.79% (range, 2.11-4.94%). However, the frequency of this subset of T cell was significantly increased in patients with detectable MRD (7.60%, range, 4.53-9.14%, P=0.008), or undergoing hematological relapse (12.96%, range, 8.62-20.49%, P<0.001) in compared to that of control group. The percentage of this subset of T cells significantly decreased (n=19) after intervention treatment. We also analyzed the reconstitution of CD4+ CD25–CD69+ T cells of 56 patients at given time points after allo-HSCT. In the follow-up, 7 patients (12.5%) got hematological or extramedullary relapse and 7 patients (12.5%) met the criteria of positive MRD. It showed that there was no time-dependent increase or decrease in this subset of T cells in patients without relapse or detectable MRD. We compared the frequency of CD4+CD25-CD69+ T cells between groups of patients with and without leukemia relapse or relapse indication at each time point after transplantation. There was significant difference in the frequency of bone marrow CD4+CD25-CD69+ T cells only at +60d (P=0.046, at other time points, P>0.05). The incidence of MRD+ or relapse in high frequency of CD4+CD25-CD69+ T cells group (>7%) was distinctly higher than that of low frequency of CD4+CD25-CD69+ T cells group at +60d, +90d and +270d after transplant. However, our preliminary data indicated that CD4+CD25-CD69+ T cells might not display immune regulatory function via cytokine secretion.

Conclusions
This study gave the first clinical evidence of correlation between non-traditional CD4+CD25-CD69+ T cells and leukemia relapse after allo-HSCT and would be beneficial to explore new methods of adoptive immunotherapy. Further research related to regulatory mechanism behind this phenomenon would be necessary.

Disclosures:
No relevant conflicts of interest to declare.