Scholarly article on topic 'A Well-Tolerated Regimen of 800 cGy TBI-Fludarabine-Busulfan-ATG for Reliable Engraftment after Unmanipulated Haploidentical Peripheral Blood Stem Cell Transplantation in Adult Patients with Acute Myeloid Leukemia'

A Well-Tolerated Regimen of 800 cGy TBI-Fludarabine-Busulfan-ATG for Reliable Engraftment after Unmanipulated Haploidentical Peripheral Blood Stem Cell Transplantation in Adult Patients with Acute Myeloid Leukemia Academic research paper on "Clinical medicine"

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{"Acute myeloid leukemia" / "Haploidentical hematopoietic stem cell transplantation" / "Total body irradiation" / "T cell replete"}

Abstract of research paper on Clinical medicine, author of scientific article — Seung-Ah Yahng, Jung-Ho Kim, Young-Woo Jeon, Jae-Ho Yoon, Seung-Hwan Shin, et al.

Abstract Eighty adult patients with acute myeloid leukemia (AML) received peripheral blood T cell–replete HLA haploidentical hematopoietic stem cell transplantation (haplo-HSCT). Disease status at transplantation was either first or second complete remission (CR, n = 69) or relapse/refractory (n = 11). Identical transplant-related procedures with conditioning regimen consisting of fractionated 800 cGy total body irradiation (TBI), fludarabine (30 mg/m2/day for 5 days), busulfan (3.2 mg/kg/day for 2 days), and antithymocyte globulin (1.25 mg/kg/day on days −4 to −1) and graft-versus-host disease (GVHD) prophylaxis with tacrolimus and methotrexate were used in all patients. Recovery of neutrophil (median, 11 days) and platelet (median, 10 days) counts was achieved in all patients with full donor chimerism (≥99%), and no delayed engraftment failure was observed. The cumulative incidence of grades III to IV acute GVHD and moderate to severe chronic GVHD was 11.2% and 26.3%, respectively. A donor CD8+ and CD4+ T cell dose above the median value was significantly associated with the incidences of grades II to IV acute GHVD and moderate to severe chronic GVHD, respectively. After a median follow-up of 28 months for survivors, the 2-year cumulative incidences of relapse (n = 20) and nonrelapse mortality (n = 10) were 26.6% and 12.2%, respectively. Although all but 1 patient in relapse/refractory status died, the 2-year overall and progression-free survival of patients in first CR was 82.5% and 75.1%, respectively. We suggest the strategy of fractionated 800 cGy TBI-based conditioning with unmanipulated peripheral blood stem cell grafts seems feasible with favorable outcomes for adult patients with AML undergoing haplo-HSCT in CR.

Similar topics of scientific paper in Clinical medicine , author of scholarly article — Seung-Ah Yahng, Jung-Ho Kim, Young-Woo Jeon, Jae-Ho Yoon, Seung-Hwan Shin, et al.

Academic research paper on topic "A Well-Tolerated Regimen of 800 cGy TBI-Fludarabine-Busulfan-ATG for Reliable Engraftment after Unmanipulated Haploidentical Peripheral Blood Stem Cell Transplantation in Adult Patients with Acute Myeloid Leukemia"

Accepted Manuscript

A well-tolerated regimen of 800 cGy TBI-fludarabine-busulfan-ATG for reliable engraftment after unmanipulated haploidentical peripheral blood stem cell transplantation in adult patients with acute myeloid leukemia

Seung-Ah Yahng, Jung-Ho Kim, Young-Woo Jeon, Jae-Ho Yoon, Seung-Hwan Shin, Sung-Eun Lee, Byung-Sik Cho, Ki-Seong Eom, Yoo-Jin Kim, Seok Lee, Chang-Ki Min, Seok-Goo Cho, Dong-Wook Kim, Jong-Wook Lee, Woo-Sung Min, Chong-Won Park, Hee-Je Kim

PII: S1083-8791(14)00610-7

DOI: 10.1016/j.bbmt.2014.09.029

Reference: YBBMT 53609

To appear in: Biology of Blood and Marrow Transplantation

Received Date: 30 April 2014 Accepted Date: 30 September 2014

Please cite this article as: Yahng SA, Kim JH, Jeon YW, Yoon JH, Shin SH, Lee SE, Cho BS, Eom KS, Kim YJ, Lee S, Min CK, Cho SG, Kim DW, Lee JW, Min WS, Park CW, Kim HJ, A well-tolerated regimen of 800 cGy TBI-fludarabine-busulfan-ATG for reliable engraftment after unmanipulated haploidentical peripheral blood stem cell transplantation in adult patients with acute myeloid leukemia, Biology of Blood and Marrow Transplantation (2014), doi: 10.1016/j.bbmt.2014.09.029.

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Title page

Journal section designations

- Scientific section designations: Transplantation

Title: A well-tolerated regimen of 800 cGy TBI-fludarabine-busulfan-ATG for reliable engraftment after unmanipulated haploidentical peripheral blood stem cell transplantation in adult patients with acute myeloid leukemia

Names of author: Seung-Ah Yahng,1 Jung-Ho Kim2, Young-Woo Jeon2, Jae-Ho Yoon,2 Seung-Hwan Shin,2 Sung-Eun Lee,2 Byung-Sik Cho,2,3 Ki-Seong Eom,2,3 Yoo-Jin Kim,2 Seok Lee,2 Chang-Ki

2 2 2,3 2 2 2

Min, Seok-Goo Cho, Dong-Wook Kim, , Jong-Wook Lee, Woo-Sung Min, Chong-Won Park, and Hee-Je Kim2,3

Author's affiliation:

department of Hematology, Incheon St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Republic of Korea; 2Catholic Blood and Marrow Transplantation Center, Department of Hematology, College of Medicine, The Catholic University of Korea, Seoul, Republic of Korea; 3Cancer Research Institute, Seoul St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Seoul, Republic of Korea

Short title: T-cell replete haploidentical HSCT in AML

Key words: acute myeloid leukemia; haploidentical hematopoietic stem cell transplantation; total body irradiation; T cell replete

Corresponding author: Hee-Je Kim, M.D., Ph.D.

Address:

Department of Hematology, Catholic Blood and Marrow Transplantation

Center, Cancer Research Institute, Seoul St. Mary's Hospital, College of

Medicine, The Catholic University of Korea, 222, Banpo-daero, Seocho-

gu, Seoul, 137-701, Republic of Korea

Phone:

82-2-2258-6054

82-2-599-3589

E-mail:

cumckim@catholic.ac.kr

Word count for text and abstract: words 5030(text), words 249(abstract) Figure/table count: 4 figures/ 3 tables & 1 Supplementary table Reference count: 77 references

Authorship Contributions

HJK was the principal investigator and takes primary responsibility for the paper; SAY and HJK collected data and performed the statistical analysis, interpreted the results, and wrote the manuscript; JHY, SEL, and BSC participated in the clinical data management; KSE, YJK, SL, and CKM participated in performing the laboratory work for this paper; YWJ and JHY participated in the statistical analysis; SGC, DWK, JWL, WSM, and CWP provided patient care and commented on the manuscript.

