Sibling Stem Cell Transplantation Research Articles

More than two courses of pre-transplant consolidation therapy benefits patients with acute myeloid leukemia in the first complete remission who underwent human leukocyte antigen-matched sibling allografts: a multicenter study.

Although the need for consolidation chemotherapy after successful induction therapy is well established in patients with acute myeloid leukemia (AML) in first complete remission (CR1), the value of consolidation chemotherapy before allogeneic hematopoietic stem cell transplantation remains controversial. We retrospectively compared the effect of the number of pre-transplant consolidation chemotherapies on outcomes of human leukocyte antigen-matched sibling stem cell transplantation (MSDT) for patients with AML in CR1 in multicenters across China. In our study, we analyzed data of 373 AML patients in CR1 from three centers across China. With a median follow-up of 969 days, patients with ≥ 3 courses of consolidation chemotherapy had higher probabilities of leukemia-free survival (LFS) (85.6% vs . 67.0%, P < 0.001) and overall survival (89.2% vs . 78.5%, P = 0.007), and better cumulative incidences of relapse (10.5% vs . 19.6%, P = 0.020) and non-relapse mortality (4.2% vs . 14.9%, P = 0.001) than those with ≤ 2 courses of consolidation chemotherapy. Pre-transplantation minimal residual disease-negative patients with AML in CR1 who received MSDT with ≥ 3 courses of consolidation chemotherapy had a higher probability of LFS (85.9% vs . 67.7%, P = 0.003) and a lower cumulative incidence of relapse (9.6% vs . 23.3%, P = 0.013) than those with ≤ 2 courses. Our results indicate that patients with AML in CR1 who received MSDT might benefit from pre-transplant consolidation chemotherapy.

The Comparision of Immune Reconstitution after HLA-Haploidentical Stem Cell Transplantation and HLA-Identical Sibling Stem Cell Transplantation

Title: The comparision of immune reconstitution after HLA-haploidentical Stem Cell Transplantation and HLA-identical sibling Stem Cell Transplantation.Background: HLA-haploidentical Stem Cell Transplantation is widely used in allogeneic hematopoietic stem cell transplantation in recent years. Previous studties reported that HLA-haploidentical Stem Cell Transplantation(HRD) has comparable GVHD and similar overall survival (OS) to HLA-identical sibling stem cell transplantation(MSD). While the immune reconstruction after transplantation was less studied and the effect of early and successful immunologic reconstitution on transplantation outcome still need further research.Methods: A total of 162 patients with hematologic malignancies who underwent allo-HSCT at Nanfang Hospital between January 2013 and December 2015 were enrolled in the retrospective study. Of the 162patients, 52 cases received HRD, 110 MSD. We sequentially monitored the reconstitution kinetics of CD3+T cells, total CD4+T cells, CD4+CD45RA+ T cells, CD4+CD45RO+ T cells, CD8+ T cells, CD8+CD45RA+ T cells, CD4+CD25+FOXP3+Treg cells, CD19+ B cells, CD3-CD56+ NK cells in all patients by flow cytometry.Results: Our results showed that in the first 30 days after grafting, counts of T cell subsets (total CD4+ T cells, CD45RA+CD4+ T cells, CD45RO+CD4+ T cells, total CD8+ T cells, CD45RA+CD8+ T cells,CD4+CD25+FOXP3+Treg cells) and CD3-CD56+ NK cell subsets were all signifcantly lower in haploidentical transplant recipients than those of HLA-matched transplant recipients, though no sigificant difference in the counts of B cells in two groups. The total CD8+ T cell numbers expanded dramatically and reach the level of that of HLA-identical recipients at day 90 post transplantation, though CD45RA+CD8+ T cells was still signifcantly lower in haploidentical transplant recipients than those of HLA-matched transplant recipients. Since day 180 post transplantation, CD8+ T cells were signifciantly higher than that of HLA-identical recipients.Conclusion: HLA-haploidentical Stem Cell Transplantation show superior CD8+ T cells and NK cells reconstitution than HLA-identical sibling Stem Cell Transplantation, which might be associated with a superior GVL effect compared to MSD for high-risk patients.similar immune reconstitution as HLA-identical sibling Stem Cell Transplantation. DisclosuresNo relevant conflicts of interest to declare.

Safety and Efficacy of Ex Vivo Expanded Healthy Donor-Derived Double Negative T Cells for the Treatment of AML Relapsed after Allogeneic Stem Cell Transplantation: A First in-Human Phase I/IIa Clinical Trial

