Outcomes of haploidentical bone marrow transplantation in patients with severe aplastic anemia-II that progressed from non-severe acquired aplastic anemia

Abstract

Severe aplastic anemia II (SAA-II) progresses from non-severe aplastic anemia (NSAA). The unavailability of efficacious treatment has prompted the need for haploidentical bone marrow transplantation (haplo-BMT) in patients lacking a human leukocyte antigen (HLA)-matched donor. This study aimed to investigate the efficacy of haplo-BMT for patients with SAA-II. Twenty-two patients were included and followed up, and FLU/BU/CY/ATG was used as conditioning regimen. Among these patients, 21 were successfully engrafted, 19 of whom survived after haplo-BMT. Four patients experienced grade II–IV aGvHD, including two with grade III–IV aGvHD. Six patients experienced chronic GvHD, among whom four were mild and two were moderate. Twelve patients experienced infections during BMT. One was diagnosed with post-transplant lymphoproliferative disorder and one with probable EBV disease, and both recovered after rituximab infusion. Haplo-BMT achieved 3-year overall survival and disease-free survival rate of 86.4%±0.73% after a median follow-up of 42 months, indicating its effectiveness as a salvage therapy. These promising outcomes may support haplo-BMT as an alternative treatment strategy for patients with SAA-II lacking HLA-matched donors.

Keywords

severe aplastic anemia / non-severe acquired aplastic anemia / haploidentical bone marrow transplantation / outcomes

1 Introduction

Acquired severe aplastic anemia (SAA) is a rare, life-threatening, bone marrow failure syndrome characterized by peripheral pancytopenia and hypoplasia of the bone marrow [1,2]. In this immune-mediated destruction of hematopoietic cells, an attack by immunologically dysfunctional T cells depletes the hematopoietic stem cell (HSC) pool, impairs its compartments [3], and eventually leads to the deficient development of multi-lineage blood cells [4].

Acquired aplastic anemia (AA) can be classified as SAA or non-severe AA (NSAA) [5], the former is most frequently newly diagnosed in pediatric patients. Advances in allogeneic bone marrow transplantation (BMT), immunosuppressive therapy (IST), and supportive care over the past two decades have markedly improved the overall survival (OS) of patients with SAA [6]. As an initial therapy, allogeneic BMT or IST has been associated with long-term OS in patients with SAA [7]. For example, treatment with combinations of methylprednisolone, anti-thymocyte globulin (ATG), and/or cyclosporine has resulted in remission rate (complete remission+ partial remission) of 70% at 6 months and an OS rate of 80% at 41 months [8,9]. In addition, BMT with HLA-matched donors has resulted in a 3-year OS of 87%–92% [7,10]. By contrast, most pediatric patients with newly diagnosed SAA experience unfavorable outcomes with supportive care alone—without IST or BMT—resulting in a 2-year mortality rate near 80%. However, 9.2%–67% of pediatric and 18.8% of adult patients with NSAA who received supportive care or cyclosporine and/or androgen agents finally progressed to meet the criteria for SAA (termed as “SAA-II”) at a median of 26.5-66 months after diagnosis [11–14]. Patients with SAA-II are characterized by “empty” marrow replaced by fat cells, leading to red blood cell (RBC) and/or platelet transfusion dependency. Therefore, they have high serum ferritin levels in addition to low performance status and are vulnerable to secondary infections due to neutropenia during disease progression. Owing to the longer interval from diagnosis to BMT and their substantially older age than patients with newly diagnosed SAA, patients with SAA-II usually experience poorer outcomes.

Most patients who fulfill the criteria for SAA do not survive for more than 1 year without IST or BMT [15]. Moreover, age≥40 years and>3-month interval from diagnosis to BMT have been associated with high mortality risk in patients with SAA [10]. Haploidentical BMT (haplo-BMT) has yielded promising results in pediatric patients newly diagnosed with SAA with a 3-year OS rate of 84.5% [16]. Therefore, this strategy may be effective in patients with SAA-II who lack HLA-matched siblings and unrelated HLA-matched donors, although its specific efficacy and safety remain unclear. This study aimed to investigate the efficacy of haplo-BMT for patients with SAA-II that progressed from NSAA.