Disclosure of Conflicts of Interest

The authors report no potential conflicts of interest to disclose. Abbreviations

PBSC, peripheral blood stem cell; haplo-HSCT, HLA-haploidentical hematopoietic stem cell transplantation; TCR, T cell-replete; TCD, T cell-depleted

Abstract

Eighty adult patients with acute myeloid leukemia (AML) received peripheral blood T cell-replete (TCR) HLA-haploidentical hematopoietic stem cell transplantation (haplo-HSCT). Disease status at transplantation was in first or second complete remission (CR, n=69) or in relapse/refractory (n=11). Identical transplant-related procedures with conditioning regimen consisting of fractionated 800 cGy total body irradiation (TBI), fludarabine (30 mg/m2/day for 5 days), busulfan (3.2mg/kg/day for 2 days), and antithymocyte globulin (1.25mg/kg/day on days -4 to -1), and graft-versus host disease (GVHD) prophylaxis with tacrolimus and methotrexate was used in all patients. Recovery of neutrophil (median, 11 days) and platelet (median, 10 days) counts was achieved in all patients with full donor chimerism (> 99%), and no delayed engraftment failure was observed. The cumulative incidence of grade III-IV acute (aGVHD) and moderate-severe chronic GVHD (cGVHD) was 11.2% and 26.3%, respectively. A donor CD8+ and CD4+ T cell dose above the median value was significantly associated with the incidences of grades II-IV aGHVD and moderate-severe cGVHD, respectively. After a median follow-up of 28 months for survivors, the 2-year cumulative incidences of relapse (n = 20) and non-relapse mortality (n = 10) were 26.6% and 12.2%, respectively. Although all but one of the patients in relapse/refractory status died, the 2-year overall and progression-free survival of patients in first CR was 82.5% and 75.1%, respectively. We suggest that the strategy of fractionated 800 cGy TBI-based conditioning with unmanipulated peripheral blood stem cell graft seems feasible with favorable outcomes for adult patients with AML undergoing haplo-HSCT in CR.

Introduction

Although hematopoietic stem cell transplantation (HSCT) provides a curative opportunity for patients with acute myeloid leukemia (AML), identifying a compatible related or unrelated donor requires considerable searching time and costs, especially for high-risk patients in urgent need of transplantation [1-3]. Haploidentical HSCT (haplo-HSCT) is an attractive strategy for these patients offering advantages such as readily available donors, control of preferred composition of graft, and timely application of repeated stem cell donation [4, 5]. Until recently, however, haplo-HSCT has been limited by high rates of graft loss and acute graft-versus host disease (aGVHD) [6-10].

Starting in the early 1990s, strategies for partial T cell depletion (TCD) were adopted to overcome the HLA barrier and control of GVHD in haplo-HSCT setting [11, 12]. Aversa and co-workers were the pioneers of using mega-dosed peripheral blood stem cells (PBSCs) with ex vivo TCD following myeloablative conditioning (MAC) in haplo-HSCT, which led to a major advance in the achievement of a high engraftment rate across the HLA barrier in the absence of GVHD [13]. Delayed immune reconstitution due to the few T cells in the graft can be challenging [14, 15]. Therefore, the recent developments in the approach using an adoptive transfer of donor T regulatory cells followed by donor conventional T cells seem very promising [16].

An alternative approach, developed at Johns Hopkins University, of using reduced-intensity conditioning (RIC) with post-transplantation cyclophosphamide in unmanipulated marrow-graft haplo-HSCT showed low non-relapse mortality (NRM) [17]. Outcomes were promising when the use of high dose, post-transplantation cyclophosphamide was combined with MAC and T cell-replete (TCR) graft for advanced/refractory hematologic malignancies [18, 19]. Recent studies have also shown the feasibility of substituting PBSC as a graft source in this setting [20, 21]. A small prospective phase II trial of MAC haplo-HSCT using TCR PBSCs showed engraftment in all patients with relatively favorable 1-year NRM (50%)[22], whereas a report by Kurokawa et al. showed NRM approaching 70% after TCD haplo-HSCT using 1 Gy total body irradiation (TBI)-based regimen [23]. Investigators from Peking University have consistently reported promising results of MAC haplo-HSCT using TCR granulocyte-colony stimulating factor (G-CSF) mobilized peripheral blood and bone marrow grafts [24, 25]. Favorable outcomes of TCR haplo-HSCT with a non-TBI based RIC regimen was reported by Lee et al. on 83 patients with AML or myelodysplastic syndromes [26].

TBI-based conditioning regimen for patients with leukemia receiving allogeneic HSCT are used with the intent of eradicating residual leukemic cells and/or reducing leukemic burden as well as for potential

immunosuppressive effects [27]. And yet, data regarding optimal dose requirements and fractionation employed during TBI for various transplantation settings are relatively limited. While 12 Gy TBI delivered twice daily over 3 days in combination with chemotherapy have been most commonly used for TBI-based MAC regimen, we had experienced poor survival outcome after haplo-HSCT following our earlier conditioning regimen, consisted of a fractionated 12 Gy TBI plus fludarabine, busulfan, and antithymocyte globulin (ATG) [10]. Therefore, to lower the risk of radiation-induced toxicity but allow adequate immunosuppression and leukemic cell eradication in the setting of unmanipulated G-CSF-mobilized PBSC haplo-HSCT, we have employed TBI-based conditioning regimen delivering 800 cGy, twice daily over 2 days followed by fludarabine, intravenous busulfan, and ATG with the standard GVHD prophylaxis. In this report, we analyzed 80 consecutive adult patients with AML undergoing haplo-HSCT using this approach to evaluate the efficacy and safety of this regimen.

