Introduction
The prognosis of acute leukemia or high-grade lymphoma in relapsed/refractory status is dismal [
1,
2]. More recently, gene mutations, such as
TP53,
DNA-methyltransferase-3a (
DNMT3a) or
ten-eleven translocation-2 (
TET2) mutation, have been identified by the next-generation sequencing technique as very high-risk molecular markers for acute leukemia with low rate of remission and short survival [
3‐
5]. Though the targeted therapy developed in recent years have resulted in an increased rate of remission and improved survival in a small subset of these very high-risk patients, allogeneic hematopoietic stem cell transplantation (allo-HCT) is still the most effective way for cures. However, the recurrence of the underlying disease after transplantation remains the leading cause of treatment failure. The rate of relapse of the leukemia/lymphoma that was in refractory/relapsed status prior to transplantation and those with very high-risk gene mutations ranged from 50 to 60% and the long-term relapse-free survival (RFS) was less than 30% after allo-HCT [
6,
7].
A reasonable approach to improve the survival of the very high-risk leukemia/lymphoma is to explore prophylactic strategies after transplantation to reduce the relapse rate. Donor lymphocyte infusion (DLI) has been proved to be effective to stimulate graft-versus-leukemia (GVL) reaction in patients with minimal residual disease (MRD) or hematologic relapse after transplantation [
8,
9]. However, the utility of DLI is limited by the toxicity of fatal GVHD or pancytopenia and subsequent infections resulting in an increase in non-relapse mortality (NRM). We have shown that using granulocyte colony-stimulating factor (G-CSF)-primed peripheral blood cells (G-PB) for DLI with substitution of the steady-state lymphocytes could reduce the DLI-associated fatal GVHD without counteracting its GVL effect [
10]. Reduced GVHD with improved RFS has been established in the unmanipulated haploidentical-HCT with bone marrow and peripheral blood (BM + PB) as grafts for therapeutic, preemptive and prophylactic use of DLI [
11‐
13]. However, all of these studies were in the setting of G-CSF-primed BM + PB as the graft source. It is well known that the components of the graft have an influence on the development of GVHD. In general, the peripheral blood stem cell transplantation (PBSCT) could be associated with a higher incidence of GVHD compared with allo-HCT with BM as grafts. The safety and efficacy of prophylactic G-PB DLI in the setting of unmanipulated haploidentical PBSCT (haplo-PBSCT) have not been determined. In this study, we used prophylactic G-PB DLI for the very high-risk leukemia/lymphoma patients after haplo-PBSCT with modifications intending to alleviate DLI-associated GVHD. The modifications included that the dose of infused CD3
+ cells was reduced to less than 2 × 10
7/kg (BM + PB setting, 4 × 10
7/kg), and the time interval between haplo-PBSCT and DLI was postponed to 60~90 days (BM + PB setting, 45~60 days). The data of tolerance and efficacy of prophylactic G-PB DLI in 31 patients with very high-risk features was presented.
Discussion
The recurrence of disease is the primary cause of treatment failure and mortality after allo-HCT for patients with very high-risk hematologic malignancies. Prophylactic DLI has been proved effective for treatment, intervention, or prophylaxis of relapse when targeted therapy is lacking post-transplantation. Here, we showed the feasibility of G-PB DLI in the unmanipulated haplo-PBSCT setting with G-CSF primed PB as the graft source for prophylaxis of relapse in the very high-risk patients.
In previous study of transplantation with BM or with BM combined with PBSC as graft, we have shown that the incidence of severe GVHD after G-PB DLI was less compared with that after traditional DLI with steady-state lymphocytes [
19]. In subsequent series of studies, the safety and efficacy of G-PB DLI were demonstrated in the treatment of relapse, preemptive therapy for MRD-positive patients or prophylaxis before relapse occurs after unmanipulated haploidentical HCT with PB + BM as grafts. It was reported that in 56 patients (29 after haplo-HCT) who were MRD-positive and received G-PB DLI, the incidence of acute GVHD grades 2–4 was 27.9% and that of chronic GVHD was 42.9% which were similar to those in the MRD negative patients (30.2 and 38.8%) who did not receive the intervention [
20]. The use of G-PB DLI was based on the laboratory findings that G-CSF-priming could induce hypo-responsiveness of T cells for polarization from Th1 to Th2 and downregulation of the CD28/B7 pathway [
10,
21] and augment NK-T cell dependent CD8
+ cytotoxicity which might enhance GVL without GVHD [
22]. Further, there was no less activity with G-CSF-primed as compared to untreated T cells [
23]. Nevertheless, the evidence for G-CSF-priming retaining GVL activity is sparse. In the current transplantation setting without in vitro T cell depletion, there is little evidence that NK cells could replace the hindered T cells. Therefore, the delay of DLI for 60 days might be the most important reason for its tolerance, as the cytokine storm caused by myeloablative conditioning was over.