2 Patients and methods

2.1 Patients

Twenty-two patients with SAA-II who underwent haplo-BMT at the Second Hospital of Dalian Medical University and Air Force General Hospital of the Chinese People’s Liberation Army (Beijing, China) between February 1, 2015 and May 31, 2018 were recruited and followed up until December 31, 2019. The treatment protocol was approved by the institutional review boards of participating hospitals.

2.2 Criteria for SAA

AA severity was based on the Camitta criteria [1,17]. Individuals with NSAA exhibit hypocellular marrow with cytopenia in the peripheral blood and thus do not fulfill the criteria for SAA. SAA-II is defined as marrow hypocellularity with pancytopenia meeting the criteria for SAA and progressing from NSAA to SAA. SAA is defined as bone marrow hypoplasia (cellularity<25%) and at least two of the following peripheral blood abnormalities: hemoglobin (HB) level<90 g/L, corrected reticulocyte count<1%; neutrophil count<0.5 × 10⁹/L, or platelet count<30 × 10⁹/L. Very severe AA (vSAA) was defined as HB concentration<80 g/L, reticulocyte count<20 × 10⁹/L, neutrophil count<0.2 × 10⁹/L, and platelet count<20 × 10⁹/L. Patients were excluded if they were diagnosed with other conditions that can cause pancytopenia prior to SAA diagnosis, including congenital Fanconi anemia, myelodysplastic syndrome, paroxysmal nocturnal hemoglobinuria, leukemia, or metastasis of solid tumor(s) into the bone marrow

2.3 Criteria for RBC and platelet transfusion dependence

RBC transfusion dependence is defined as a requirement for≥2 U of RBCs over a 28-day period and an HB level<85 g/L over a 12-week period according to the International Working Group for Myelofibrosis Research and Treatment consensus criteria for RBC transfusion dependence in patients with myeloproliferative neoplasm-associated myelofibrosis [18,19]. Platelet transfusion dependence is defined as a requirement for≥1 U platelets every 8 weeks over a period of 4 months [20].

2.4 Conditioning regimen

FLU/BU/CY/ATG conditioning regimen is a modification of the previous BU/CY/ATG regimen used in haplo-BMT for SAA [16] with fludarabine added to enhance immunosuppression. Fludarabine (FLU) (30 mg/m²) was administered intravenously once a day on days -10, -9, -8, and -7; 0.8 mg/kg busulfan (BU) was given four times a day on days -7 and -6; 50 mg/kg cyclophosphamide (CY) once a day on days -6, -5, -4, and -3; and 2.5 mg/kg rabbit ATG (SangStat, Lyon, France) once a day on days -5, -4, -3, and -2. The conditioning regimen FLU/CY/ATG for the second haplo-BMT consisted of 30 mg/m² FLU once a day on days -7, -6, -5, and -4; 50 mg/kg CY once a day on days -4, -3, and -2; and 2.5 mg/kg of ATG once a day on days -2 and -1.

2.5 Granulocyte colony stimulating factor-primed stem cell mobilization and harvesting

All healthy donors were subcutaneously injected with 5 mg/kg granulocyte colony stimulating factor (G-CSF; Kirin, Japan) once a day for 5 days. Bone marrow grafts containing target stem cell counts (1×10⁶/kg−3×10⁶/kg recipient body weight) were harvested on day 0, and peripheral blood stem cells were harvested by apheresis using a Spectra Optia Cell Separator (Terumo BCT, Lakewood, CO, USA) on the following days until the harvest of HSC counts≥3 × 10⁶/kg body weight or mononuclear cells counts≥5 × 10⁸/kg body weight of the recipient.