Methods Patients

Eighty consecutive adult patients with AML who received haplo-HSCT between August 2008 and July 2013 at Catholic Blood and Marrow Transplantation Center were analyzed. Haplo-HSCT was offered to patients if they had no available > 7/8 HLA antigen-matched unrelated donor. Twenty-three patients previously reported were included with further follow-up for this analysis [28]. Median age of patients was 41 years (range, 16-69), with 19% > 60 years of age (Table 1). For the patients with better cytogenetic-risk [29] transplanted in first complete remission (CR1; 10 of 17 subjects with inv(16)(13.1q22)/ t(16;16)(p13.1;q22) or t(8;21)(q22;q22)), reasons for transplantation were myeloid sarcoma involving nervous system (n = 1), c-kit (n = 5) or FLT3/ITD (n = 1) mutations, secondary AML (n = 1), high aberrant expression of CD19 (n = 1), and presence of minimal residual disease after repetitive chemotherapy (n = 1). Among twenty-one (58%) of 36 patients with intermediate cytogenetic-risk in CR1 (normal karyotype, n = 24; cytogenetic abnormalities not classified as favorable or adverse, n = 12), factors associated with high-risk AML included: secondary AML (n = 2), refractoriness to first- (n = 3) or second- (n = 2) line chemotherapy, hyperleukocytosis (> 50 x 109/L, [n = 14]), and FLT3/ITD mutation (n = 4). Patients with t(9;22) or t(15;17) were excluded. All patients and donors had certified their fully informed consent as per the Declaration of Helsinki. This study was approved by the institutional review board. Donors

Donors had a median age of 33 years (range, 7-61). All donor-recipient pairs were typed for HLA-A, -B, -C and -DR loci by high-resolution DNA techniques. For each pair, the patient shared one haplotype and mismatched the unshared haplotype with some difference in degree. Among 42 non-inherited maternal antigens-(NIMA) complementary transplantations [30], offspring and maternal donors were 19 (45%) and 23 (55%), respectively. Because parental HLA typing was not available for sibling recipients, NIMA-mismatched sibling donor could not be identified. Donors who were classified as NK-alloreactive against patients possessed HLA-B and -C killer cell immunoglobulin-like receptor (KIR) ligand(s) that were missing in the patient. All but one of the donors and all recipients were cytomegalovirus (CMV) seropositive at HSCT. In cases of multiple available donors, the mother had priority under consideration of other factors, such as donor age, co-morbidity, and ABO matching (Table 1).

Conditioning regimen and GVHD prophylaxis

All patients received an identical conditioning regimen comprised of 800 cGy TBI (fraction size of 200 cGy given twice a day on days -9 and -8), fludarabine (30 mg/m2/day intravenously on days -7 to -3), busulfan (3.2 mg/kg/day intravenously in four divided doses on days -6 and -5), and rabbit ATG (Thymoglobulin®, Sanofi/Genzyme, Cambridge, MA, USA; 1.25 mg/kg/day on days -4 to -1). A combination of tacrolimus (intravenously continuous infusion with starting dose of 0.03 mg/kg from days -1 to +21, and then orally at a total of .12 mg/kg/day) plus short-course methotrexate (10 mg/m2 intravenous bolus injection on days +1, +3, +6, and +11) was used for GVHD prophylaxis in all patients [10]. Individualized dosage adjustment of tacrolimus was based on plasma level to maintain a target dose of 8-12 ng/mL. Tacrolimus was tapered off from day +90 after transplant in the absence of aGVHD. Supportive care

Recombinant human G-CSF (5 ^g/kg/day) was administered subcutaneously from day +7 until neutrophil recovery. Intravenous ganciclovir (5 mg/kg twice daily) was administered during the conditioning period for CMV prophylaxis. Serial weekly monitoring for CMV viremia using quantitative polymerase chain reaction (PCR) assays was initially started from neutrophil recovery during the first 3 months followed by monitoring every 2 weeks or at each follow-up visit. Ganciclovir or foscarnet was initiated with evidence of CMV reactivation defined by PCR > 500 copies/mL on two consecutive assays. Recovery of lymphocyte subpopulations was assessed by flow cytometry of peripheral blood samples 1, 3, 6, 9, and 12 months after transplantation. The strategy for other general supportive care followed a previously described institutional

protocol [31, 32]. Hematopoietic stem cell harvest

All donors were primed with G-CSF (10 ^g/kg/day) subcutaneously for 4-6 consecutive days from day -4. PBSC harvesting was repeated on the next day or two if the target CD34+ cell count (a minimum of 5 x 106 cells/kg of recipient body weight) was not reached. The collected unmanipulated graft was infused into the recipient on the day of harvesting. The composition of infused PBSCs was evaluated for cell surface markers using monoclonal antibodies.

Definition

Myeloid and platelet engraftment was defined as the first of 3 consecutive days with an absolute neutrophil count (ANC) of > .5 x 109/L and the first of 7 consecutive days with a platelet count > 20 x 109/L without transfusion, respectively. Failure to neutrophil engraftment by day +28 was considered primary engraftment failure. Secondary engraftment failure was defined as an initial engraftment with documented donor-derived hematopoiesis followed by loss of graft function. Complete (at least 95%) donor chimerism was evaluated at 1, 3, 6, and 12 months by PCR DNA finger printing of short tandem repeats on recipient peripheral T lymphocytes. GVHD was diagnosed and graded according to the clinical consensus criteria [33, 34]. Statistical analysis

Categorical and continuous variables were compared by the x2 test and Mann-Whitney's test, respectively. Spearman's rank correlation coefficient was used to associate graft composition and engraftment. Graft cell doses were analyzed as categorical variables (< versus > median value) for transplant outcome. Overall survival (OS) and progression-free survival (PFS), assessed from the day of transplant, were estimated by the KaplanMeier method and compared by the log-rank test. Probability of relapse, NRM, and aGVHD and chronic GVHD (cGVHD; severity measures referring to peak) were estimated using the cumulative incidence method and compared by the Gray test. P values < .05 were considered significant. Analyses were conducted using the Statistical Package for the Social Sciences, ver. 13.0 (SPSS, Inc, Chicago, IL, USA) and R software (http: //cran. r-proj ect. org/).

Results

Engraftment, graft composition, and recovery of T cell subsets

Sustained myeloid engraftment with median 11 days (range, 9-42) was achieved in all patients (> 60 years,