It has been shown that G-PB DLI with immunosupressants prophylaxis more than 6 weeks were associated with a lower incidence of acute GVHD grades 3–4. Further, DLI-associated acute GVHD grades 3–4 was the only risk factor for OS and NRM but not for relapse after DLI [
11]. Therefore, in the current study, CsA was reduced and discontinued after the 6-week prophylaxis if no GVHD occurred. PBSCT with HLA-identical sibling donors was considered to be associated with an increased incidence of chronic GVHD compared with allo-HCT with BM [
24]. Therefore, we postponed the timing for DLI and reduced the dose of infused CD3+ cells concerning the potential higher incidence of GVHD in the haplo-PBSCT setting. The incidences of DLI-associated acute GVHD grades 2–4, 3–4, chronic GVHD, and NRM in our study were 55.3%, 10.2%, 52.0,%, and 33.1%, respectively. In the patients who received haplo-PBSCT in our unit before January of 2015 [
14], the incidences of acute GVHD grades 2–4, 3–4, chronic GVHD, and NRM were 36.1%, 14.5%, 38.4%, and 24.0%, respectively. Though the incidences of GVHD between these two cohort studies should not be compared directly, it seemed that G-PB DLI did not result in an intolerable toxicity in terms of GVHD and NRM. In addition, the tolerance of this procedure might be related to the following mastery of contraindications for prophylactic DLI: (1) the elimination of the patients with intermittent GVHD and (2) all prophylactic DLI was given after stable response of treatment of the previous GVHD.
An optimal timing of prophylactic DLI should be in a balance between GVL and GVHD because increasing the time interval between transplantation and DLI will lead to a decrease in the risk of DLI-associated toxicity but an increase in the likelihood of relapse. Because the median time to post-allo-HCT relapse or progression was 2 to 3 months for adult patients with high-risk acute myeloid leukemia and T cell leukemia/lymphoma [
25,
26], it is reasonable to administrate the prophylactic strategy before day + 90 after HCT. The median time of the occurrence of acute GVHD in the unmanipulated haplo-PBSCT was + 30~ + 60 days after graft infusion. That means the prophylactic DLI candidates would face superimposed risk of GVHD if the time of DLI is before day + 60. Even though several patients, at the beginning stage of the current study, had received DLI before day + 60, it is reasonable and feasible to give the prophylactic DLI after day + 60 if no intermittent GVHD occurred or GVHD was stably controlled.
Considering the different kinetics and sensitivity of GVL response in hematologic malignancies enrolled in this study, the relapse and survival could not have been compared with other reports. In the current study, a total of six patients with a disease in NR status prior to transplantation relapsed at a median of 114 days after DLI, and the statistical analysis revealed that disease in NR status prior to transplantation was an independent risk factor for relapse after DLI. Reasons for the treatment failure might be associated with the poor GVL effect of DLI. The high proliferative kinetics of leukemia cells is one cause for poor GVL effect of DLI. Nevertheless, the immune evasion of leukemia cells by mechanisms such as loss of the patient-specific HLA haplotype represents another cause for poor GVL effect of DLI [
27]. Therefore, it is likely the future of DLI should involve more strategies to enhance the GVL effect of the infused donor cells via using cytokines like interferon [
28], selection or depletion of specific donor lymphocytes subsets [
29,
30], genetically modified donor lymphocytes targeting of tumor-specific antigens, second prophylactic DLI in case of no GVHD occurrence, or combination with target therapy.
In summary, the data from this study suggested the tolerance and efficacy of prophylactic DLI in patients with very high-risk leukemia/lymphoma in the unmanipulated haplo-PBSCT setting with PB as grafts. Further study is required to determine kinetics of relapse in each subtype of high-risk malignancies and the optimal long-term prophylactic strategy.
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