2.6 Prophylaxis and management for GvHD

aGvHD and cGvHD were diagnosed and graded based on MAGIC criteria (the Mount Sinai Acute GvHD International Consortium) [21] and National Institute of Health Severity score diagnostic criteria (NIH 2014 criteria) [22], respectively. GvHD prophylaxis consisted of intravenous tacrolimus (FK506, 0.03 mg/kg/day) and oral mycophenolate mofetil (MMF, 1.0 g/day) beginning on day -9 and intravenous methotrexate (MTX) at doses of 15 mg/m² on day+1 and 10 mg/m² on days+3, +6, and+11. FK506 blood levels were maintained at 10-20 ng/mL for nearly 3 weeks until the recipient’s bowel function recovered normal status. Thereafter, intravenous FK506 was replaced by FK506 tablets until 12 months after BMT and was discontinued at 15 months. MMF was tapered, beginning four weeks after HSCT, and was discontinued at 8 weeks. Patients who developed aGvHD were treated with 1-2 mg/kg/day methylprednisolone, and those who developed steroid-refractory aGvHD were treated with CD25 monoclonal antibody (CD25Mab) and/or mesenchymal stem cells, the latter was administered particularly to patients with intestinal tract grade III/IV aGvHD.

2.7 Virus reactivation, surveillance, and management

Epstein Barr virus (EBV) reactivation is defined as an EBV DNA level>50 equivalents per milliliter (Eq/mL) and positive EBV serology prior to transplantation. Recurrent EBV reactivation is defined as a positive polymerase chain reaction (PCR) result after at least two consecutive negative PCR results following a reactivation episode [23]. Cytomegalovirus (CMV) reactivation is defined as the detection of≥500 copies CMV DNA/mL [24]. As stated above, polyomavirus BK virus (BKV)-induced hemorrhagic cystitis (HC) is defined as the presence of hematuria concomitant with urinary BKV excretion and the absence of any other known causes of hematuria. Episodes of BKV HC were graded as follows: grade 1, microscopic hematuria; grade 2, macroscopic hematuria; grade 3, macroscopic hematuria with small blood clots; and grade 4, massive macroscopic hematuria requiring the instrumentation of the urinary tract or causing urinary retention [25].

Virus concentrations were monitored using Taqman real-time PCR (qPCR) performed weekly for 3 months, biweekly until 6 months, and monthly until 1 year after haplo-BMT [23,26,27]. EBV concentrations were measured weekly, however, in patients with cGvHD [28]. Nucleic acid was extracted from the plasma samples of recipients [27], and virus concentrations were measured using commercial diagnostic kits for EBV DNA (7756-2014, Wuhan Bio-tech Gene Engineering Co., China), human CMV DNA (20173401206, Wuhan Bio-tech Gene Engineering Co., China), and BKV DNA (OD-0102-02, Shanghai ZJ Bio-tech Co., China).

2.8 Management of EBV reactivation and proven (or probable) EBV disease

Proven EBV disease (post-transplant lymphoproliferative disorder (PTLD)) is defined based on lymph node histology and/or cytology [29,30]. Probable EBV disease is defined as serious lymphadenopathy (or other end-organ damage) with high blood EBV concentration. Immunosuppressive agents FK506 and glucocorticoids were tapered or discontinued in patients with proven or probable EBV disease due to the possibility of EBV reactivation and were treated using 200 mg of rituximab (Roche Pharma, Switzerland) per week for 2–4 consecutive weeks; when necessary, rituximab plus CHOP chemotherapy regimen (rituximab, cyclophosphamide, doxorubicin, vincristine, prednisolone) was used as a second-line treatment [31]. Recipients with>1000 CMV DNA copies/mL were treated with intravenous ganciclovir (10 mg/kg, once a day) or foscarnet (180 mg/kg, once a day) [32,33]. BK-associated HC was treated with ganciclovir and foscarnet sodium, which are usually combined with bladder irrigation and symptom relief [34].