median 10 days [range, 6-47]), and all but one (early NRM) of them had platelet recovery at median 10 days (range, 6-47)(Figure1). Donor chimerism of > 99% was confirmed in 75 of all 80 (94%) patients by day +30, and in the remaining 5 patients within the next month. No graft loss has occurred in the survivors so far. The median number of CD34+ cells transplanted was 6.48 x 106 (range, 2.22-39.7 x 106)/kg of recipient body weight, the median numbers of donor CD4+ cells and CD8+ cells were 253 x 106 (range, 102-942 x 106)/kg and 172 x 106 (range, 51.7-643 x 106)/kg, respectively. The median number of CD3-CD56+ cells was 96.7 x 106 (range, 16.9-366 x 106)/kg. A strong concordance was found between the graft cell doses of CD3+ and CD4+ (P < .0001) or CD8+ (P < .0001) as well as between CD4+ and CD8+ (P < .0001). The CD34+ cell dose was less than the targeted dose in 42 donors (53%) after first apheresis (more than 2 aphereses were performed in 16 donors). Higher CD34+ cell dose correlated with significantly less time for ANC (R2 = -.365, P = .001) recovery, but only with borderline significance with platelet (R2 = -.203, P = .071) recovery. Other factors including patient or donor age showed no significant correlation with time to engraftment. On the other hand, we found a significantly higher number of CD4+ cells (R2 = .295, P = .008) with increasing donor age, while cell doses of other graft composition showed no correlation. This may be related to less yield of CD34+ cell collection after the first apheresis with a tendency towards increasing donor age (R2 = -.196, P = .082), which led to subsequent aphereses to achieve the target dose. The mean values of lymphocyte subsets are illustrated in Figure 2. Briefly, the median number of absolute CD8+ and CD4+ T cells among evaluable patients were 22/^L and 3/^L at 1 month posttransplantation (n = 64), 539/^L and 117/^L at 3 months (n = 44), 1227/^L and 203/^L at 6 months (n = 23), and 1364/^L and 318/^L at 1 year posttransplantation (n = 16), respectively. As expected, the median number of CD3-CD56+ NK cells showed higher proportions during the first 6 months posttransplantation: 198/^L (n = 64), 404/^L (n = 44), and 486/^L (n = 23) at 1, 3, and 6 months after transplant, respectively. We found no significant impact of pre-transplant variables on lymphocyte reconstitution (data not shown).

The cumulative incidences of grades II-IV (n = 38) and grades III-IV (n = 9) de novo aGVHD at 100 days was 47.5% (95% CI, 36.2-58.0) and 11.2% (95% CI, 5.4-19.2), respectively, occurring at a median time of 20 days (range, 12-84) after transplantation with no late-onset. Of the nine patients with severe aGVHD, the disease status at transplantation was standard (CR1) in 4 patients, and advanced in 5 (second [CR2, n = 2] or third [CR3, n = 1] remission; relapsed/refractory, n = 2). The patients with grade III aGVHD (n = 6) were all manageable with steroids and mycophenolate mofetil (MMF) added on tacrolimus, although three of them later showed

chronic features consistent with moderate cGVHD. Of the patients with grade IV aGVHD (n = 3), one patient who received etanercept after failing triple combination of tacrolimus, steroid and MMF is currently alive without signs of GVHD, whereas two other patients died of severe infection during treatment of GVHD. After a median time of 3.1 months (range, 0.7-7.5), 36 patients developed cGVHD (overlap syndrome, n = 14; classic chronic, n = 22) with the cumulative incidence of 45.0% (95% CI, 33.8-55.6), while that of moderate-severe cGVHD (n = 21) was 26.3% (95% CI, 17.1-36.3). Of the 21 patients with moderate (n = 19) or severe (n = 2) cGVHD, 2 patients with severe cGVHD died from severe infections, and 5 patients eventually relapsed of AML during cGHVD management. The rest of the patients (n = 14) had received the combinations of tacrolimus, systemic steroid and/or MMF, together with localized therapy including phototherapy or topical agents. Whereas, 6 of 46 disease-free survivors with follow-up duration of more than 1 year needed immunosuppression after the first year post-transplant, while 22 of the 50 patients who are alive in disease-free status went back to their works.

As shown in Table 2, donor CD8+ and CD4+ T-cell doses above the median value were significantly associated with the incidences of grades II-IV aGHVD (< vs. > median value of CD8+, 35% vs. 60%; P = .021) and moderate-severe cGVHD (< vs. > median value of CD4+, 13% vs. 40%; P = .007), respectively (Figure 3). Because there were no significant associations between any of the other pre-transplant variables and acute or chronic GVHD, we arbitrarily performed multivariate analyses by choosing donor age, previous transplantation, and patient age as confounding correlates, which showed that CD8+ and CD4+ cell doses dichotomized by the median values were independently associated with grades II-IV aGHVD (HR 2.4, 95% CI 1.2-4.9; P = .014) and moderate-severe cGVHD (HR 4.0, 95% CI 1.5-11.1; P = .007), respectively. Of the 42 recipients, evaluable for NIMA effects, lower incidence of grades II-IV aGVHD was seen in the group of female patients with offspring donors (n = 19; NIMA-mismatched) compared to 23 patients with maternal donors without statistical significance (36.8% ± 11.0% vs. 52.2% ± 10.7%, P = .242). Survival

After a median follow-up of 28.0 months (range, 7.2-67.1) for survivors, 55 patients were alive and 25 patients had died from NRM (n = 10) or relapse (n = 15). After the post-transplant relapse, reinduction chemotherapy followed by second allogeneic transplantation (double-cord blood transplantation, n = 3; HLA-matched unrelated transplantation, n = 1) were performed which left one patient dead due to pneumonia, another relapsed after the second transplantation, and two other patients are currently alive in disease-free status. The remaining

one patient decided to receive conservative care and is deceased. The 2-year OS and PFS were 66.0% ± 5.8% and 61.1% ± 5.80%, respectively, in all patients, while the OS and PFS were significantly different according to the disease status (Figure 4A, 4B). At 2 years, OS and PFS of 52 patients in CR1 (n = 45) or iCR1 (n = 7) were 82.5% ± 5.7% and 75.1% ± 7.1%, respectively, and 78.2% ± 6.8% and 71.1% ± 7.1%, respectively, at 3 years. Univariate analyses showed that advanced disease status at HSCT (HR 3.9, 95% CI 1.9-8.1; P < .0001), donor age (HR 1.03, 95% CI 1.00-1.05; P = .029), and occurrence of grades III-IV aGVHD (HR 3.1, 95% CI 1.3-7.3; P = .009) were associated with worse PFS (Supplementary Table 1). Similar factors were also significantly associated with OS (Supplementary Table 1). Multivariate analyses revealed that the advanced disease status (OS, P = .001; PFS, P = .001) and occurrence of aGVHD (OS, P = .046; PFS, P = .016) were the only independent prognostic factors for OS and PFS (Table 3). Relapse

Of all 80 patients, 20 patients had relapsed after HSCT with the 2-year cumulative incidence of 26.6% ± 5.2%. Relapse was first detected on bone marrow examination routinely performed every 3 months during the first 2 years after transplant (n = 10), or by circulating leukemic blasts (n = 8) or development of myeloid sarcoma (n = 2) confirmed by subsequent bone marrow examination (n = 8). The incidence according to disease status was 16.6% ± 5.5% in CR1/iCR1, 18.2% ± 9.9% in CR2/CR3, and 77.3% ± 17.3% in relapsed/refractory status (P < .0001; Figure 4C). Nine patients with relapsed/refractory status relapsed at median 5.5 months (range, 1.0-8.5), whereas the patients in CR1 (n = 8) and CR2/CR3 (n = 3) relapsed at a median time of 8.9 months (range, 3.816.1) and 5.8 months (range, 5.3-9.4), respectively. No significant difference in 3-year relapse incidence according to cytogenetic-risk, whether better (24.2% ± 10.9%), intermediate (23.6% ± 6.4%), or poor (41.0% ± 15.4%) was shown (P = .413). In further univariate analysis, advanced disease status (HR 3.7, 95% CI 1.6-8.9; P = .003) and white blood cell (WBC) count of > 50x109/L at diagnosis (HR 2.2, 95% CI 0.9-5.3; P = .07) were significantly associated with incidence of relapse (Supplementary Table 1). The multivariate analysis showed advanced disease status (P = .001) and WBC count of > 50x109/L (P = .029) were independently associated with higher incidence of relapse (Table 3).