BACKGROUND Relapse after allogeneic hematopoietic stem cell transplantation (allo-HSCT) remains the main cause of treatment failure for patients with acute myeloid leukemia (AML). Conventional treatments such as donor lymphocyte infusions (DLI) or secondary allo-HSCT as a monotherapy or in combination with novel agents or immunotherapy showed limited improvements on long-term survival. Our previous studies have shown that a rare subset of CD4- and CD8- double negative T cells (DNTs) expanded from periphery of healthy donors are effective in targeting AML cells in vitro and in xenograft models. Also, allogeneic DNTs can be used as an off-the-shelf adoptive cellular therapy (ACT) for AML. We undertook a first in-human phase I/IIa trial to assess the safety and efficacy of escalating doses of DNTs for the treatment of AML patients relapsed post allo-HSCT. This study protocol was approved by Chinese Ethics Committee of Registering Clinical Trials. Informed consent was obtained in accordance with the Declaration of Helsinki. The trial was registered on the Chinese Clinical Trial Registry (ChiCTR-IPR-1900022795). METHODS From June 2019 to June 2020, 12 patients with relapsed AML after allo-HSCT were enrolled in the trials. Three patients received HLA-matched sibling stem cell transplant, and 9 patients received umbilical cord blood transplantation (UCBT). Hematological relapse was found in 9 cases, and molecular relapsed in 3 cases. None of the patients were able to obtain DLI from previous donors for various reasons. Clinical-grade DNTs from peripheral blood of healthy donors were expanded. After lymphodepleting chemotherapy with fludarabine and cyclophosphamide, each patient received three infusions of HLA-mismatched DNTs expanded from healthy volunteers at escalating doses of 5×107, 1×108, or 2-4×108 per kilogram of body weight at one week interval. Numbers of total DNTs and donor-derived DNTs, inflammatory cytokines levels in vivo were measured according to the study protocol. Post-remission therapy was permitted 14 days after the last DNTs infusion according to treating physician's discretion. RESULTS All surviving patients were followed until August 1st, 2020. At a median follow-up of 4.3 months (range 1.7 to 12.2 months), 1 patient withdrew from the trial after the second DNT infusion for a personal reason. 6/11 patients (54.5%) achieved complete response (CR) with 5 of these patients negative for (45.4%) minimal/measurable residual disease (MRD). 4/11 patients (36.3%) achieved partial remission. Up to the last follow-up, 4/11 patients remain CR. Increase in the levels of serum inflammatory cytokines, including interleukin (IL)-6, TNF-alpha, MCP-1, and MIP-1-beta over baseline were observed without signs of cytokine release syndrome, neurotoxicity, or graft-versus-host disease. The most common adverse events related to DNTs treatment were fever and headache. None of the patients showed &amp;gt;grade 2 adverse event (AE). The maximum tolerated dose was not reached. Donor DNT cell expansion was detected as early as 6 hours after infusion and reached the peak at 24 hours after the 1st infusion in most patients, slightly increased after the 2nd and 3rd infusions, and lasted for 2 weeks after 3rd infusion. However, total DNTs increased and reached the peak at 2 weeks after 3rd infusion. CONCLUSIONS Our results show that allogeneic DNTs therapy is safe, well tolerated, and potentially effective in treating AML patients who relapsed after allo-HSCT. The total number of DNTs increased after infusion. Compared to other agents, DNTs more effectively induced complete responses in this patient population. Our on-going study is to elucidate the mechanism of DNTs-mediated leukemia clearance, and explore the use of DNTs as carrier for Chimeric Antigen Receptor or transgenic TCR to further enhance the efficacy targeting resistant AML cells. Disclosures Zhang: AbbVie: Current equity holder in publicly-traded company; University Health Network: Consultancy, Current equity holder in private company, Other: Principal Investigator, Patents &amp; Royalties.

Non-myeloablative matched sibling stem cell transplantation with the optional reinforced stem cell infusion for patients with hemoglobinopathies.

The NIH protocol for non-myeloablative (NMA) conditioning allogeneic stem cell transplantation (alloSCT) with alemtuzumab and low-dose total body irradiation corrected the abnormal sickle cell disease (SCD) phenotype without the risk of graft-versus-host disease. However, alloSCT using NMA conditioning had been rarely applied to β-thalassemia major (β-TM) patients. To avoid prolonged immunosuppression, we developed a two-stage strategy. Mixed donor chimerism was initially achieved using the protocol developed by the NIH protocol. Thereafter, we facilitated donor chimerism using the optional reinforced stem cell (SC) infusion in cases requiring protracted immunosuppression or experiencing impending graft failure. In this study, β-TM (n=9) and SCD (n=4) patients were equally effectively treated with eradicating the abnormal hemoglobin phenotype. Five patients, including four β-TM, achieved stable mixed chimerism without receiving optional reinforced SC infusion. All patients that received optional reinforced infusion achieved complete (n=4) or mixed chimerism (n=1). The overall survival rate and event-free survival at 4years were 91.7% (95% CI; 53.9-98.8) in both groups, with a thalassemia-free survival rate in β-TM patients of 87.5% (95% CI; 38.7-98.1). This study is the first to report successful NMA conditioning alloSCT to achieve stable mixed chimerism correcting the abnormal hemoglobin phenotype in adult β-TM patients.

Post-transplant Hemophagocytic Lymphohistiocytosis in Benign Hematological Disorders: Experience of 4 Cases with Review of Literature.

Post transplant Hemophagocytic lymphohistiocytosis (HLH) is a form of secondary HLH, which can be either early onset or late onset and is associated with significant morbidity and mortality. With the increasing popularity of post transplant cyclophosphamide based haploidentical stem cell transplantation (SCT), post transplant HLH is becoming a significant complication especially in benign hematological disorders. Methods: We present 4 cases of post transplant HLH occurring in 2 cases of severe aplastic anemia (post haploidentical SCT) and 2 cases of thalassemia major (post matched sibling SCT). All 4 cases had early onset variety with dismal prognosis. Conclusion: Post-transplant HLH is an important entity in benign hematological disorders, which needs to be identified early and treated promptly with steroids, monoclonal agents or immunosuppressive therapy. Serum ferritin levels are an important biomarker and help in monitoring response.

Survival Outcomes in Patients with Intermediate Risk Acute Myeloid Leukemia: A Single-Center Retrospective Review

