Abstract
We present a comparative study on 124 patients with hematologic malignancies who had undergone reduced-intensity conditioning and then received a transplant from an HLA-matched related (MRD), an HLA-matched unrelated (MUD), or an HLA-haploidentical related (HAPLO) donor. The conditioning regimen, which consisted of fludarabine, melphalan or busulfan, and alemtuzumab was administered to patients with lymphoid (n = 62) or myeloid disease (n = 62). Mycophenolate mofetil was used as prophylaxis for graft-versus-host disease (GVHD), and 38, 58, and 33 patients received transplants from MRD, MUD, and HAPLO donors, respectively. Only 2 patients experienced primary graft failure (GF) after melphalan-based regimen, whereas 8 of the 17 patients who received a transplant from HAPLO donors experienced a primary GF after busulfan-based regimen. The cumulative incidence of grade III to IV acute GVHD in engrafted patients who had received transplants from MRD, MUD, or HAPLO donors was 3%, 11%, and 27%, respectively, and the 2-year overall survival (OS) rates were 51%, 22%, and 23%, respectively. According to multivariate analysis, transplantation from either MUD or HAPLO donors compared with MRD were adverse factors that affected the OS (P = .006 and P = .002, respectively). In conclusion, the reduced-intensity regimen that included fludarabine, busulfan, or melphalan and alemtuzumab using only mycophenolate mofetil as the GVHD prophylaxis conferred favorable outcomes in the MRD group but lower survival rates in the MUD and HAPLO groups. The busulfan-based regimen led to a high incidence of GF in the HAPLO group, suggesting the need for modification or intensification of immunosuppression.
Keywords: Reduced-intensity, Alemtuzumab, Matched-related, Matched-unrelated, Haploidentical
INTRODUCTION
With the development of nonmyeloablative and reduced-intensity conditioning, allogeneic hematopoietic stem cell transplantation (SCT) can be offered to older and more infirm patients who have hematological malignancies and are at a high risk of relapse under conventional chemotherapy [1,2]. Our group and others previously showed that facilitation of engraftment and reduced incidence of graft-versus-host disease (GVHD) and treatment-related mortality (TRM) were achieved in allogeneic SCT from HLA-matched sibling donor or unrelated donor by using alemtuzumab: a humanized monoclonal antibody directed against human CD52 that is expressed on many T and B cells and some dendritic and NK cells [3–6]. We expanded this strategy further to include haploidentical SCT [7]. The combined use of alemtuzumab (100 mg), fludarabine, and cyclophosphamide as a conditioning of haploidentical transplantation resulted in low cumulative incidence of acute GVHD, low TRM, and an acceptable incidence of primary graft failure (GF). However, the high incidences of infection and relapse due to slow immune reconstitution have been significant obstacles to these strategies.
In the present prospective study, we reduced the dose of alemtuzumab from 100 mg to 80 mg and replaced cyclophosphamide with either busulfan or melphalan for myeloid and lymphoid malignancies, respectively, to decrease incidence of infectious complications and increase antitumor activity in reduced-intensity peripheral blood stem cell transplantation from HLA-matched related (MRD), HLA-matched unrelated (MUD), and HLA-haploidentical related (HAPLO) donors.
METHODS
Patient and Donor Selection
The patient inclusion criteria were as follows: (1) patients at least 18 years of age; (2) patients not considered candidates for myeloablative allogeneic SCT as a result of having other comorbidities, known recent Aspergillus infection, advanced age, or prior high-dose therapy; (3) patients who have pulmonary function test with single-breath diffusing capacity at least 40% of the predicted value, cardiac ejection fraction at least 40%, and Eastern Cooperative Oncology Group performance status of 2 or less; and (4) fulfillment of the disease status described below. For the lymphoid cohort, the target patient population exhibited a high likelihood for progressive lymphoid or myelomatous disease: (1) acute lymphoid leukemia with no more than 3 hematological remissions, (2) relapsed Hodgkin or non-Hodgkin lymphoma that are chemosensitive to salvage chemotherapy, and (3) myeloma or myelomatous disease that had persisted or progressed after the use of at least 1 regimen. For the myeloid cohort, the target patient population exhibited a high likelihood of progressive myeloid disease or myeloproliferative disease (MPD): (1) myeloid leukemia with no more than 3 hematological remissions, (2) myelodysplastic syndrome (MDS) with a history of at least intermediate-1 risk according to the International Prognostic Scoring System criteria, and (3) MPD.