NRM and complications

Ten patients (10%) died from transplantation-related complications at a median of 3.8 months (range, 1.4-32.4), resulting in the 2-year cumulative NRM incidence of 12.2% (95% CI, 5.9-20.1). Causes were refractory GVHD (one aGVHD, two cGVHD), veno-occlusive disease (n = 2), and infections, whether bacterial (one sepsis from

pyogenic spondylitis, one bacterial pneumonia) fungal (one invasive aspergillosis), and viral (one meningitis, one hepatic failure from hepatitis B virus reactivation). In univariate analysis, the occurrence of grades III-IV aGVHD (P = .054) and donor age (P = .054) were associated with NRM with borderline significance (Supplementary Table1). Multivariate analysis showed that only the occurrence of grades III-IV aGVHD was associated with higher incidence of NRM with borderline significance (Table 3). CMV reactivation requiring antiviral therapy for preemptive purpose (n = 53) and/or CMV disease (n = 11) occurred in 72%. In univariate analysis, donor CD8+ cell count (HR 1.79, 95% CI 1.03-3.11; P = .039), male donor (HR 1.67, 95% CI 0.952.96; P = .077), and previous HSCT (HR 2.04, 95% CI 1.15-3.62; P = .015) were associated with CMV reactivation requiring pre-emptive therapy. When adjusted in multivariate analysis, previous HSCT remained the only significant predictor of CMV reactivation. KIR ligand(s)-mismatch and GVHD had no impact on CMV reactivation (data not shown).

Discussion

The results of our study are noted in particular by rapid and sustained engraftment with achievement of complete donor chimerism in the total of all eighty patients with AML after haplo-HSCT. The use of G-CSF primed PBSC ensured adequate yield of donor CD34+ cell dose, while fractionated 800 cGy TBI-based conditioning with low-dosed ATG might have provided sufficient room for donor hematopoietic stem cells to engraft. By using unmanipulated graft, we observed only 6% mortality from infectious causes, with low incidence of NRM (12%). Our data represents a homogeneous group of patients in terms of disease type and transplantation-related procedures.

The use of "mega-dose" of PBSC content with effective T cell depletion (median: 2 x 104 CD3+ cells/kg) for haplo-HSCT resulted in very low rates of aGVHD [35, 36], while reported incidence of grades II-IV aGVHD after unmanipulated graft haplo-HSCT ranges around 30%-40% [19, 22, 25]. A possible potential cause for relatively higher incidence of overall GVHD in our study using TCR graft may have come from our policy of rapid tapering-off of tacrolimus starting from day +90 in the absence of severe aGVHD features. All but two patients with severe aGVHD (grade III aGVHD in 6 patients and grade IV aGVHD in 3 patients) were manageable with restart of tacrolimus and/or addition of systemic steroid and/or MMF. Nevertheless, PFS was affected by the occurrence of severe aGVHD, and we found that a donor graft CD8+ cell dose was significantly related to its incidence. While significant correlation was seen between doses of CD4+ and CD8+ cells, higher

number of CD4+ cell dose was associated with incidence of cGVHD in our study. Higher donor CD3+ cells in G-CSF-primed PBSC products have been suggested to be related to aGVHD or cGVHD in some studies [38] but not in others [39]. Efficient TCD, typically targeting less than the critical threshold of 1 x 105 CD3+ cells/kg of recipient body weight, is beneficial by means of negligible severe aGVHD at the expense of delayed immune recovery with high NRM from infectious complications [13, 40]. Although it has been suggested that increasing the T-cell dose beyond the critical threshold does not necessarily result in higher incidence of GVHD, the findings had been demonstrated in HLA-matched HSCT setting [41]. Because our transplantation protocol does not restrict the number of donor T-cells, the median dose of CD3+ cells infused was relatively higher than other studies [22, 24, 42, 43]. In this aspect, it is expected that the recent innovative tryouts of "designed" haplo-grafts to not only separate the graft-versus-leukemia (GVL) effect from GVHD but also to facilitate expansion of donor T cells for long-term immunity will lead to promising result [16, 44, 45].

At least until the present time, almost all Korean patients, as well as the donors, are seropositive for CMV IgG, a similar situation in other Asian populations [24, 26, 46], and show high incidence of CMV reactivation after HSCT [47, 48]. Therefore, the use of extensive TCD graft or intensified GVHD prophylaxis such as posttransplantation cyclophosphadmide may raise concern in terms of fatal CMV infections in populations with prevalent CMV seropositivity. By infusing large numbers of peripheral blood progenitor cells from TCR graft and using G-CSF after transplantation, a fast hematopoietic recovery could be achieved in our study. On the other hand, although the grafts were unmanipulated and tacrolimus was discontinued as early as possible, the median CD4+ cell count at 12 months since transplant in evaluable recipients (n = 16) was less than 400/^l in our study, showing a similar finding to what has been reported after an extensive TCD mismatched transplant. The use of G-CSF after allogeneic bone marrow transplantation has been met with concerns mainly because of the reported risks of GVHD and TRM in some studies including a retrospective study of the European Blood and Marrow Transplantation group [49] whereas the majority of studies on its use after allogeneic PBSC transplantation did not find an increase in the incidence of GVHD [50]. On the other hand, earlier studies in animals and human volunteers showed that G-CSF attenuates the production of proinflammatory cytokines such as tumor necrosis factor alpha, interleukin-12, and interferon gamma [51], and exerts anti-inflammatory influence by increasing interleukin-10 production [52] and promoting T-helper-2 immune deviation [53, 54]. Therefore, it may be argued that G-CSF administered after transplantation could negatively influence on the quality of posttransplantation immune reconstitution. In fact, in the setting of TCD haploidentical transplants,

posttransplantation G-CSF administration was stopped subsequent to the observation by the Perugia group that it promoted T-helper-2 immune deviation leading to functional impairment of immune recovery [15, 55-57]. On the contrary, the shift towards an anti-inflammatory cytokine pattern by the use of G-CSF could be considered as an advantage in the context of non-TCD PBSC transplantation setting because it may attenuate the risk of aGVHD [53, 54]. Although earlier studies on TCR transplantations found no benefit or disadvantage of giving G-CSF after transplantations [58, 59], most of the recent studies reported on transplantation outcome of TCR haplo-HSCT from numerous centers [19, 21, 22, 26, 37, 60-62] have used G-CSF after transplantation to ensure faster neutrophil recovery, suggesting that it is safe to use postgrafting G-CSF in TCR haplo-HSCT. Future experimental and clinical evidence on functional data of immune recovery to evaluate the anti-CMV response in a time dependent manner and also to correlate prospectively the T cell receptor diversity and specificity with acquisition of immune-competence in presence/absence of GVHD and/or infection in the setting of TCR haplo-HSCT are needed to provide better understanding in immune reconstitution after our transplant approach including the use of post-grafting G-CSF.