Objectives Acute Myeloid leukemia (AML) is a heterogeneous disease with high morbidity and mortality. These patients can be stratified into favorable, intermediate and adverse-risk groups based on their cytogenetic and molecular profile. We analyzed the outcomes of the intermediate risk patients in our center to evaluate the results and to report the differences between the intensity of the treatments, chemotherapy alone versus allogeneic stem cell transplantation (SCT). Methods A total of 53 patients diagnosed in Hospital Universitari Son Espases between 2012 and 2019 with AML, classified as intermediate risk disease (based on ELN 2017 criteria), and candidates to intensive treatment were analyzed. Results The median age at diagnosis was 61 years (range 17-74), 28 males and 25 females. Regarding the diagnosis, almost 50% were AML with mutated NPM1, AML with myelodysplasia-related changes and AML therapy-related. Thirty percent of the patients had FLT3-ITD mutation (78.6% of them with high ratio). Twenty eight percent had NPM1 mutation. The 75.5% received cytarabine and anthracycline induction chemotherapy and the remaining 24.5% received a combination of cytarabine, anthracycline plus etoposide. Twenty percent of the patients (11) died during the induction therapy, the main of them due to infections. Of the 42 patients alive, 36 (85.7%) achieved a complete remission, 4 a partial response (9.5 %) and 2 patients (4.8%) were refractory. All of the patients went under consolidation: 39 (73.5%) with chemotherapy alone and 14 (33 %) with SCT (4 of them immediately after salvage therapy without hematopoietic recovery). Three patients received a HLA identical sibling SCT (21.4%), 3 a matched unrelated donor (21.4%), 5 (35.7%) an haploidentical SCT, 1 patient (7.1%) a cord blood stem cell transplantation and 2 patients an autologous SCT (14.3%). The median follow-up of the whole group of patients was 22 months (2.6-86). The median overall survival (OS) was 18 months (CI 95%: 10-25) and the median progression free survival (PFS) was 9 months (CI 95%: 4- 14). Patients who went under transplantation achieved a PFS of 18.5 months compared with 7.8 months in no transplantation patients (p=0.159). Patients with non-mutated FLT3-ITD have a median PFS of 14 months compared with 7 months in patients with mutated FLT3-ITD (p=0.333). Excluding the patients who died during induction therapy, 11 patients with mutated FLT3-ITD went into consolidation therapy. Three received SCT and 8 chemotherapy alone. The median OS of the SCT group was 18.5 versus 13.6 months in the chemotherapy group (CI 95% 4.3-22.9; p=0.4). The median PFS of the SCT group was 6.17 months (CI 95% 2.06-10.3), similar to the chemotherapy group where was 7.6 months (CI 95% 5.03-10.66; p=0.7). Twenty-eight patients with non-mutated FLT3-ITD received consolidation therapy, 6 of them went into SCT. OS in the transplant group was 46.4 months (CI 95%: 44.6-48.2; p= 0.4) compared with 35.3 months (CI 95%: 11.7- 58.3; p=0.4) in the non-transplantation group. PFS of transplant group was 18.1 month (CI 95%: 9.5-26.22) superior to PFS in chemotherapy group that was 9.4 months (CI 4.65-14; p=0.7). Patients with mutated FLT3-ITD who did not received an FLT3 tyrosine kinase inhibitor (TKI) had a PFS of 6 months (0-13.8) compared with those that received an FLT3 TKI who did not reach the median survival rate. Conclusions Our outcomes are similar to the ones described in the literature. We confirm that patients with intermediate risk who underwent transplantation have better outcomes that the ones treated with chemotherapy alone. Moreover, we confirm that the prognosis is better in patients with non-mutated FLT3-ITD. SCT increase the OS in FLT 3-ITD mutated patients, although the PFS is similar in both groups (SCT vs chemotherapy), taking into account the small size of our sample and the short follow up period in many patients. The better outcome observed in FLT3 tyrosine kinase inhibitor treated patients indicates that this can be a promising therapy in AML patients. Disclosures No relevant conflicts of interest to declare.

HLA-Matched Sibling Stem Cell Transplantation Using Post-Transplantation Cyclophosphamide as Graft-Versus-Host Disease Prophylaxis: A Single Center Experience

HLA-Matched Sibling Stem Cell Transplantation Using Post-Transplantation Cyclophosphamide as Graft-Versus-Host Disease Prophylaxis: A Single Center Experience

Effects of two doses of anti-T lymphocyte globulin-Fresenius given after full-match sibling stem cell transplantation in acute myeloblastic leukemia patients who underwent myeloablative fludarabine/busulfan conditioning

Objective/backgroundAnti-T lymphocyte globulin Fresenius (rATG-F; ATG-Fresenius) and antithymocyte globulin (thymoglobulin), which are included in transplant protocols, are used to reduce the risk of chronic graft-versus-host disease (cGVHD) or suppress allograft rejection. Available clinical studies have been conducted in heterogenous patient populations and with different administration protocols including stem cell sources. Additionally, the pharmacokinetics of ATG is variable, and the clinically effective dose of rATG-F, in particular, is not exactly known. The aim of the study was to investigate the clinical outcomes of acute myeloid leukemia (AML) patients who underwent hemopoietic peripheral stem cell transplantation from full-matched sibling donors and given two different doses of r-ATG-F. MethodsThis was a single-center, retrospective chart review conducted between July 2005 and July 2016. Sixty-nine consecutive AML patients who underwent transplant with fludarabine- and busulfan-based conditioning were included in the study. Patients in Group 1 received 15 mg/kg body weight rATG-F to 2013 (n = 46), and Group 2 received 30 mg/kg of rATG-F dose begining in 2013 to reduce to cGVHD (n = 23). Cyclosporine and methotrexate were used to treat acute GVHD (aGVHD) prophylaxis. Outcome parameters were compared between the groups. ResultsAlthough the recommended dose r-ATG-F had led to a decrease in the cumulative incidence of cGVHD (27 [58.7%] vs. 8 [34.8%]; p = .03), it also increased the infection rate at 1 year (3 [6.5%] vs. 4 [17.4%]; p = .02). The two groups were similar in terms of engraftment time, aGVHD, relapse, nonrelapse mortality, and rATG-F-related toxicity. A Cox regression model revealed that aGVHD III–IV was associated with increased nonrelapse mortality at 1 year (hazard ratio = 18.2; 95% confidence interval, 1.667–199.255; p = <.02). No patients developed rATG-F-related severe adverse events (Common Terminology Criteria grade 4 or 5). ConclusionDose difference of rATG-F did not influence survival parameters; however, increasing the dose to 30 mg/kg seems to be effective for reducing cGVHD with an increase in infection rate requiring close monitoring of infections in AML patients who received myeloablative fludarabine/busulfan conditioning.

Transplantation in patients with iron overload: is there a place for magnetic resonance imaging? : Transplantation in iron overload.

In iron overload diseases (thalassemia, sickle cell, and myelodysplastic syndrome), iron is deposited in all internal organs, leading to functional abnormalities. Hematopoietic stem cell transplantation (HSCT) is the only treatment offering a potential cure in these diseases. Our aim was to describe the experience in the field and the role of magnetic resonance imaging in the evaluation of iron overload before and after HSCT. Magnetic resonance imaging (MRI), using T2*, is the most commonly used tool to diagnose myocardial-liver iron overload and guide tailored treatment. Currently, HSCT offers complete cure in thalassemia major, after overcoming the immunologic barrier, and should be considered for all patients who have a suitable donor. The overall thalassemia-free survival of low-risk, HLA-matched sibling stem cell transplantation patients is 85-90%, with a 95% overall survival. The problems of rejection and engraftment are improving with the use of adequate immunosuppression. However, a detailed iron assessment of both heart and liver is necessary for pre- and post-transplant evaluation. In iron overload diseases, heart and liver iron evaluation is indispensable not only for the patients' survival, but also for evaluation before and after HSCT.