The donor selection algorithm included a 5/6 to 6/6 matched sibling as the first choice, an available matched unrelated donor as the second choice, or a 3/6 to 5/6 partially matched family member (if 5/6, the donor is not a sibling, which would be first choice) as the third choice. The protocol was approved by the institutional review board of the Duke University School of Medicine. Written informed consent was obtained from all patients and donors. This protocol was registered at ClinicalTrials.gov (NCT00597714).
Treatment Plan
The conditioning regimen used for myeloid disease consisted of fludarabine (40 mg/m2/day) infused over a period of 30 minutes on days −5 through −2; busulfan (130 mg/m2/day) infused over a period of 3 hours on days −3 through −2; and alemtuzumab (20 mg/day) infused over a period of 3 hours on days −4 through −1. The conditioning regimen used for lymphoid diseases consisted of fludarabine (40 mg/m2/day) infused over a period of 30 minutes on days −5 through −2; melphalan (140 mg/m2/day) infused over a period of 15 minutes on day −2; and alemtuzumab (20 mg/day) infused over a period of 3 hours on days −4 through −1. Peripheral blood stem cells were mobilized from related or unrelated donors. The target goals for related or unrelated donor harvest were 15 to 20 × 106 and 5 × 106 CD34+ cells/kg, respectively. GVHD prophylaxis consisted of mycophenolate mofetil (1000 mg) administered orally or intravenously twice daily beginning on day −2 and continuing until day +60 post transplantation. Granulocyte colony-stimulating factor was not used routinely, except in patients who showed no signs of hematopoietic recovery. Of patients who had received transplants from MRD, 3 received subsequent unmanipulated donor lymphocyte infusion (DLI) and 14 received NK cell–enriched DLI infusions. Of patients who received transplants from MUD, 1 received DLI, and of patients who had received transplants from HAPLO donors, 2 received DLI and 2 received NK cell–enriched DLI infusions. T/NK DLIs were mostly given as planned on other post-transplantation protocols, except for 1 given for mixed chimerism and another for relapsed disease.
Assessment of Engraftment and Toxicity
Bone marrow aspiration and/or biopsy were performed 3 to 5 weeks after transplantation to assess donor-cell engraftment or determine the cause for delayed neutrophil recovery. Samples of bone marrow or peripheral blood were used to assess donor cell chimerism. Recipient and donor chimerism was determined by PCR amplification and subsequent size comparison of multiple short tandem repeats. Primary GF was defined as a neutrophil count below 500/μL or the absence of donor-derived hematopoiesis (<5% donor cells) before relapse, death, or retransplantation [8]. Secondary GF was defined as the achievement of primary engraftment and a subsequent decrease in neutrophils to 3 consecutive counts of less than 100/μL or the absence of donor-derived hematopoiesis (<5% donor cells) before relapse, death, or retransplantation. Neutrophil engraftment was defined as the achievement of a neutrophil count of at least 500/μL on or before day 50 post transplantation. Acute or chronic GVHD was diagnosed and graded according to standard criteria [9,10]. Toxicity was formally graded using the National Cancer Institute Common Toxicity Criteria (version 3.0; http://ctep.cancer.gov/protocolDevelopment/electronic_applications/docs/ctcaev3.pdf).
Measurement of Immune Recovery
Quantification of CD3+, CD4+, and CD8+ T cells was performed by flow cytometry on fresh peripheral blood at approximately 1 month before transplantation and then 1.5, 3, 6, and 12 months after transplantation.