To lower the risk of radiation-induced toxicity but allow adequate immunosuppression and leukemic cell eradication, we have consistently used a multiple daily fractionated 800 cGy TBI-based conditioning regimen for adult patients with hematologic disorders receiving haplo-HSCT. Previous randomized studies showed that increasing the TBI dose from 1200 cGy to 1575 cGy showed lower relapse rate in AML patients at the expense of a significantly higher early NRM [63, 64]. Lowering the TBI dose to 990 cGy was associated with a higher relapse rate without the benefit of reducing NRM [65]. The intensity of 800 cGy is the lowest dose of TBI in MAC according to some definitions [66-69], while an operational definition used by the Center for International Blood and Marrow Research and the National Marrow Donor Program identified any regimen with TBI dose of 800 cGy as RIC, if fractionated [70]. Based on data from several animal models [71, 72], a 800 cGy TBI in a single fraction in combination with thiotepa, cyclophosphamide, and rabbit ATG had been successfully used in recipients of haploidentical bone marrow and G-CSF-mobilized peripheral blood TCD HSCT, although lung toxicity was substantial [13, 73]. We decided to adopt four doses of 200 cGy TBI (800 cGy) subsequent to observing poor survival outcome of patients who received haplo-HSCT employing a conditioning regimen consisting of fractionated 1200 cGy TBI plus fludarabine and busulfan reported in our previous study [10]. In another previous report, we reported the feasibility and favorable outcomes of a RIC regimen consisting of 400 cGy TBI (with a fraction size of 200 cGy on day -1) with the same doses of fludarabine and busulfan described

in this study for AML patients with old age and/or co-mobidities [74]. Although the rationale for using the dose of fractionated 800 cGy TBI was not confirmed in preclinical tests, we ought to select an "intermediate" intensity effect from irradiation based on these clinical experiences. When fractionated dosage of 800 cGy TBI combined with cyclophosphamide (120 mg/kg) was adopted prior to unrelated HSCT in adult patients with severe aplastic anemia, we also observed relatively high OS (88.0%) [31]. Furthermore, the outcome of a very similar TBI-based conditioning regimen consisting of four 200 cGy doses of TBI (800 cGy) on days -3 and -2 with 30 mg/m2/day fludarabine daily for 4 days for AML patients was reported by the Cooperative German Transplant Study Group, showing that this RIC regimen was feasible with low NRM and preserved antileukemic activity in allogeneic HSCT from related or unrelated donors [75]. Also, the results of prospective randomized phase 3 trial comparing this conditioning regimen with standard conditioning of six doses of 200 cGy of TBI and 120 mg/kg cyclophosphamide showed a significantly lower rate of non-relapse mortality in 800 cGy TBI conditioning arm in patients aged 41-60 years, although significant difference was not found in younger patients [76]. Low NRM (12.2%) with sustained complete donor chimerisim achieved in all patients was demonstrated in this study. Therefore, although somewhat hypothesis-generating, we suggest that our unique conditioning regimen of fractionated 800 cGy TBI plus fludarabine/ busulfan followed by low-dosed ATG with standard GVHD prophylaxis is safe, and may have provided a well balanced effect between patient T-cell suppression and donor T-cell immune reconstitution through a possible mechanism of tolerable in vivo TCD in unmanipulated graft-using haploidentical PBSC transplantation.

Most importantly, the safety of our haplo-HSCT strategy was not offset by high relapse rate, demonstrated by 3-year incidence of relapse of < 20% in patients receiving transplant in remission. The 2-year relapse incidence of 16% in patients in CR1 seems much promising considering that about 53% of them were high-risk AML. Among the 52 patients in CR1, lower incidence of relapse was seen in a small number of patients (n = 12) with KIR-alloreactivity defined by ligand-ligand model [77], but without statistical difference (11.1% vs. 18.4%, P = .481; data not shown). There is a possibility that KIR-alloreactivity would work less prominently under TCR setting. On the other hand, the GVL effect of patients with cGVHD in our study may have been masked from possible but obscure (at least in our study) influence of other immune cells such as natural killer T-cells or regulatory T-cells on decreasing disease recurrence in patients without signs of cGVHD. As for effects of NIMAs and inherited paternal antigens, we observed no significant influence of either NIMA-mismatched donor on GVHD or maternal donor on relapse (data not shown). Due to the small number of patients, we cannot

currently draw any conclusion on a role of the immune cell subsets or fetomaternal microchimerism in relation to GVHD as well as GVL reaction from this study. Despite the favorable transplant outcomes in patients with remission at the time of HSCT, only one of the patients transplanted in relapsed/refractory status survived without an event. Future novel strategy is demanded for this group of patients.

In conclusion, the overall outcomes of HLA-mismatched related HSCT using unmanipulated PBSCs and our novel conditioning regimen consisting of 800cGy TBI, fludarabine and busulfan with standard GVHD prophylaxis added by ATG were favorable in terms of no primary and secondary engraftment failure and low incidence of severe GVHD with favorable PFS, prominently seen in patients with standard-risk disease status. We are currently on the way of a clinical trial to prospectively evaluate and demonstrate the efficacy of our regimen for adult patients with AML receiving allogeneic HSCT from HLA-haploidentical donor, compared to that from HLA-matched unrelated donor (#NCT01751997).

Acknowledgments

The authors acknowledge all members at Catholic Blood and Marrow Transplantation Center, particularly the house staff, for their excellent care of the patients.

Authorship

H.J.K. was the principal investigator and takes primary responsibility for the paper; S.A.Y. collected, analyzed, and interpreted data and wrote the manuscript; K.H.J. recruited and cared patients, designed the research, collected, analyzed and interpreted data, and wrote the manuscript; J.H.K, Y.W.J., J.H.Y., S.H.S., S.E.L provided patient care and commented on the manuscript; B.S.C, K.S.E., Y.J.K. provided patients care and contributed to the writing of the paper; S.L., C.K.M. contributed to data analysis; S.G.C., D.W.K., J.W.L., W.S.M. and C.W.P commented on the manuscript. All authors have read and approved the final manuscript.

