HLA-matched Sibling Hematopoietic Stem Cell Transplantation Research Articles

Clonal Dynamics after Haematopoietic Stem Cell Transplantation Using Genome-Wide Somatic Mutations

Background Allogeneic hematopoietic stem cell transplantation (HSCT) is a potentially curative treatment received by over 40,000 patients/year. Despite being an established treatment for 50 years, fundamental questions remain about the dynamics of long-term stem cell engraftment and immunological reconstitution in humans. How many of the >108 transplanted CD34+ cells contribute to long-term hematopoiesis? How does increasing age of donor and recipient reshape clonal dynamics of HSCT? Are the same HSCs responsible for both myeloid and lymphoid reconstitution? Does HSCT increase mutation rates or clonal evolution? Methods We used spontaneous somatic mutations occurring genome-wide in single hematopoietic stem and progenitor cells (HSPCs) to reconstruct high-resolution lineage trees of the hematopoietic system after HSCT. We analysed blood samples from both the donor (D) and recipient (R) for 10 HLA-matched sibling HSCTs - the duration of time elapsed since transplant ranged from 9 to 31 years, meaning that we could study genuinely long-term reconstitution in the recipient, compared to the unperturbed hematopoiesis in the donor. For each individual (D and R), peripheral blood CD34+ HSPC-derived colonies were grown on methylcellulose medium. Whole genome sequencing was performed on 100-300 colonies per individual, generating a final dataset of 3,413 genomes. Somatic mutations from each colony's founder cell were identified and used to build phylogenies for each D/R pair (Figure). For 7/10 pairs, we then used ultra-deep targeted sequencing to assess clonal contributions in sorted mature D/R cell subsets, including monocytes, B-cells and T-cells. Approximate Bayesian computation was used to estimate numbers of engrafting HSCs. RESULTSIn most cases, 10,000 - 50,000 HSCs engrafted and contributed to long-term reconstitution during HSCT. However, grafts from older donors, or those with lower CD34+ cell doses tended to have lower estimates, some as low as <1,000 cells.Recipients had decreased clonal diversity compared to their donors, equivalent to 8-10 years of additional ageing.We detected 93 individual clones that contributed >1% of the population of at least one mature cell type. Each of these expanded clones was detectable in both the donor and the recipient, confirming that they had originated prior to HSCT.Only ~10% of these expanded clones carried mutations in genes known to contribute to clonal hematopoiesis, suggesting that there are many other genes that can influence stem cell dynamics after HSCT.The sizes of these expanded clones often varied dramatically between donors and recipients, up to 25-fold differences in some instances.The population bottleneck caused by HSCT was not sufficient to explain the differences in clonal composition between recipient and donor, suggesting that there are additional HSCT-specific selective pressures during stem cell harvest, in vitro manipulation, infusion, homing and engraftment.While large proportions of mature myeloid and B-cell populations usually derived from the identified clones, this was not true for T-cell populations. This suggests that following HSCT, T-cells are largely derived from stem cells that do not contribute substantially to myeloid reconstitution.HSPC mutation burdens in recipients were similar to their donors, contradicting previous hypotheses that the proliferative stress of HSCT would lead to excess mutations. CONCLUSION Our findings reveal that tens of thousands of HSCs typically engraft during HSCT, but with significant loss of clonal diversity in recipients compared to their unperturbed donors. HSCT-specific selective pressures shape dynamics of hematopoietic reconstitution in the recipient, with substantial differences in clonal composition between myeloid and T-lymphoid populations. This study shows the power of using somatic mutations as clonal marks for lineage tracing in real-world HSCT in humans, an approach we believe will be a new gold standard for understanding the impact of different stem cell sources and transplant protocols. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Nonmyeloablative Transplant for Sickle Cell Disease Does Not Lead to Kidney Dysfunction Post-HSCT