Statistical Analysis
Descriptive statistics were used for summarizing variables related to patient demographics and transplantation characteristics. Comparisons among the groups were performed by use of the chi-square statistic for categorical variables and the Kruskal-Wallis test for continuous variables. The median ± standard error values of the percentages were used to describe the donor cell chimerism for the CD15+ and CD3 + cells of the bone marrow or peripheral blood. These chimerism data were available in 34 and 35 patients of the lymphoid and myeloid cohorts, respectively. Disease-free survival (DFS), progression-free survival (PFS), and overall survival (OS) rates after SCT were estimated using the Kaplan-Meier method. We performed univariate comparisons by using the log-rank test. DFS was defined as the period of time between the day of transplantation and either disease relapse or death due to the disease; DFS was measured in all patients who, based on all measureable criteria, had attained a complete response after transplantation. PFS was defined as the period of time between the day of transplantation and either the day underlying disease progression was documented or death occurred by any cause. OS was defined as the period of time between the day of transplantation and death. To eliminate the effects of competing risk, we assessed cumulative incidence by methods described elsewhere [11], and we conducted univariate comparisons using the Gray test. For the analysis of relapse and acute and chronic GVHD, we defined a competing event as death without an event of interest. For the analysis of TRM, relapse was defined as a competing risk. Acute GVHD was analyzed among the patients who achieved and maintained donor cell engraftment, and chronic GVHD was analyzed among the patients who achieved and maintained donor cell engraftment and survived more than 100 days. Cox proportional hazards regression was used to evaluate variables that potentially affected DFS, PFS, and OS. The following variables were considered as confounders: recipient age, recipient sex, disease (myeloid or lymphoid), disease risk (standard or high), and donor type (MRD, MUD, or HAPLO donor). The following were defined as standard-risk diseases: acute leukemia in complete remission (CR), chronic myelogenous leukemia in the chronic phase, non-Hodgkin and Hodgkin lymphoma in CR, MDS or MPD in CR or untreated, and multiple myeloma in CR. Other diseases were defined as high-risk diseases. Donor cell chimerism was compared by performing Student t-test. The immune recovery rates of the CD3+, CD4+, and CD8+ T cells in the MRD, MUD, and HAPLO groups were compared by performing an analysis of variance at each time point after transplantation. All tests were 2 sided and a P value less than .05 indicated statistical significance. All statistical analyses were performed using Stata version 12 (Stata Corp.; College Station, TX, USA).
RESULTS
Patients and Graft Characteristics
A total of 124 patients with a median follow-up of 26 months (among surviving patients) were included in this study (Table 1). The lymphoid cohort included 62 patients who had acute lymphoid leukemia (n = 5), chronic lymphocytic leukemia (n = 12), lymphoma (n = 40), or myeloma (n = 5). The myeloid cohort included 62 patients with acute myelogenous leukemia/MDS (n = 56), chronic myelogenous leukemia (n = 3), or MPD (n = 3). Median ages of MRD (n = 38), MUD (n = 53), and HAPLO donors (n = 33) were 57 (range, 20 to 69), 59 (range, 22 to 73), and 55 (range, 23 to 70), respectively. The CD34 counts of peripheral blood stem cells for MRD, MUD, and HAPLO donors were 13.1 × 106 /kg (range, 1.9 to 25.5 × 107/kg), 7.9 × 106/kg (range, 2.7 to 21.0 × 107/kg), and 10.9 × 106/kg (range, 1.8 to 20.8 × 107/kg), respectively.
Table 1.