Multiple Brain Hemorrhages on Presentation in a Patient with Accelerated Phase Chronic Myeloid Leukemia and Subsequent CNS Blast Crisis Despite Systemic Complete Molecular Response: Possible Role of CNS Bleed in Leukemia Proliferation

BackgroundChronic myeloid leukemia (CML) is a myeloproliferative disorder which can present in one of the three disease phases; chronic phase, accelerated phase or blast crisis. Central nervous system (CNS) as a site of extramedullary blast crisis in CML is rare and when affected, it usually occurs concurrently with systemic relapse. In extremely rare cases, the blast phase can affect the CNS without bone marrow involvement. Tyrosine kinase inhibitors (TKI) have become the treatment of choice for CML. However, there is poor penetration of CNS by some of these drugs or their active metabolites. Therefore, CNS may act as a sanctuary site for malignant cells in CML patients treated with TKIs. Here we describe an unusual case of CML who presented in accelerated-phase and multiple intracranial hemorrhages, and developed isolated CNS blast crisis after one year in spite of being in systemic remission.Clinical presentationA previously healthy 27-years-old male presented with10 days history of fatigue, abdominal pain and vomiting. He also complained of weight loss and low grade fever. Abdominal examination revealed huge splenomegaly. Investigations showed a white cell count of 538Í 109/l, hemoglobin 7.2 g/dl and platelets 47Í 109/l. Blood film showed marked leukocytosis with left shift, blasts 9% and thrombocytopenia, consistent with accelerated-phase CML. A bone marrow (BM) aspirate confirmed the above findings. Cytogenetic analysis of the bone marrow revealed100% Philadelphia chromosome and BCR/ABL positivity.After admission, the patient developed headache and vomiting. MRI of brain showed multiple small hemorrhages in the supratentorial and infratentorial compartments (figure). The patient was diagnosed with accelerated-phase CML and leukostasis. Emergency leukapheresis was performed (two sessions) resulting in a decrease in his WBC count.He was also started on hydroxurea, nilotinib and cytarabine 100mg, which led to improvement of his general condition and control of leukocyte count. He was continued on nilotinib 400 mg twice daily. He achieved complete molecular remission. Brain radiotherapy was considered because of brain hemorrhages and fear of lodging of blasts in the brain but not given because of opinion against it by expert colleagues. He was also referred for allogeneic stem-cell-transplant (SCT) because of the above concerns but not considered because of excellent response to nilotinib.After 1year of initial diagnosis, he was readmitted with headache and bilateral visional loss. Fundoscopy showed retinal hemorrhages. His CBC, BM biopsy and brain MRI were normal. Lumbar puncture showed markedly increased CSF pressure (55cm) and leukocyte infiltration with many blast cells (42%).Patient was diagnosed as CNS blast crisis of CML with BM in molecular remission. He was stareted on intrathecal chemotherapy (ITC), triple regime (methotrexate 6mg, ARA-C 30 mg, hydrocortisone 30 mg) and nilotinib was changed to dasatinib. His CSF cleared after multiple sessions of ITC; unfortunately, significant visual impairment persisted. Patient also received radiotherapy to the brain. He went on to have a sibling SCT and remains well 8 months post transplant.ConclusionWe describe a young male with accelerated-phase CML and multiple brain hemorrhages on presentation and subsequent isolated CNS blast crisis. We speculate that there was a possible link between the initial brain hemorrhages causing lodging and proliferation of blasts in the brain and subsequent isolated CNS blast crisis. We suggest that treatment options in such cases should include consideration of cranial irradiation initially, a tyrosine kinase inhibitor with better CNS penetration like dasatinib, and upfront allogeneic SCT as potential therapeutic modalities. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

High incidence of engraftment syndrome after haploidentical allogeneic stem cell transplantation.

We determined the incidence, clinical manifestations, and outcomes of engraftment syndrome (ES) in haploidentical stem cell transplantation (SCT) recipients. We compared the incidence of ES between the patient group that received haploidentical SCT (n=516) and the patient group that received HLA-identical sibling SCT (n=393). The transplantations were performed in the Peking University People's Hospital in the period between October 2001 and October 2012. The ES incidence data were collected retrospectively. Patients that presented non-infectious fever or skin rash within the 24-h window before or after the beginning of neutrophil recovery were diagnosed with ES in accordance with the Maiolino criteria. ES incidence in haploidentical SCT recipients (21.9%) was significantly higher than that in HLA-identical sibling SCT recipients (2.0%; P<0.001). Major symptoms included fever (119/121, 98.3%), skin rash (98/121, 81.0%), and diarrhea (51/121, 42.1%), with the median time of +10d (range: 6-20d). The median C-reactive protein level of the ES group (99.0mg/L; n=13) was significantly higher than that of the non-ES group (13.9mg/L; n=38; P<0.001). Similarly, the results showed that the median C3 plasma concentration of the ES group (1.30g/L) was higher than that of the non-ES group (1.16g/L, P=0.003). ES was not associated with non-relapse mortality or overall survival. High incidence of ES was observed in haploidentical SCT recipients; however, ES did not predict poor clinical outcomes.

Differences in Natural Killer(NK) Reconstitution Between Unmanipulated Haploidentical and HLA Identical Stem Cell Transplantation and Relationship with Citomegalovirus and Graft Versus Host Disease (GVHD). Experience in One Centre in 22 Patients.