Hematopoietic stem cell transplantation (HSCT) is a potentially curative therapy for patients with sickle cell disease (SCD). Though HSCT can reverse the SCD phenotype, both acute kidney injury (AKI) and chronic kidney disease (CKD) have been associated with HSCT. SCD alters renal function thus the impact of HSCT on renal function in SCD patients is a critical area of exploration. Here, we report the effect of HSCT on renal function in people with SCD.This study analyzes data from 195 patients who received HLA-matched sibling or haploidentical HSCT for SCD at Imperial College London (ICL), National Institutes of Health (NIH), and University of Illinois Chicago (UIC). The former is a pediatric cohort and all sites employed a nonmyeloablative conditioning. Patients' renal function was assessed at baseline and annually thereafter for up to 3-years.We examined the prevalence of CKD before and after HSCT, estimated glomerular filtration rate (eGFR) and urine albumin to creatinine ratio (UACR) trends after HSCT, and the incidence of AKI within 100 days of HSCT. We defined and staged AKI according to Kidney Disease: Improving Global Outcomes (KDIGO) criteria; we calculated eGFR with the CKD- epidemiology collaboration (CKD-EPI) equation for adults and Bedside Schwartz for children.The median eGFR declined annually but remained within the normal range throughout the follow-up period: the baseline median eGFR was 138 ml/min/1.73m 2 and declined by 7 in the first year of follow-up and by additional decreases of 5 and 3.6 ml/min/1.73m 2 in subsequent years (p-values of 0.07, 0.0002, and 0.0002 for years 1, 2, and 3 for comparison to baseline from regression model of eGFR). No differences in eGFR were seen for covariates in the model: haploidentical vs. matched sibling, engraftment status, gender, or site. The downward eGFR trend may represent an improvement in renal function toward normal as hyperfiltration (eGFR ≥150 ml/min/1.73m 2) was present in 28% of patients at baseline and steadily declined to 7% by 3 years post-HSCT. There was a corresponding increase in patients with normal eGFR (60-149 ml/min/1.73m 2) from 59% at baseline to 88% at 3 years post-HSCT.Among the ICL and NIH cohorts (UIC excluded due to use of a different AKI determination strategy), 58% of patients developed AKI in the early post-HSCT period. 67% of AKI cases were mild, stage 1; 25% were moderate, stage 2; and 8% were severe, stage 3 AKI.This study demonstrates that HSCT in patients with SCD is associated with a transient increase in UACR but not associated with a significant increase in CKD prevalence by 3-years post-HSCT. The stability of UACR compared to baseline by the 3-year time point suggests that even more mild renal damage may stabilize after HSCT. While there is a substantial decline in eGFR from baseline to each annual follow-up, the proportion of patients whose eGFR was in the normal range increased as the prevalence of hyperfiltration decreased. Finally, while AKI occurred in more than half the patients in our cohort, the preponderance developed only mild AKI. Therefore, our data indicate that nonmyeloablative HSCT for SCD does not lead to significant kidney dysfunction post-HSCT. DisclosuresSaraf: Pfizer: Research Funding; Global Blood Therapeutics: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding. Diamantidis: United Health Group: Consultancy.

Renal Function Is Preserved after Hematopoietic Stem Cell Transplantation for Sickle Cell Disease