Patient Characteristics
| Characteristics | MRD | MUD | HAPLO | P Value |
|---|---|---|---|---|
| No. of patients | 38 | 53 | 33 | |
| Age, median (range), yr | 57 (20–69) | 59 (22–73) | 55 (23–70) | .10 |
| Sex | ||||
| Female | 20 (53%) | 22 (42%) | 12 (36%) | .36 |
| Male | 18 (47%) | 31 (58%) | 21 (64%) | |
| Diagnosis | ||||
| Lymphoid disease | 21 (55%) | 25 (47%) | 16 (48%) | |
| ALL | 2 | 1 | 2 | |
| CLL | 3 | 4 | 5 | |
| ML | 13 | 18 | 9 | |
| MM | 3 | 2 | 0 | .73 |
| Myeloid disease | 17 (45%) | 28 (53%) | 17 (52%) | |
| AML/MDS | 17 | 25 | 14 | |
| CML | 0 | 2 | 1 | |
| MPD | 0 | 1 | 2 | |
| Disease status | ||||
| Standard | 28 (74%) | 36 (68%) | 23 (70%) | .84 |
| High | 10 (26%) | 17 (32%) | 10 (30%) | |
| HLA matching, A,B,DR | ||||
| 3/6 | – | – | 21 (64%) | |
| 4/6 | – | – | 9 (27%) | – |
| 5/6 | – | – | 3 (9%) | |
| 6/6 | 38 (100%) | 53 (100%) | – | |
| KPS | ||||
| <90 | 18 (47%) | 23 (43%) | 12 (36%) | |
| 90–100 | 20 (53%) | 30 (57%) | 20 (61%) | .48 |
| Missing | 0 | 0 | 1 (3%) | |
| HCT-CI | ||||
| 0 | 10 (26%) | 8 (15%) | 8 (24%) | |
| 1–2 | 9 (24%) | 13 (25%) | 3 (9%) | .27 |
| >3 | 19 (50%) | 32 (60%) | 22 (67%) | |
| Prior history of autologous transplantation | ||||
| Yes | 6 (16%) | 10 (19%) | 3 (9%) | .47 |
| No | 32 (84%) | 43 (81%) | 30 (91%) | |
| CD34+ cells/kg, × 106, median (range) | 13.1 (1.9–25.5) | 7.9 (2.7–21.0) | 10.9 (1.8–20.8) | <.01 |
| Follow-up duration among survivors, median (range), mo | 28.6 (3.4–44.8) | 23.7 (2.7–40.0) | 26.3 (4.6–38.9) | .67 |
KPS indicates Karnofsky Performance Status; HCT-CI, hematopoietic cell transplantation-comorbidity index; MRD, HLA-matched related donor; MUD, HLA-matched unrelated donor; HAPLO, HLA-haploidentical related donor; ALL, acute lymphoblastic leukemia; CLL, chronic lymphocytic leukemia; ML, malignant lymphoma; MM, multiple myeloma; AML, acute myelogenous leukemia; MDS, myelodysplastic syndrome; CML, chronic myelogenous leukemia; MPD, myeloproliferative disease.
Data presented are n (%) unless otherwise indicated.
Engraftment
For the lymphoid cohort, all 21 patients who had received a SCT from a MRD engrafted successfully, whereas 1 of the 25 patients who had received a SCT from a MUD experienced secondary GF with complete donor chimerism that did not respond to CD34-selected DLI. Among the 16 HAPLO patients, 2 exhibited primary GF; both underwent retransplantation from the same donor and 1 was rescued. The chimerism analysis at 1 month after SCT showed a mean donor chimerism of 94.4% ± 2.6% for the CD15+ cells and 88.9% ± 3.0% for the CD3 + cells. In the myeloid cohort, 1 of the 17 patients who had received a transplant from a MRD experienced a primary GF, which was caused by cytomegalovirus (CMV) reactivation most likely, and was rescued by retransplantation from the same donor. Of the 28 patients who had received a SCT from MUD, 2 experienced a primary GF and 4 experienced a secondary GF. Four were rescued by a subsequent SCT from the same MUD or a new HAPLO donor. Eight of the 17 patients who had received transplants from HAPLO donors experienced primary GF; 3 of the 8 underwent retransplantation from the same or another HAPLO donor, and 1 of the 8 received an autologous transplantation—all 4 of these patients were rescued. One of the remaining 4 patients received a DLI and a CD34 boost, but the graft failed. One patient experienced secondary GF. The chimerism analysis at 1 month after SCT showed a mean donor chimerism of 91.6% ± 3.2% for CD15+ cells and 69.7% ± 6.0% for CD3 + cells. We used the combination of fludarabine, cyclophosphamide, and alemtuzumab as a salvage-conditioning regimen for GF in most cases [12]. We also analyzed chimerism data at 1 month after SCT for both malignancies focusing on the MUD and HAPLO group. A mean donor chimerism of patients with and without engraftment was 96.3% ± 1.0% and 78.6% ± 12.5% (P = .003) for CD15+ cells and 89.6% ± 3.2% and 39.6% ± 16.4% (P < .001) for CD3 + cells. The median time period in days for neutrophil engraftment after MRD, MUD, or HAPLO transplantation was 19 (range, 13 to 27), 19 (range, 11 to 29), and 16.5 (range, 13 to 22) (P = .06), respectively.