Introduction: It´s well known that NK cells exert an important role in postransplant immune reconstitution. Their maturation process through the contact with recipient´s bone marrow stromal cells is crucial for the development of their functional capacities. Previous studies showed the relevance of NK alloreactivity in relapse prevention and the influence of NK subpopulations and expression of activating receptors on CMV infection control, antitumor effects, and graft versus host disease (GVHD). Unmanipulated Haploidentical (Haplo) transplantation with postrasplant cyclophosphamide (Cy) is an emerging alternative for patients (pts) with no identical sibling donor with encouraging results. We previously reported preliminary data of NK cell reconstitution in 7 pts in Haplo setting. Our aim is to contrast our previous data in 22 pts comparing with conventional HLA-identical (HLAid) Stem Cell Transplantation (SCT) and to analyse the relationship between NK reconstitution and CMV reactivation and GVHD.Methods: 22 pts received an Haplo SCT and 7 pts an HLAid SCT in Gregorio Marañon Hospital between January 2013 and April 2014. Peripheral blood was used in all cases. Conditioning regimen comprised fludarabine (Flu), Cy and busulfan (Bux) for Haplo and Flu/Bux or Flu/Melphalan for HLAid SCT. Prophylaxis for GVHD consisted of high dose Cy on +3, +4, cyclosporine (CsA) A and mycophenolatemofetil for Haplo and CsA/Methotrexate for HLAid. Table1 show patients characteristics. For analysis of NK reconstitution and receptor expression multi-colour flow cytometry on FC500 and Navios Beckman Coulter® was used. Total NK cells, CD56 intensity and expression of NKG2A, NKp30, Nkp46 and NKG2D was studied at +15, +30, +60, and +90. For comparison between the two groups Mann–Whitney U-test was used.Results: Median NK cells/mm3 on +15 and +30 were significantly lower on Haplo than HLAid group ( 0 (0-5.5) and 28.5 (12.2-96.5) vs 69.5 (55-134) and 276 (151-422); p<0.05), being similar on + 60 and +90 (86 (49.5-158) and 147 (85-204.5) for Haplo vs 105 (39.2-162.5) and 117 (82-197) for HLAid; p ns). In Haplo pts NK cells reached normal levels on +60, while only 6/22 (27%) on +30. As previously reported NK high percentages of immature CD56bright (CD56b) cells and high expression of immature receptor NKG2A appears on +30, decreasing on +60 and +90 (CD56b 80% (72.5-88.5), NKG2A 88% (80-94) on +30, 39% (25-49) and 73% (62.2-86.7) on +60, 34% (23-43) and 69% (54.7-77.5) on +90. Compared with HLAid patients, the percentage of NKb cells was significantly higher (p<0.05) on Haplo group on +30, +60 and +90 and expression of NKG2A was also higher on +15, +30 and +60 (p<0.05). Interestingly, expression of activating receptors NKP46 and NKG2D linked to NK maturation are only significantly decreased in Haplo group comparing with HLAid on +15 (p<0.05), with appropriate expression and no differences on +30, +60 and +90. In fact, expression of NKP46 tended to be higher in Haplo on +30 (81% (62.5-90) vs 61% (42-71)), with no statistical significance (p 0.06). NKP30 had a low expression, similar in the 2 groups. In Haplo pts, no differences were found in total NK cell between pts with and without CMV reactivation at any time. Patients with acute GVHD had lower NK cells/mm3 at +30 (21.5 (10.7-35) vs 122 (21.7-260.7) and +60 (75(46-137) vs 156 (51-258); p 0.1), but statistical significance was not reached. Proportion of CD56b and NKG2A expression did not show any difference in pts regarding presence of CMV reactivation or GVHD.Conclusions: After Haplo SCT with postransplant Cy NK cell reconstitution seem to be delayed comparing with HLAid sibling SCT, reaching normal levels at +60. As previously reported, we confirm in our series that NK maturation also show a different pattern. In Haplo setting NK cells remains in an immature phenotype with high proportion of NK CD56b and expression of NKG2A much longer than in HLAid pts. Interestingly activating receptors expression, normally linked to maturation is only lower in Haplo pts in +15, but it is similar in both SCT types by day +30 and later, what should translate in appropriate cytotoxic capacity. In Haplo group, those patients with and without CMV reactivation had similar NK cell reconstitution. Patients with GVHD seem to have an impaired early NK reconstitution, but this fact needs to be contrasted.Acknowledgments: Fundación Mutua Madrileña. [Display omitted] [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Life Threatening Hyperhemolysis in Two Children with Beta-Thalassemia Major (β-TM) Treated with Stem Cell Transplantation (SCT)

Hyperhaemolysis is a rare, poorly understood complication of red cell transfusion. We report the outcomes of SCT in 2 boys of Middle Eastern origin who developed life-threatening hyperhaemolysis each following their third transfusion for β-TM at the ages of 2.5 and 4 years. The diagnosis of hyperhaemolysis was based on laboratory evidence for haemolysis, post-transfusion haemoglobin (Hb) levels lower than pre-transfusion, positive Coomb’s tests and no allo-antibody able to be identified. Haemolysis was intravascular, extravascular and severe. Precipitous drops in Hb made transfusion unavoidable. The first boy responded to prednisolone, intravenous immunoglobulin (IVIG) and splenectomy. Haemosidderosis and fibrosis were present on liver biopsy, he was therefore regarded as a Class 3 thalassaemia patient and received hydroxyurea, azathioprine, erythropoietin, desferrioxamine, busulfan, cyclophosphamide and fludarabine conditioning prior to an HLA identical sibling SCT. He is alive and well 7 years post BMT with 30% stable donor chimerism and a normal Hb. The second child’s hyperhaemolysis failed to respond to prednisolone, IVIG, rituximab and splenectomy. Provision of the large number of suitably matched red cell units required was problematic. After receiving hydroxyurea, azathioprine, desferrioxamine, busulfan, cyclophosphamide, fludarabine, thiotepa and antithymocyte globulin (ATG) preparation for a CD3/CD19 depleted maternal haplotype peripheral blood SCT, the haemolysis finally stopped. Post-transplant severe veno-occlusive disease and multi-organ failure (MOF) required dialysis and ventilation. The maternal graft was rejected so 28 days after the first transplant he received a 4/6 mismatched unrelated cord blood transplant (UCBT) following further fludarabine and ATG, which fully engrafted. He recovered from MOF and was discharged from hospital 47 days after the UCBT, transfusion independent. On day +86 he contracted Respiratory Syncytial Virus chest infection with acute intravascular haemolysis necessitating transfusions. Fulminant liver failure developed, presumably due to iron toxicity, and death occurred on day +102, having received 112 transfusions in the 12 months since presentation. In conclusion, avoiding red cell transfusion is not always possible in hyperhaemolysis, especially in β-TM. Patients may quickly become classified as Class 3 in terms of predicting BMT outcome. Immune modulation therapy and SCT was effective in 1 case but only temporarily stopped haemolysis in the other, despite full engraftment ultimately being achieved with a mismatched UCBT. SCT should be considered early in cases of hyperhaemolysis in β-TM because it can potentially cure both and result in transfusion independence. DisclosuresOff Label Use: Intravenous immunoglobulin and rituximab for treatment of haemolytic anaemia; hydroxyurea, azathioprine, fludarabine, erythropoeitin, busulphan, cyclophosphamide, thiotepa and antithymocyte globulin for use in stem cell transplantation in children with thalassaemia.