Conflict-of-interest disclosure

There is no conflict-of-interest to disclosure.

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Table 1.

Patient and donor characteristics

Recipient age, years, median (range) 41(16-69)

Gender, male, No. (%) 52 (65)

Age groups, years, No. (%)

16 - 29 16 (20)

30 - 49 41 (51)

50 - 59 8 (10)

60 - 69 15 (19)

AML type, No. (%)

De novo 73 (91)

Secondary 7 (9)

Previous transplant, No. (%)

No 68 (85)

Autologous 5 (6)

Allogeneic 7 (9)

AML with myeloid sarcoma at diagnosis, No. (%) 7 (9)

Chromosomal risk group, No. (%)

Favorable 17 (21)

Intermediate 50 (63)

Adverse 13 (16)

WBC at diagnosis, x 109/L

Median (range) 6.86 (0.37-250.13)

> 50, No. (%) 22 (28)

Disease status, No. (%)

CR1 or iCR1 52 (65)

CR2 17 (21)

Relapse/ refractory 11 (14)

Serum ferritin level at transplant, ng/mL, median(range) 1122(241-7192)

> 2000, No. (%) 17 (21)

Matched HLA loci, No. (%)

4/8 54 (68)

5/8 19 (24)

6/8 7 (9)

Donor age, years, median (range) 33 (7-61)

Donor-recipient gender matching, No. (%)

Male-male 29 (36)

Male-female 17 (21)

Female-female 11 (14)

Female-male 23 (29)

ABO incompatibility, No. (%)

Match 59 (74)

Mismatch 21 (26)

Donor relationship, No. (%)

Mother -son 17 (21)

Mother -daughter 6 (7)

Son-father 24 (30)

Son-mother 15 (19)

Daughter-father 3 (4)

Daughter-mother 4 (5)

Sibling-sibling 11 (14)

KIR alloreactivity, No. (%)

Yes 20 (25) No_60 (75)

AML indicates acute myeloid leukemia; WBC, white blood cell; CR, complete remission; iCR1, CR with incomplete blood count recovery; KIR, killer immunoglobulin-like receptor.

1 Table 2.

2 Univariate analysis of donor graft cell doses on transplant outcomes: recipients with donor cell dose above the median value were compared with the others.

CD34+ CD3+ CD4+ CD8+ CD19+ CD3-CD56+ CD3+CD56+

Median (range) x 106/kg 6.5 (2.2-39.7) 498 (201-1510) 253 (102-942) 172 (51.7-643) 129 (19-447) 96.7 (16.9-366) 37.2 (3.2-177)

Grades II-IV aGVHD

HR (95% CI)1 .7 (.4-1.3) 1.8 (1.0-3.4) 1.7 (.9-3.3) 2.1 (1.1-4.1) 1.6 (.9-3.1) 1.3 (.7-3.4) .8 (.5-1.6)

P .27 .07 .09 .02 .13 .45 .58

Moderate-severe cGVHD

HR (95% CI)1 .5 (.2-1.3) 2.1 (.8-5.1) 3.6 (1.3-10) 2.6 (1.0-7.0) 1.8(.7-4.2) 2.2 (.9-5.4) 1.1 (.4-2.4)

P .16 .13 .01 .05 .20 .09 .92

CMV DNAemia

HR (95% CI)1 .7 (.4-1.2) 1.6 (.9-2.7) 1.0 (.6-1.7) 1.8 (1.0-3.1) 1.6 (.9-2.7) 1.3 (.9-2.2) 1.6 (.9-2.7)

P .20 .10 .91 .04 .09 .28 .09

HR (95% CI)1 2.0 (.9-4.5) 2.1 (.9-5.0) 1.8 (.8-4.1) 1.3 (.6-2.8) 1.0 (.4-2.1) .6 (.3-1.4) .8 (.4-1.8)

P .10 .08 .17 .58 .89 .23 .64

HR (95% CI)1 1.7 (.8-3.5) 1.6 (.7-3.2) 1.4 (.7-2.8) 1.1 (.6-2.3) .8 (.4-1.6) .6 (.3-1.2) .3 (.3-1.3)

P .16 .24 .40 .70 .50 .17 .17

Relapse

HR (95% CI)1 1.7 (.7-4.0) 1.2 (.5-3.0) 1.3 (.5-3.1) 1.0 (.4-2.4) .8 (.3-1.9) .7 (.3-1.6) .2 (.2-1.3)

P .26 .64 .55 .99 .58 .33 .15

HR (95% CI)1 1.5 (4.4-5.4) 2.3 (.6-8.8) 1.4 (.4-4.9) 1.5 (4.3-5.4) 1.0 (.3-3.2) .6 (.2-2.2) .9 (.3-3.3)

P .50 ^.22 .57 .52 .94 .48 .94

"3 aGVHD indicates acute graft-versus-host disease; HR, hazard ratio; CI, confidence interval; cGVHD, chronic graft-versus-host disease; CMV, cytomegalovirus; OS, overall survival; PFS, progression-free survival; NRM,

4 non-relapse mortality

5 ^Hazard ratio of recipients with donor cell dose above the median value compared to the others with cell dose < median.

8 Table 3.

9 Multivariate analysis for transplant outcomes.

Risk factors No./total %±SD at 2 years HR(95% CI) P

Disease status at HSCT 12

Standard 9/52 82.5% ± 5.7% 1 13

Advanced 16/28 33.6% ± 10.4% 4.35 (1.87-10.08) .001

Occurrence of aGVHD 14

No aGVHD 19/71 71.1% ± 5.9% 1 15

Grades III-IV aGVHD 6/9 29.6% ± 16.4% 2.58 (1.02-6.53) .046

Donor age (continuous variable) - - 1.02 (1.00-1.05) .098 16

Disease status at HSCT 17

Standard 13/52 75.1% ± 6.3% 1 18

Advanced 17/28 34.2% ± 10.0% 3.58 (1.68-7.63) .001

Occurrence of aGVHD 19

No aGVHD 23/71 66.6% ± 6.4% 1 20

Grades III-IV aGVHD 7/9 22.2% ± 13.9% 2.89 (1.22-6.88) .016

Donor age (continuous variable) - - 1.02 (.99-1.04) .186 ?1

Relapse Disease status at HSCT 22

Standard 8/52 16.6% ± 5.5% 1 23

Advanced 21/28 45.8% ±10.5% 4.22 (1.77-10.07) .001

WBC at diagnosis 24

< 50000 11/57 19.8% ± 5.4% 1 25

> 50000 9/23 44.1% ± 11.7% 2.69 (1.11-6.54) .029

NRM Occurrence of aGVHD 26

No aGVHD 7/71 12.4% ± 4.8% 1 27

Grades III-IV aGVHD 3/9 33.3% ± 17.7% 3.35 (.81-13.87) .096 28

Donor age (continuous variable) - - 1.03 (.99-1.06) .110

29 SD indicates standard deviation; HR, hazard ratio; HSCT, hematopoietic stem cell transplantation; OS, overall survival; aGVHD, acute graft-versus-host disease; PFS, progression-free survival; WBC, white blood cell; NRM,

30 non-relapse mortality.