GVHD
Among the engrafted patients who had received a transplant from a MRD, MUD, or HAPLO donor, the cumulative incidence of grades II to IV acute GVHD on day 180 post transplantation was 16% (95% confidence interval [CI], 7% to 30%), 26% (95% CI, 14% to 39%), and 46% (95% CI, 24% to 65%), respectively (Gray test, P = .044); and the cumulative incidence of grades III to IV acute GVHD on day 180 was 3% (95% CI, 0.2 to 12%), 13% (95% CI, 5% to 25%), and 27% (95% CI, 11% to 47%), respectively (Gray test, P = .016) (Figure 1). Among the engrafted patients who survived more than 100 days, the incidence of chronic GVHD 2 years after transplantation from MRD, MUD, or HAPLO donors was 22% (95% CI, 9% to 37%), 35% (95% CI, 20% to 51%), and 39% (95% CI, 17% to 61%), respectively (Gray test, P = .166) (Figure 1).
Figure 1.
Cumulative incidence of acute and chronic GVHD. Black line shows HLA-matched related donor (MRD), dotted line shows HLA-matched unrelated donor, and gray line shows HLA-haploidentical related donor (HAPLO).
Infections
Table 2 shows the incidence of infection (Grades ≥ 2 according to National Cancer Institute Common Toxicity Criteria) according to donor type. In total, 55% of patients experienced CMV reactivation and 9% developed CMV disease. Other infectious complications included polyoma virus in 25% of patients, bacteria in 38%, Herpes simplex virus in 10%, varicella zoster virus in 6%, human herpesvirus 6 in 19%, respiratory syncytial and parainfluenza viruses in 13%, and fungi in 9%. The incidences of polyoma virus, fungal, and human herpesvirus 6 infection were low in the MRD group, whereas those of Herpes simplex virus and fungal infection were high in the HAPLO group.
Table 2.
Infections According to Donor Type
| Toxicity | MRD | MUD | HAPLO | Total |
|---|---|---|---|---|
| CMV reactivation | 24 (63%) | 26 (49%) | 18 (55%) | 68 (55%) |
| CMV disease | 2 (5%) | 6 (11%) | 3 (9%) | 11 (9%) |
| Bacteria | 12 (32%) | 22 (42%) | 13 (39%) | 47 (38%) |
| Clostridium difficile | 3 (8%) | 4 (8%) | 1 (3%) | 8 (6%) |
| EB virus | 1 (3%) | 0 | 0 | 1 (1%) |
| HSV | 3 (8%) | 3 (6%) | 7 (21%) | 13 (10%) |
| VZV | 1 (3%) | 6 (11%) | 1 (3%) | 8 (6%) |
| HHV-6 | 1 (3%) | 12 (23%) | 11 (33%) | 24 (19%) |
| AFB | 2 (5%) | 1 (2%) | 0 | 3 (2%) |
| Polyoma virus | 6 (16%) | 15 (28%) | 10 (30%) | 31 (25%) |
| RS virus | 3 (8%) | 3 (6%) | 2 (6%) | 8 (6%) |
| Parainfluenza | 4 (11%) | 2 (4%) | 3 (9%) | 9 (7%) |
| Fungus | 1 (3%) | 5 (9%) | 5 (15%) | 11 (9%) |
| Aspergillus | 1 (3%) | 4 (8%) | 0 | 5 (4%) |
| Candida species | 0 | 0 | 1 (3%) | 1 (1%) |
| Other fungus | 0 | 1 (2%) | 4 (12%) | 6 (5%) |
MRD indicates HLA-matched related donor; MUD, HLA-matched unrelated donor; HAPLO, HLA-haploidentical related donor; CMV, cytomegalovirus; EB, Epstein-Barr; HSV, herpes simplex virus; VZV, varicella zoster virus; HHV-6, human herpesvirus 6; AFB, acid-fast bacilli.
Data presented are n (%).