HLA- Matched Sibling Stem Cell Transplantation in Children with Thalassemia Major with ATG As Part of Preperative Regimen: A Suitable Choice Even for High Risk Cases

▪Background:Myeloablative chemotherapy using busulfan(BU) & cyclophosphamide(CY) is a standard form of conditioning for thalassemia transplantation. But this is associated with substantial incidence of graft rejection. This is much more common in high risk patients .Methodology:This study retrospectively analyses the outcome of 50 (34 boys & 16 girls) consecutive children, who underwent HLA matched stem cell transplantation from family donors at our centre All patients were classified into 3 risk groups on the basis of inadequate chelation, hepatomegaly and the presence of portal fibrosis in the liver. Pesaro Class 1 patients had none of these, class 2 patients had one or two and class 3 had all three.30 patients received oral BU 1 mg/kg, Q6H for 4 days followed by CY 60 mg/kg/D for two days. & rabbit ATG (2.5 mg/kg/day from Day-5 to Day-1). Patients with advanced disease (class 3) received BU 0.8mg/Kg & CY 50 mg/kg. with same dose of ATG. All patients received stem cells harvested from G-mobilized bone marrow Standard MTX & Cyclosporin based GVHD prophylaxis , infection prophylaxis & supportive care as per standard protocols were given as per standard protocols. Chimerism was routinely monitored at 30, 60, 90, 120 and 365 days post transplant using PCR-based analysis of selected polymorphic STRs loci.Results :OS and thalassemia-free-survival were estimated according to the Kaplan-Meier method. Probabilities of rejection, GVHD and infections were calculated by using the cumulative incidence estimator. The impact of the following parameters on OS and TFS was examined: (1) age of the recipient (above vs below the median value) (2) serum ferritin (above vs, the median value) (3) Pesaro class, (4) total nucleated cells per kg (contained in the graft) (above vs below the median), (5) CD34+ cells contained in the graft (above vs below the median) and (6) BU (oral vs i.v. BU). Both univariate and multivariate analysis of prognostic factors were made according to the log-rank test and to a stepwise Cox proportional hazards regression model, respectively. With a median follow-up of 11 years (range 1—17 years), the overall survival (OS) and thalassemia free survival (TFS) rates were 94% and 90%, respectively. The estimated TRM was 4% while the cumulative incidence of rejection/failure were 2%. One patient ( Class III, & HIV infection) died of severe veno-occlusive disease. ,The patient who rejected the graft after 10 months, received II transplant & is transfusion free with 98% donor chimerism post second transplant. The cumulative incidences of acute GVHD grade II & grade III were 15% & 8 %, respectively. 1 patient developed acute GVHD grade IV. None of the patients developed chronic GVHD. The estimated OS and TFS for patients with Class 1 and 2 disease according to Pesaro criteria were 94% and 92%, whereas for patients with Class 3 disease they were 90% and 85%, respectively.Conclusions: Myeloablative( BU-CY) conditioning with ATG as conditioning from matched sibling donors with innovative modifications can be used to achieve excellent survival , with a low incidence of TRM and graft rejection in low as well as high risk patients. DisclosuresNo relevant conflicts of interest to declare.

Umbilical Cord Blood Transplantation in Hematologic Diseases in Patients Over 15 Years Old: Long-term Experience at the Pontificia Universidad Católica de Chile

Most patients who require a sibling stem cell transplantation do not have a matched donor. In our experience, only 1/3 patients have a matched unrelated donor (MUD); therefore, the majority of the patients will require umbilical cord blood (UCB). Patients treated for hematologic diseases with UCB transplants were included. UCB selection and conditioning regimens were performed according to the Minnesota group. Graft-versus-host disease (GVHD) prophylaxis, infection prevention, and patient care were performed according to institutional guidelines. We analyzed patients and graft demography, neutrophil and platelet recovery, chimerism kinetics, GVHD incidence, overall (OS), progression-free survival (PFS) and transplant-related mortality (TRM). We included 29 patients with a median age of 34.8 years (range 15–55). Eighteen were male and the median weight was 72.6 kg (range 54–100). Nineteen patients had acute leukemia. Myeloablative (MA) conditioning was used in 27 patients. Seventeen received double UCB (DUCB) grafts. Median total nucleated cell (107/kg) was 4.2 (range 3.9–4.9) and 4.4 (range 2.8–6.3) for single UCB (SUCB) and DUCB transplants, respectively. Median time for neutrophil engraftment was 24.7 (range 14–43) and 25.8 days (range 14–52) after SUCB and DUCB transplants, respectively. Median time for platelet engraftment was 147 (range 30–516) and 81 days (range 37–200) after SUCB and DUCB transplants, respectively. All the patients receiving MA conditioning had >95% chimerism shortly after transplant. Cumulative incidence of grades II–IV and III–IV acute GVHD was 41% and 20%, respectively. Localized chronic GVHD was seen in 14% of the patients. Median follow-up was 16.7 months (range 1–63). Five-year OS and PFS were 38% and 39%, respectively. One-year TRM was 42%. UCB transplantation is associated with potential cure of hematologic malignancies and our results are similar to other series. Studies are needed to decrease mortality and improve immune reconstitution.