2.9 Engraftment and immune reconstitution

Engraftment is defined as the recovery of neutrophil and platelet counts [35]. Neutrophil recovery is defined as a neutrophil count≥0.5 × 10⁹/L for 3 consecutive days, and platelet recovery as a platelet count≥20 × 10⁹/L for 7 consecutive days. Engraftment failure is defined as a neutrophil count that never reaches 0.5 × 10⁹/L or that reaches 0.5×10⁹/L, followed by a subsequent, non-transitory reduction in neutrophil count or loss of donor chimerism. Hematopoietic chimerism was evaluated using PCR amplification of short tandem repeats in extracted DNA. Full donor chimerism is defined as>95% donor hematopoietic cells and mixed chimerism as 5%–95% donor cells [16,36]. Lymphocyte counts, cellular immune reconstitution, and immunoglobulins from peripheral blood were monitored prior to HSC transplantation on days 30, 60, 90, 180, 365, and 730 after HSC transplantation.

2.10 Prevention of other complications and supportive cares

During conditioning, the recipients were administered with two doses of 10 mg/kg/day phenytoin sodium as prophylaxis for epilepsy on days -9 to -3 and an oral posaconazole suspension (4 mg/kg, three times a day) as fungal prophylaxis from day -10 to day+100 [37]. Sulfamethoxazole at a dosage of 1.0 g was administered twice per week as prophylaxis for Pneumocystis carinii from day -10 to day+100 [38]. Ganciclovir (5 mg/kg) was administered intravenously twice a day from day -10 to day -2, and acyclovir from day -1 to the time of cyclosporine discontinuation as prophylaxis for herpes simplex virus and varicella zoster virus infections. When necessary, RBCs were transfused to maintain the HB levels at 50-60 g/L and platelets to maintain the levels at 10×10⁹/L -20×10⁹/L prior to successful engraftment. All recipients were administered with G-CSF (5 mg/kg) from day+6 after transplantation until myeloid recovery.

2.11 Statistical methods

OS and disease-free survival (DFS) are expressed median±standard deviation (SD) and were evaluated using the Kaplan–Meier method. OS was measured from the date of HSC infusion until death from any cause. DFS was calculated from the date of HSC infusion to the first day of either recurrence or graft failure or poor graft function or death. SPSS version 13.0 (SPSS Inc., Chicago, USA) was used to perform all statistical analyses. Differences with P<0.05 were considered statistically significant.

3 Results

3.1 Patients

The 22 patients had a median age of 17 years (range, 5-35 years) and a median disease duration of 8 years (range, 1-22 years) from diagnosis until haplo-BMT (Table 1). No patient had a history of treatment with ATG. Median values were recorded for the following parameters prior to haplo-BMT: white blood cell (WBC) count, 1.84×10⁹/L (range, 0.74×10⁹/L-2.90×10⁹/L); neutrophils, 0.60 × 10⁹/L (range, 0.11×10⁹/L-0.95×10⁹/L); HB, 56.5 g/L (range, 23-82 g/L); platelets, 7 × 10⁹/L (range, 1×10⁹/L-18×10⁹/L); and serum ferritin, 2716 ng/mL (range, 512-9559 ng/mL). All patients were negative for donor-specific antibodies. Prior to haplo-BMT, the 22 patients received packed RBC transfusions at a median of 76 U (range, 14–192 U) for a median of 3 years (range, 1-6 years) and platelet concentrate transfusions at a median 42 U (range, 26–110 U) for a median of 2 years (range, 1-3 years) in attempt to correct anemia and/or thrombocytopenia prior to transplantation.