DFS, PFS, and OS
The 2-year DFS rate after SCT from MRD, MUD, and HAPLO donors was 47% (95% CI, 29% to 63%), 23% (95% CI, 12% to 36%), and 16% (95% CI, 5% to 33%), respectively (log-rank test, P = .021); the 2-year PFS rate was 43% (95% CI, 26% to 59%), 22% (95% CI, 11% to 34%), and 15% (95% CI, 5% to 30%), respectively (log-rank test, P = .042); and the corresponding 2-year OS rate was 51% (33% to 66%), 22% (12% to 35%), and 23% (10% to 39%), respectively (log-rank test, P = .004) (Figure 2). Multivariate analysis revealed that SCT from MUD or HAPLO donors in comparison to MRD was the only adverse factor that affected the OS rate (hazard ratio [HR] for MUD, 2.23 [95% CI, 1.26 to 3.93], P = .006; HR for HAPLO, 2.63 [1.42 to 4.87], P = .002) (Table 3). However, the OS rate after SCT from HAPLO donors did not differ significantly from the OS rate after SCT from MUD (P = .520). Other variables (recipient age, sex, disease, and disease risk) were not significantly associated with the outcome.
Figure 2.
Probability of disease-free survival, progression-free survival, and overall survival. Black line shows HLA-matched related donor (MRD), dotted line shows HLA-matched unrelated donor, and gray line shows HLA-haploidentical related donor (HAPLO).
Table 3.
Multivariate Analysis of Factors Potentially Affecting Overall Survival
| Variables | HR (95% CI) | P Value |
|---|---|---|
| Age (by 1 year) | 1.01 (.99–1.03) | .212 |
| Sex | ||
| Male | 1.00 | |
| Female | 1.20 (.77–1.87) | .419 |
| Diagnosis | ||
| Myeloid disease | 1.00 | |
| Lymphoid disease | .85 (.53–1.38) | .523 |
| Disease risk | ||
| Standard | 1.00 | |
| High | 1.28 (.79–2.10) | .317 |
| Donor type | ||
| MRD | 1.00 | |
| MUD | 2.23 (1.26–3.93) | .006 |
| HAPLO | 2.63 (1.42–4.87) | .002 |
MRD indicates HLA-matched related donor; MUD, HLA-matched unrelated donor; HAPLO, HLA-haploidentical related donor; CI, confidence interval; HR, hazard ratio.
Relapse and TRM
No significant differences in relapse incidence occurred among recipients who received SCT from MRD, MSD, or HAPLO donors (Gray test, P = .776). The relapse incidence 2 years after SCT was 28% (95% CI, 14 to 44%) for MRD, 28% (16 to 42%) for MUD, and 24% (10% to 41%) for HAPLO donors (Figure 3). On the other hand, nonrelapse mortality after SCT was significantly higher among recipients who received SCTs from MUD and HAPLO donors (Gray test, P = .018). The nonrelapse mortality 2 years after SCT was 29% (95% CI, 15% to 45%) for MRD, 50% (35% to 64%) for MUD, and 61% (41% to 76%) for HAPLO transplantation (Figure 3). The most common causes of death were progressive disease (27% of death by all causes) and infection (39%) (Table 4).
Figure 3.
Cumulative incidence of relapse and nonrelapse mortality. Black line shows HLA-matched related donor (MRD), dotted line shows HLA-matched unrelated donor, and gray line shows HLA-haploidentical related donor (HAPLO).
Table 4.
Causes of Death According to Donor Type
| Cause of Death | MRD (n = 38) |
MUD (n = 53) |
HAPLO (n = 33) |
Total |
|---|---|---|---|---|
| Progressive disease | 5 (28%) | 13 (33%) | 5 (19%) | 23 (27%) |
| Infections | 10 (56%) | 15 (38%) | 8 (31%) | 33 (39%) |
| Rejection | 0 | 0 | 3 (12%) | 3 (4%) |
| GVHD | 0 | 2 (5%) | 3 (12%) | 5 (6%) |
| Organ failure | 0 | 2 (5%) | 3 (12%) | 5 (6%) |
| PTLD | 1 (6%) | 0 | 1 (4%) | 2 (2%) |
| Others | 1 (6%) | 3 (8%) | 1 (4%) | 5 (6%) |
| Undetermined | 1 (6%) | 5 (13%) | 2 (8%) | 8 (10%) |
MRD indicates HLA-matched related donor; MUD, HLA-matched unrelated donor; HAPLO, HLA-haploidentical related donor; GVHD, graft-versus-host disease; PTLD, post-transplantation lymphoproliferative disorder.