Successful Stem Cell Tranplant Following HLA Identical Sibling Selection Using PGD Combined With HLA Typing In a Case Of CD40 Ligand Deficiency

Pre-Implantation genetic diagnosis and pre implantation Human Leukocyte Antigen (HLA) typing has been successfully used to conceive unaffected HLA identical siblings to provide stem cells for Hematopoietic Stem Cell Transplant (HSCT) (Goussetis, Evgenios, et al. Biology of Blood and Marrow Transplantation 16.3 (2010): 344-349). This technique has been effective in treatment of disease such as Leukemia, Diamond-Blackfan anemia, Thallasemia etc. With improvement of molecular techniques it has been possible to do gene testing from a single cell. We report first case of X-linked Hyper CD40 ligand deficiency where preimplantation genetic diagnosis (PGD) and pre implantation HLA typing were used to achieve successful HSCT from the resulting HLA identical sibling donor. We used dual hematopoietic stem cell sources from cord blood and bone from the nine month old sibling. Currently, patient is 34 months post transplantation doing well with full immunologic reconstitution. Patient was a 14 month old male who presented with a 2 month history of intermittent fever &amp; poor oral intake. He was drooling excessively and tired on appearance. He had also developed mouth sores and later worsening stridor. Chest x-ray revealed right-sided patchy infiltrates. A laryngoscopy was performed that showed supraglottic stenosis. He was transferred to the PICU where he was intubated, ventilated and treated with 10 days of Meropenem. He was later extubated when it was confirmed that there was healing of the supraglottic area. Patient had Leukopenia and received GCSF. Labs also showed decreased level of IgG, IgA and elevated IgM with low titers to tetanus, H flu and diphtheria vaccines. HIV test was negative. His CD40 ligand test revealed deletion of Exon 2 (Figure 1) confirming the diagnosis. Patient's mother was also confirmed to be a carrier for the mutation. He was started on IVIG supplementation and later subcutaneous Ig. When the patient was diagnosed, he had no siblings but his mother was in the first trimester of a pregnancy. HLA typing was done utilizing chorionic villus sampling. Unfortunately, the fetus was found not to be a match and efforts were directed at finding an unrelated donor through multiple donor drives. A well matched unrelated donor was not available after multiple attempts and donor drives for two years. Cord blood transplant and mismatched unrelated donor transplant were considered. Transplants utilizing mismatched unrelated donors for CD40 ligand deficiency are rare and are associated with an increased risk of complications as well as lower overall survival rate. Due to these factors, the parents declined these options for transplant. Parents then decided to pursue PGD with Preimplantation HLA Testing. After PGD and combined HLA testing an unaffected embryo that was a HLA match to the patient was selected after multiple attempts (Figure 2). The mother underwent IVF with the selected embryo and delivered a male sibling. The genetic testing for CD40 ligand deficiency and HLA typing was repeated on the proposed donor baby after birth. The repeat tests confirmed that the proposed donor baby was unaffected and a 10 of 10 HLA match for the patient. The patient was subjected to a conditioning regimen consisting of Busulfan, Cytoxan and GVH prophylaxis with Methotrexate and Tacrolimus. Following the regimen, the patient was infused with stem cell dose consisting of fresh bone marrow (1.86x108 TNC/Kg) harvested from the matched sibling donor and cryopreserved cord blood (0.29x108 TNC/Kg) obtained during the sibling donor's birth. The patient was found to be well engrafted (98% on day 18, 100% at 9 months) post transplant. The CD40L test showed 60% CD40L on T cells after stimulation with ICOS (inducible co-stimulator) increased from 0% pre-transplant. Most patients with hyper CD40 ligand deficiency inherit the causative gene mutation from one of their parents. The average family size in the US is 3.14 (Daphne Lofquist, et al. Households and Families: 2010 Census Briefs).This results in a low probability of finding a sibling donor that is both unaffected and a HLA match. Undergoing multiple pregnancies with low probabilities of conceiving an unaffected HLA matched sibling is stressful, time consuming and ineffective. PGD combined with HLA typing provides an effective way of choosing an embryo that result in a matched HLA donor sibling and subsequent successful stem cell transplant. Disclosures: No relevant conflicts of interest to declare.

The Comparison Of Clinical Outcomes Between Matched Sibling Stem Cell Transplantation and Immunosuppressive Treatment As a First-Line Treatment For Adult Patients With Severe Aplastic Anemia

▪ BackgroundAllogeneic stem cell transplantation (SCT) from matched-sibling donor (MSD) and immunosuppressive treatment (IST) are the most widely used first-line treatments for patients with severe aplastic anemia (SAA). Overall long-term survival rates are comparable between the two groups. However, patients with age of over 40 have not been generally considered as candidates of SCT from MSD (MSD-SCT) due to higher transplant-related mortality. Recent improvements in MSD-SCT such as less intensive fludarabine-based conditioning, and use of rabbit anti-thymocyte globuline (ATG) instead of horse ATG as first-line IST may change these results. Therefore, we compared the clinical outcomes between MSD-SCT with fludarabine-based conditioning and IST with rabbit ATG and cyclosporine A (CsA). MethodsWe analyzed the clinical results of 54 adult SAA patients who were treated with MSD-SCT and 93 with IST as a first-line treatment from March 2006 to May 2012 at Seoul St. Mary's Hospital, Seoul, Korea. The patients who were treated with MSD-SCT received conditioning with fludarabine (30 mg/m2/day × 6 days), cyclophosphamide (50 mg/kg/day × 2 days), and rabbit ATG (Thymoglobulin®, 2.5 mg/kg/day × 4 days). Those who were treated with IST received rabbit ATG (2.5 mg/kg/day × 5 days) with CsA. ResultsThe median ages were not significantly different between the MSD-SCT group and IST group (38.5 years vs. 43.0 years; P=0.103). Other baseline characteristics were comparable except the interval from diagnosis to treatment (100 days vs. 40 days; P=0.013), absolute lymphocyte count (0.68 × 109/L vs. 0.93 × 109/L; P=0.013), and platelet count (10.0 × 109/L vs. 11.0 × 109/L; P=0.035). In the IST group, overall response and complete response rates at 1 year were 44.1% (95% CI, 33.8-54.8) and 10.8% (95% CI, 5.3-18.9). Treatment failure developed in 55 (59.1%) patients due to non-response in 34 (36.6%), relapse in 5 (5.4%), clonal evolution in 3 (3.2%), and treatment-related mortality in 13 (14.0%) patients. After treatment failure, 17 (18.3%) patients received SCT from MSD or unrelated donor. In the MSD-SCT group, 10 (18.5%) patients experienced treatment failure due to secondary graft failure in 5 (9.3%), clonal evolution in 1 (1.9%), and treatment-related mortality in 4 (7.4%) patients. Among the patients who experienced secondary graft failure, 4 (7.4%) patients received secondary SCT, which resulted in sustained graft function. Consequently, overall survival (OS) at 3 years in the MSD-SCT group was not significantly different compared to that in the IST group (90.7% vs. 81.0%; P=0.139). However, the MSD-SCT group showed significantly higher failure-free survival (FFS) at 3 years compared to the IST group (80.2% vs. 46.6%; P<0.001). When we analyzed the patients with age of over 40 years, OS at 3 years in the MSD-SCT group was not significantly different compared to that in the IST group (87.5% vs. 74.7%; P=0.251), whereas FFS at 3 years in the MSD-SCT group was significantly higher compared to that in the IST group (84.0% vs. 43.0%; P=0.001). ConclusionsOur data suggest that MSD-SCT is more favorable than IST as a first-line treatment, considering the curative nature of MSD-SCT even in patients over 40 years of age. Disclosures:No relevant conflicts of interest to declare.