Figure legends

Figure 1. Cumulative incidence of myeloid (Left) and platelet (Right) engraftment.

Figure 2. Reconstitution of lymphocyte subsets (CD8+ T-cells [CD3+CD8+], CD4+ T-cells [CD3+CD4+], CD56+ T-cells [CD3-CD56+], and B-cells [CD19+]) following T-cell replete haplo-identical hematopoietic stem cell transplantation after fractionated 800 cGy TBI-based conditioning regimen with low-dosed ATG . Curves represent means ± standard error of the mean.

Figure 3. Cumulative incidence of GVHD by donor graft cell dose. (Left) shows incidence of grades II-IV aGVHD according to graft CD8+ T-cell dose: < (dotted line) versus > (continuous line) median value of CD8+ (172 cells x 106/kg of recipient body weight). (Right) shows incidence of moderate to severe NIH cGVHD according to graft CD4+ T-cell dose: < (dotted line) versus > (continous line) median value of CD4+ (253 cells x 106/kg of recipient body weight). Figure 4. Transplantation outcomes according to disease status: Probability of (A) OS and (B) PFS and cumulative incidence of (C) relapse and (D) NRM. Shown are patients with CR1 (n = 52, continuous line), CR2 (n = 17, dotted line), or relapsed/refractory status (n = 11, dashed line) at the time of transplant.

c 11) E a a i_ u> c

c 11) E a

csj Ö

csj ö

Platelet

10 20 30

Days after transplant

10 20 30

Days after transplant

0 1 3 6 9 12

Months after transplant

Months after transplant

Months after transplant

Months after transplant

20 40 60

Days after transplant

Months after transplant

P < .0001

10 20 30 40 50 60

Months after transplant

10 20 30

P < .0001

Months after transplant

02 - .I

P < .0001

20 30 40 50

Months after transplant

P= .240

20 30 40

Months after transplant

72 Supplementary Table 1. Univariate analysis for transplant outcomes

OS PFS Relapse NRM

Risk factors HR(95% CI) P HR(95% CI) P HR(95% CI) P HR(95% CI) P

Patient age .99(.96-1.02) .596 .99(.96-1.02) .445 .98(.95-1.01) .200 1.01(.95-1.07) .710

Gender, male 1.02(.45-2.31) .958 .89(.43-1.84) .743 .85(.35-2.05) .720 .88(.26-3.02) .840

Type AML

De novo 1 1 1

Secondary 1.96(.26-14.53) .512 .84(.20-3.55) .817 1.42(.32-6.37) .650 not feasible

WBC at diagnosis

< 50000 1 1 1

> 50000 0.99(.42-2.39) .995 1.25(.58-2.67) .569 2.22(.94-5.27) .070 .26(.03-1.96) .190

Cytogenetic risk

Better 1 1 1 1

Intermediate 1.27(.42-3.83) .671 1.26(.47-3.38) .642 .88(.28-2.77) .830 2.75(.34-22) .340

Poor 2.09(.59-7.40) .255 1.78(.54-5.84) .341 1.80(.48-6.72) .380 1.31(.08-21) .850

Serum ferritin at HSCT

< 1000 1 1 1 1

> 1000 2.24(.89-5.62) .085 1.49(.70-3.18) .307 1.03(.43-2.47) .950 2.90(.63-13.3) .170

Previous HSCT

No 1 1

Yes 1.05(.36-3.07) .925 .88(.31 -2.51) .805 not feasible not feasible

Disease status at HSCT

Standard 1 1 1 1

Advanced 5.04(2.20-11.52) < .0001 3.90(1.87-8.12) < .0001 3.74(1.56-8.93) .003 2.15(.65-7.12) .210

Donor age 1.03(1.01-1.06) .018 1.03(1.00-1.05) .029 1.02(.99-1.05) .210 1.03(1.00-1.07) .054

Gender matching

Others 1 1 1 1

Female-male 1.51(.67-3.43) .322 1.18(.54-2.58) .676 .87(.31-2.43) .780 1.72(.50-5.87) .390

ABO matching

Match 1 1 1 1

Mismatch .98(.39-2.46) .961 1.08(.48-2.43) .858 1.27(.50-3.24) .610 .75(.16-3.40) .710

Relation of donor

Child 1 1 1 1

Sibling 1.51 (.48-4.74) .482 1.23(.41-3.75) .712 1.43(.41-5.03) .580 .81(.09-7.19) .850

Parent 2.23(.95-5.27) .067 2.12(.98-4.8) .057 2.14(.83-5.53) .120 1.63(.46-5.77) .450

HLA matching

5-6 loci 1 1 1 1

4 loci .92(.41-2.09) .846 1.02(.48-2.18) .956 1. 13(.43 -3.01) .800 1.25(.49-3. 17) .640

KIR alloreactivity

Yes No

Occurrence of aGVHD Grades 0-II Grades III-IV Occurrence of cGVHD None-mild Moderate-severe

.76(.32-1.82) .536 .43-2.32 .988 1.36(.46-3.99) .570

3.19(1.26-8.01) .014 3.10(1.33-7.26) .009 2.02(.73-5.60) .180

.70(.28-1.76) .449_.75(.27-2.06) .574_0.85 (.32-2.25) .740

.68(.18-2.63) 1

3.71(.99-14.0) 1

.58 (.14-2.33)

77 .440

OS indicates overall survival; PFS, progression-free survival; NRM, non-relapse mortality; HR, hazard ratio; CI, confidence interval; AML, acute myeloid leukemia; WBC, white blood cell; HSCT, hematopoietic stem cell transplantation; KIR, killer-like immunoglobulin receptor; aGVHD, acute graft-versus-host disease; cGVHD, chronic graft-versus-host disease.

Highlights

1) We analyze the outcomes of T cell-replete haploidentical transplantation in AML.

2) The conditioning regimen is based on a fractionated 800 cGy total body irradiation.

3) All patients achieved prompt and sustained neutrophil and platelet engraftment.

4) Favorable outcome was seen in patients with first complete remission at transplant.