3.2 Engraftments

Among the 22 patients, 21 (95.5%) had achieved successful engraftment, 20 of whom engrafted used FLU/BU/CY/ATG conditioning regimen and one used FLU/CY/ATG in the second haplo-BMT. These patients exhibited a median donor-derived chimerism of 97.5% (range, 79.7%-100%), and 19 (86.4%) had survived after haplo-BMT. The median time for the neutrophil recovery of recipients was 11 days (range, 10-15 days), and that for platelet recovery was 16.5 days (range, 14-39 days). A total of 100 days after haplo-BMT, the median values of relevant parameters were as follows: WBC count, 3.83 × 10⁹/L (range, 2.52 × 10⁹/L–5.52 × 10⁹/L); neutrophils 2.11 × 10⁹/L (range, 0.99×10⁹/L–4.09×10⁹/L); HB, 93 g/L (range, 47–130 g/L); platelets, 106 × 10⁹/L (range, 17×10⁹/L–180×10⁹/L); percentage of reticulocytes, 2.17% (range, 0.63%–4.28%); and absolute reticulocyte count, 52.03 × 10⁹/L (range, 20.2×10⁹/L–129.7×10⁹/L).

A 17-year-old patient with a primary graft failure underwent a second haplo-BMT with FLU/CY/ATG through the transfusion of unrelated umbilical cord blood (UCB) on day 0 at 4 h before his father’s bone marrow grafts [16]. This strategy resulted in a successful engraftment with donor-derived chimerism remaining at>99.5% until day+100 and for>1 year after haplo-BMT. The modifications to the second haplo-BMT included the use of FLU/CY/ATG supplemented with UCB. FLU/CY/ATG may have reinforced the immunosuppression and eliminated the autoreactive T cells, and the mesenchymal stem cells from UCB may have promoted the repair of hematopoietic microenvironment, resulting in successful engraftment. These findings indicate that a second haplo-BMT supplemented with UCB may be an effective option after engraftment failure [39].

3.3 Liver and renal function

Liver transaminases, including aspartate transaminase (AST) and alanine aminotransferase (ALT), are useful biomarkers of liver injury in patients with intact liver function to some degree. During the conditioning regimen, three patients experienced liver dysfunction with elevated enzyme levels. One patient with severe liver dysfunction showed an increased ALT (550 IU/mL) and slightly high AST and gamma-glutamyl transpeptidase levels on day -5. The remaining two doses of CY in the conditioning regimen were halved to 25 mg/kg once a day, resulting in normal liver enzyme levels after 8 days (on day+3). One patient exhibited an increase in creatinine level—from 120.3 µmol/L on day+10 to 169.3 µmol/L on day+14. A reduction in FK 506 dose returned the creatinine levels to normal after 1 week. Five recipients experienced headache and hypertension after the infusion of bone marrow graft; however, these symptoms were alleviated after the treatment with oral nifedipine for 48 h. No one experienced recurrence.

3.4 Infection and immune reconstitution

During conditioning and engraftment, 12 out of the 22 (54.5%) patients experienced infections; three were caused by multidrug-resistant Klebsiella pneumoniae, and one by Escherichia coli as confirmed by bacterial culture of peripheral blood samples. Four patients experienced febrile neutropenia with unlocated infected sites. One had skin and soft tissue infection of the left upper arm, and two exhibited peri-anorectal abscesses. All these patients recovered after treatment with appropriate antibiotics. However, one patient died of Stenotrophomonas maltophilia-induced septic shock, in which the infection originated from the left maxillofacial space and rapidly expanded into the soft tissues around the neck. The patient died within 72 hours.

The median lymphocyte counts returned to normal level in nearly 1 year after HSCT (Table 2). Median NK and CD8⁺ cells recovered faster than CD4⁺ and CD19⁺ ones. For immunoglobulins, initial recovery occurred in IgM levels, followed by in IgG and IgA levels. However, these values did not return to normal level even after 2 years of HSCT. Therefore, the cellular and humoral immunocompromises may have been associated with the increase in infection rate.