Data presented are n (%).
Immune Recovery
The immune recovery data for the CD3+, CD4+, and CD8+ T cells are shown in Figure 4. The number of these T cells was very low at 1.5 months after transplantation and then gradually recovered through 12 months after transplantation. Although the median values of these T cells were lower in the MUD and HAPLO groups than in the MRD group, no significant difference was found among the 3 groups.
Figure 4.
Immune recovery of the (A) CD3+, (B) CD4+, and (C) CD8+ T cells. The median values before transplantation and 1.5, 3, 6, and 12 months after transplantation are shown with the standard error bars. Black line shows HLA-matched related donor (MRD), dotted line shows HLA-matched unrelated donor, and gray line shows HLA-haploidentical related donor (HAPLO).
DISCUSSION
There has been no standard conditioning regimen for nonmyeloablative or reduced-intensity transplantation, particularly from a HAPLO donor. Use of ex vivo CD34 selection or T cell depletion methods decrease incidence of severe acute GVHD, but render patients at high risk of infection and relapse, as well as graft failure. Therefore, we expanded our previous study of nonmyeloablative transplantation using fludarabine, melphalan or busulfan, a lower dose of alemtuzumab, and only mycophenolate mofetil as a GVHD prophylaxis. In this protocol, we observed favorable outcomes for the MRD group either in the myeloid or lymphoid cohort. However, high incidence GF in the myeloid cohort of the MUD and HAPLO groups was observed after busulfan-based regimen. This suggests that the use of busulfan and fludarabine in combination with alemtuzumab does not ensure donor engraftment, which indicates the need for additional immunosuppression. Further, the most common causes of death remain the same as our previous study: progressive disease and infection. Although we reduced the dose of alemtuzumab and used mycophenolate mofetil as a single GVHD prophylaxis agent without increasing the occurrence of severe acute GVHD, the reduction in the alemtuzumab dose from 100 mg to 80 mg may be inadequate for reducing infection and relapse rates and actually did not improve T cell recovery in comparison with our previous study [7]. Chakraverty et al. [4] tested a reduced-intensity conditioning regimen with dose deescalated alemtuzumab after HLA-identical sibling SCT using cyclosporine as GVHD prophylaxis. They reported that a 20-mg dose of alemtuzumab was associated with a greater risk of severe GVHD. In contrast, the 30-mg dose was associated with comparable clinical outcomes and improved lymphocyte recovery compared to the 40- or 60-mg dose groups that had acceptable levels of severe GVHD incidence. This study suggests that the alemtuzumab dose can be reduced to 30 mg on day −1 before transplantation from an HLA-identical sibling if cyclosporine is used as the single GVHD prophylactic agent. Therefore, the dose of alemtuzumab may be reduced to 30 mg after the MRD transplantation under a less intensified post-transplantation GVHD prophylaxis. If a conventional GVHD prophylaxis, such as calcineurin inhibitors with methotrexate or mycophenolate mofetil is used, the dose of alemtuzumab may be further reduced, or the administration may be discontinued, at least after the MRD transplantation. However, because we observed an incidence of grade III to IV acute GVHD in 27% after HAPLO transplantation, the dose of alemtuzumab should be reduced moderately, perhaps to 60 mg or less, for HAPLO transplantation.