Birth Order and Transplantation Outcome in HLA-Identical Sibling Stem Cell Transplantation: An Analysis on Behalf of the Center for International Blood and Marrow Transplantation

Allogeneic stem cell transplantation (SCT) is the most effective treatment option for many hematologic malignancies, but graft-versus-host disease (GVHD) remains a major cause of treatment failure. Along with well-established risk factors for transplantation outcomes, recent single-center studies have identified a birth order effect in HLA-identical sibling SCT, with lower rates of acute and chronic GVHD and improved overall survival when the donor is younger than the recipient. One hypothesized mechanism for this effect is microchimerism due to fetomaternal and transmaternal sibling cell trafficking during pregnancy as the donor is exposed to recipient antigens in utero. The aim of the present study was to validate previously reported single-center data in a large, multicenter cohort provided by the Center for International Blood and Marrow Transplantation. All adult and pediatric patients (n = 11,365) with a hematologic malignancy who underwent allogeneic SCT with a graft from an HLA-identical sibling donor between 1990 and 2007 were included. When donors were younger than recipients, there was a significantly lower rate of acute GVHD grade II-IV and chronic GVHD in children, as well as a lower rate of chronic GVHD in adolescents. However, the hypothesized overall positive effect of lower relapse and better survival when donors are younger than recipients was not observed. Our data suggest that if otherwise equally matched, a graft from a younger sibling may be superior to a graft from an older sibling for children and adolescents undergoing SCT.

Comparison of Outcomes Between Unrelated Donor and Sibling Donor Transplant in Adult Patients with Severe Aplastic Anemia: A Single Center Retrospective Study

AbstractAbstract 4151For the patient with severe aplastic anemia (SAA) who do not have a sibling donor and had failed to immunosuppressive therapy, allogeneic stem cell transplantation (SCT) from an unrelated donor is therapeutic option. Although unrelated SCT (u-SCT) has shown less favorable outcomes, there were few studies comparing unrelated donor to sibling donor transplant. The aim of the present study was to evaluate the feasibility of u-SCT in adult SAA patients compared with sibling SCT (s-SCT).This study examined 157 consecutive patients who underwent allogeneic SCT at our institution between April 2001 and December 2011. In this study, for a homogenous cohort, patients who received second allogeneic SCT were excluded. The diagnosis of acute GVHD or chronic GVHD was made as the NIH published consensus criteria.There were 71 male and 86 female patients with a median age of 31 years (range, 13–59 years). The median interval from the diagnosis to transplantation was 24 months (range; 1–346 months). The median transfusions prior to SCT were 47 units (range; 4–680 units). Patients with SAA (n=114) or VSAA (n=43) had received SCT from sibling donor (n=82) or unrelated donor (n=75). The conditioning regimen for s-SCT consisted of fludarabine (180 mg/m2) + cyclophosphamide (CY, 100 mg/kg) + ATG (10 mg/kg), and the conditioning regimen for u-SCT consisted of TBI (fractionated, 800 cGy) + CY (100–120 mg/kg) ± ATG (2.5 mg/kg) (Lee JW, et al, Biol BMT 2011, 17:101). GVHD prophylaxis consisted of CsA + MTX in s-SCT and FK506 + mini-MTX in u-SCT, respectively. After a median follow-up of 45 and 50 months for s-SCT and u-SCT, respectively, the 3-year estimated OS rates were 92.7 and 89 % for s-SCT and u-SCT (P=0.650), respectively. 15 patients (7 in s-SCT, 8 in u-SCT) died of transplant toxicities, including in s-SCT, acute GVHD (n=1), infections (n=4), and occurrence of MDS or AML (n=2) and in u-SCT, acute GVHD (n=4), chronic GVHD (n=1), infection (n=1), thrombotic microangiopathy (n=1). Nine patients (8 in s-SCT, 1 in u-SCT) developed secondary engraftment failure. The cumulative incidence of acute GVHD (≥grade II) was 4.9% in s-SCT and 44.0% in u-SCT (P<0.001). The cumulative incidence of chronic GVHD also showed higher in u-SCT than s-SCT (4.2% vs 44.4%, P<0.001).This study showed that survival of u-SCT is comparable to that of s-SCT. Therefore, early application of u-SCT should be considered in patients with SAA/VSAA in case of availability of well matched donor. Because the incidence of GVHD is significantly higher in u-SCT than s-SCT, further study is mandatory to reduce the incidence of GVHD in u-SCT setting. Disclosures:No relevant conflicts of interest to declare.

Overview of T-cell depletion in haploidentical stem cell transplantation.

Overview of T-cell depletion in haploidentical stem cell transplantation.