3.5 Epilepsy

The first two transplanted patients presented with new-onset generalized tonic–clonic seizures on days+1 and+3, respectively, after the infusion of>1200 mL of bone marrow solution. Epilepsy was attenuated by diazepam and lasted for nearly 3 days. Oral levetiracetam (20 mg/kg/day, twice daily) was administered for 1 month to prevent the onset of epilepsy. Prior to hospitalization, these two patients had HB levels<40 g/L, which increased to>80 g/L following the infusion of bone marrow solution. Both were infused with mononucleated cells to maintain their HB levels at<60 g/L after stem cell infusion. Subsequently, none of the recipients experienced any episodes of epilepsy prior to successful engraftment.

3.6 Viral reactivation and virus monitoring

EBV was reactivated in nine recipients at a median of 18 days (range, 12-53 days) with median EBV loads of 3.39 × 10³ copies/mL (range, 873×10⁷-1.24×10⁷ copies/mL) after BMT. One recipient developed probable EBV disease with an EBV load of 2.4×10⁶ copies/mL on day+47, a persistent fever, and enlarged lymph nodes around the neck. Another was diagnosed with PTLD according to lymph node biopsy on day+80. These two patients were treated with rituximab (200 mg/week) for 2–3 weeks, which normalized body temperature and lymph node size and reduced the EBV load to 8.1 × 10³ copies/mL. Prior to BMT, none of the recipients were seropositive for CMV. After BMT, six recipients experienced CMV reactivation at a median of 26 days (range, 19-39 days) with a median viral load of 1.69 × 10³ copies/mL (range, 1.57 × 10³-7.85 × 10³ copies/mL). All six recovered after ganciclovir or foscarnet was administered. Nineteen recipients experienced hemorrhagic cystitis at a median 27 days (range, 14-33 days) after BMT. Among them, nine had grade III-IV HC, concomitant with a median BKV load of 6 × 10⁸ copies/mL (range, 3.28 × 10⁷-6 × 10¹¹ copies/mL). HC was alleviated by treatment with antiviral agents, and all patients recovered their immune function.

3.7 aGvHD and cGvHD

According to the MAGIC criteria for aGvHD, 4 out of 20 patients (19.0%) experienced grade II–IV aGvHD, two of which had grade III–IV aGvHD. The skin and intestinal tract were the organs mainly affected. The patients with grade II–IV aGvHD were successfully treated with methylprednisolone (1–2 mg/kg/day), anti-CD25 monoclonal antibody and/or infusion of 5 × 10⁶/kg mesenchymal stem cells once a week for 4 consecutive weeks when necessary. Gradual recovery was observed. According to the NIH 2014 criteria for cGvHD, 6 out of the 20 patients developed cGvHD with cumulative incidence of 28.6%. Among them, four had mild cGvHD, two had moderate cGvHD, and none had severe cGvHD during the follow-up period (as shown in Table 1). All these patients recovered after corticosteroid and/or FK506 treatment.

3.8 Bone marrow biopsy

Bone marrow biopsies were obtained before and after BMT from eight patients. Their hematopoietic trilineage pattern and cellularity were assessed using immunohistological staining. Bone marrow cellularity was<10%, which increased to 30%–60% 1 year after haplo-BMT (Table 3). All samples before BMT were negative for CD34⁺, a marker of HSCs; CD61, a marker for megakaryocytic precursor cells; and CD42b, a marker for differentiated megakaryocytic cells. By contrast, all samples post BMT were positive for these markers, suggesting that the HSCs had recovered, and trilineage hematopoiesis occurred in the bone marrow after BMT. A year after haplo-BMT, all these patients recovered with normal peripheral blood cell counts.

3.9 OS and DFS

Among the 22 patients with SAA-II who underwent haplo-BMT, one died of severe infection 5 days before HSC transfusion, and two died of fungal lung infection and thrombotic microangiopathy, respectively, after haplo-BMT. At the end of follow-up (median, 42 months), 19 out of the 22 patients remained alive, corresponding to the estimated 3-year OS and DFS rates of 86.4%±0.73% (Fig. 1A and 1B).