A high incidence of GF was noted in the myeloid cohort, who were conditioned with a fludarabine, busulfan, and alemtuzumab regimen and who received mycophenolate mofetil as GVHD prophylaxis. The chimerism analysis showed a lower donor chimerism of CD3 + cells in the busulfan-based regimen than in the melphalan-based regimen (69.7% versus 88.9%). Further, low donor chimerism of CD3 + cells was associated with graft failure. This finding suggests that these combinations were inadequate for the suppression of recipient T cells. We previously reported a high incidence of GF after single unrelated cord blood transplantations that used a conditioning regimen of fludarabine and busulfan (16 mg/kg) [13]. Based on these findings, the combined use of fludarabine and busulfan inadequately ensures successful donor cell engraftment particularly after cord or haploidentical transplantation. In our previous study using fludarabine, cyclophosphamide, and alemtuzumab (100 mg), primary GF was seen in only 6% of recipients after HAPLO transplantation [7]. The addition of an alkylating agent (eg, melphalan or cyclophosphamide) instead of busulfan and/or the addition of 2 to 4 Gy total body irradiation will be needed to ensure engraftment following transplantation from a MUD and HAPLO donor. In addition, donor-specific HLA antibodies should be screened to further decrease the risk of GF after HAPLO transplantation [14,15]. Relatively low doses of CD34 cells in the MUD group compared with the MRD group may have affected the GF rate of the MUD group. However, it is practically difficult to collect more CD34 cells from unrelated donors to ensure donor cell engraftment. Another strategy to enhance donor cell engraftment is increased intensity of GVHD prophylaxis. Ogawa et al. [16] reported a high incidence of donor cell engraftment after treatment with fludarabine, busulfan, and anti T-lymphocyte globulin, as well as GVHD prophylaxis that consisted of tacrolimus and methylprednisolone. However, because the high incidence of infection remains a problem in the protocol, these prophylaxes may not improve the OS rate. Although the high GF rate was observed in the myeloid cohort, disease itself (myeloid versus lymphoid disease) was not associated with other outcomes, such as acute and chronic GVHD, relapse, TRM, and OS in the multivariate analysis (data not shown). This suggests that choice of donor source rather than treatment regimen had a strong effect on other outcomes.
Recently, the use of post-transplantation cyclophosphamide was demonstrated to be effective for reducing the incidence of severe acute GVHD and TRM in haploidentical transplantation [17,18]. Brunstein et al. [18] performed a phase II trial of haploidentical transplantation using post-transplantation cyclophosphamide and reported 1-year cumulative incidence rates of 7% and 45% for nonrelapse mortality and relapse, respectively. The overall and PFS rates at 1 year were 62% and 48%, respectively. Bashey et al. [19] retrospectively compared the outcomes of HAPLO transplantation with the use of post-transplantation cyclophosphamide and MRD and MUD transplantations. They reported comparative results among the MRD, MUD, and HAPLO groups. Compared with their approach, our approach resulted in higher rates of TRM for the MUD and HAPLO groups and higher rates of severe acute GVHD for the HAPLO group, although the relapse rate was relatively lower. The higher TRM in the MUD and HAPLO groups was partly due to the high GF rates after the busulfan-based regimen, the GVHD-associated complications, and delay in the immune recovery after transplantation. These results suggest that effective GVHD suppression without inhibiting graft-versus-leukemia effects or delaying immune recovery remains a major issue in our approach. We need an enhanced strategy to boost immune recovery without increasing the risk of severe GVHD and TRM by reducing the dose of alemtuzumab, using additional immunosuppression with cyclophosphamide or other agents, and discontinuation of the use of busulfan as a conditioning regimen in this approach. However, the results of this study should be cautiously interpreted because of the relatively small sample size and heterogeneity of the underlying diseases.
In conclusion, the reduced-intensity regimen that included fludarabine, busulfan or melphalan, and alemtuzumab (80 mg), using only mycophenolate mofetil as the GVHD prophylaxis, conferred favorable outcomes in the MRD group but lower survival rates in the MUD and HAPLO groups. The busulfan-based regimen led to a high incidence of GF in the MUD and HAPLO groups, suggesting the need for modification or intensification of immunosuppression of the conditioning regimen. The high incidences of disease recurrence and infection suggest that the development of strategies to improve immune recovery remains a challenge.
Acknowledgments
We thank the nurse practitioners, physician’s assistants, ward and clinic nurses, and staff of the Duke Adult Stem Cell Transplant Program for their outstanding care of the patients described in this report.
Financial disclosure statement: This work was supported in part by the National Cancer Institute (NIH) P01-CA047741-19 (N. J. C.).
Footnotes
Conflict of interest statement: There are no conflicts of interest to report.
References
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