4 Discussion

ATG-based IST and HLA-matched allogeneic transplantation are considered standard treatment for SAA. In the absence of HLA-matched sibling donors [16], haplo-BMT has yielded promising outcomes. In a study for haplo-BMT performed at a median interval of 3 months (range, 1–8 months) after diagnosis, the 3-year OS rate was 84.5%±5.0% in 52 patients, and the 10-year OS rate was 89.3%±5.8% in 25 patients [40]. Haplo-BMT has recently emerged as an alternative treatment for newly diagnosed pediatric patients with SAA/vSAA. However, SAA-II, which progresses from NSAA, usually has a long disease duration accompanied by worsening general health and various types of medical management, including treatment with glucocorticoids, cyclosporine, and androgens; supportive care including RBC and platelet transfusion and antibiotics to treat recurrent infections; and even Chinese herbs with unknown components. Therefore, haplo-BMT maybe an alternative salvage therapy in patients with SAA-II.

In this study, 12 out of the 22 patients who underwent haplo-BMT developed bacterial infections. One had experienced intermittent infections during a history of NSAA for 21 years and finally died of severe sepsis caused by multidrug-resistant S. maltophilia in conditioning regimen. This finding suggested that SAA-II should be treated as early as possible to reduce the risk for secondary infections by multidrug-resistant microorganisms.

A large-scale retrospective study reported that BU/CY/ATG, FLU/CY/ATG, and CY/ATG are mostly used in patients with SAA who underwent haplo-BMT. These conditioning regimens had similar effects with 5-year OS rates of 68.5%–93.8% [41]. In the present work, 22 patients with SAA-II progressed from NSAA after a median of 8 years, most of whom were young with a median age of 17 years. Low-dose total body irradiation (TBI) carries a high risk for causing late effects in pediatric patients after allogeneic-BMT, such as growth impairment, cataracts, hypothyroidism, and secondary malignancies and therefore was not added to the conditioning regimen. FLU and ATG were incorporated to prevent graft failure through enhanced immunosuppression and lymphoablative effect similar to that of low-dose TBI to ensure that the recipients achieved a high level of donor engraftment and a low rate of aGvHD. Therefore, the conditioning regimen was modified to FLU/BU/CY/ATG to improve engraftment and resulted in successful primary engraftment in 20, and engraftment in 1 with the second transplantation, out of 22 patients. Moreover, no one experienced severe liver or renal dysfunction, indicating that haplo-BMT with FLU/BU/CY/ATG is safe and effective in patients with SAA-II.

Most SAA-II patients have a history of blood transfusion for long-term anemia and iron overload prior to transplantation and therefore are likely to die of infection and organ failure following allogeneic HSCT [42]. Although the 22 included patients had a high median serum ferritin level (2716 ng/mL) prior to transplantation, 19 patients remained alive with a 3-year OS rate of 86.4%±0.73%, indicating that a high serum ferritin level does not affect the outcomes of haplo-BMT in patients with SAA-II.

Haplo-BMT has yielded a 10-year OS rate of 73.4%±12.6% in children<17 years of age [41,43] and therefore may be a good front-line treatment for children with SAA or a valid alternative option for SAA patients<40 years of age without HLA-matched sibling donors [41]. Nearly all of these patients were children with newly occurring SAA and had a median interval from diagnosis to HSCT of only 3 months [41,43]. This study was the first to assess the treatment outcomes of haplo-BMT in patients with SAA-II and revealed their 3-year OS rate of 86.4%±0.73%, indicating that haplo-BMT may be a promising salvage option for patients with SAA-II.

The outcomes in these 22 patients with SAA-II who underwent haplo-BMT may provide clues to improve the treatment strategies for SAA-II in the absence of HLA-matched sibling donors and other effective treatments. Additional multi-center prospective studies are required to evaluate the efficacy of haplo-BMT for SAA-II treatment.

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