Chronic kidney disease (CKD) is a common complication and a significant contributor to morbidity and mortality for patients with sickle cell disease (SCD). Hematopoietic stem cell transplantation (HSCT) is a curative strategy for SCD and potentially life-saving therapy. Though HSCT can reverse the SCD phenotype, acute and chronic kidney dysfunction may be an unintended consequence. The incidence of acute kidney injury (AKI) and the effect of different HSCT regimens on progression or amelioration of CKD in adults with SCD have not previously been reported. This cohort study analyzes prospectively collected data from 106 patients transplanted for SCD at the National Institutes of Health (NIH) Clinical Center, including HLA-matched sibling (N=71) and haploidentical HSCT (N=26) where nonmyeloablative conditioning was given, as well as gene therapy (N=9), where a myeloablative regimen was employed. Patients' renal function was assessed at baseline and annually thereafter for up to 3-years. We sought to define 1) the prevalence of CKD before and after HSCT 2) the incidence of CKD after HSCT 3) estimated glomerular filtration rate (eGFR) trends after HSCT 4) urine albumin to creatinine ratio (UACR) change after HSCT and 5) the incidence of AKI within 100 days of HSCT. We defined and staged AKI according to Kidney Disease: Improving Global Outcomes (KDIGO) criteria (stage 1: 1.5-1.9 times baseline serum creatinine increase or ≥0.3 mg/dl increase; stage 2: 2-2.9 times baseline; stage 3: 3 times baseline, increase in creatinine to ≥4.0 mg/dl, or initiation of renal replacement therapy). The baseline prevalence of CKD was 6% when defined conservatively (eGFR &amp;lt;60 ml/min/1.73m2); this prevalence did not change significantly post-HSCT. When using a more comprehensive definition of CKD (urine albumin to creatinine ratio (UACR) ≥30 mg/g OR eGFR &amp;lt;60 ml/min/1.73m2), 48% had CKD at baseline, 62% at 1 year, 55% at 2 years, and 55% at 3 years (with p=0.02, 0.26, 0.54 for longitudinal model comparing post-HCST prevalence to baseline). The incidence of CKD any time after HSCT was 1% and 11% according to each of these CKD definitions, respectively. 25% of participants had follow-up of ≤2 years, median follow-up was 3 years. The median eGFR remained within the normal range throughout the follow-up period but declined annually: the baseline median eGFR was 139.7 ml/min/1.73m2 and declined by 7.6, 11.6, and 20.9 ml/min/1.73m2 each year post-HSCT (p&amp;lt;0.0001). In an adjusted regression analysis, there was no association of gender, engraftment status, transplant regimen, or conditioning agent- specifically use of pentostatin, total body irradiation, or post-transplant cyclophosphamide- with worsening eGFR (p&amp;gt;0.05 for each variable). This eGFR trend represents an improvement in renal function toward the normal range: there was a statistically significant decrease in hyperfiltration (eGFR ≥150 ml/min/1.73m2), which was present in 34% patients at baseline but steadily declined to 12% by 3 years post-HSCT (p = 0.0006 for adjusted longitudinal model). There was a corresponding increase in patients with normal eGFR (60-149 ml/min/1.73m2) from 60% at baseline to 80% at 3 years post-HSCT. UACR showed an increase in the year following transplant that subsequently dissipated. Median UACR at baseline was 24.3 mg/g with annual follow-up median values of 62.2, 38.9 and 36.2 mg/g (p-values of 0.007, 0.41, and 0.69 for comparison to baseline from regression model of log transformed UACR). In our cohort, 58% of patients experienced AKI in the first 100 days post-HSCT; which is consistent with previous data. 77% of those were mild, stage 1; 13% were moderate, stage 2; and 10% were severe, stage 3 AKI. This study demonstrates that HSCT in patients with SCD is not associated with a significant increase in CKD incidence or prevalence at 2 and 3 years post-HSCT. While there is a substantial decline in eGFR from baseline to each annual follow-up, the proportion of patients whose eGFR was in the normal range increased and the prevalence of hyperfiltration decreased. The stability of UACR after an initial increase at the 1-year time point further suggests that even more mild renal damage may stabilize after transplant. Finally, AKI occurred in over half of the patients in our cohort, though the preponderance developed mild AKI. Disclosures Diamantidis: United Health Group: Membership on an entity's Board of Directors or advisory committees.

Analysis of Factors Influencing Overall Survival of MDS Patients Transplanted with HSCs

To analyze the effect of clinical features, routine laboratory examination and related gene mutation on the OS of patients with myelodysplastic syndrome (MDS) after hematopoietic stem cell transplantation (HSCT). 121 patients diagnosed as MDS and underwent hematopoietic stem cell transplantation in the First Affiliated Hospital of Soochow University from October 2013 to August 2018 were selected. Basic information of the patients was collected, and blood cells, bone marrow blasts at initial diagnosis, chromosomal karyotypes and gene mutations of the patients were detected.The effect of different factors on overall survival (OS) was analyzed by statistical method. Kaplan-Meier univariate analysis shows that OS was significanly different among different age groups. The 3-year OS rate of patients aged 0-29 years was (83.3±7.7) %, the 3-year OS rate in patients aged 30-49 years was (58.1±7.7 %), and the 3-year OS rate of patients aged 50-69 years was (31.0±22.6) %, which was statistically different (P＜0.05) between different groups. There were also significant differences in OS among patients with different transplantation types. 3-year OS rate: HLA-matched sibling HSCT＞unrelated HLA-matched HSCT＞haploidentical HSCT＞micro HSCT. The OS rate of patients with bone marrow blasts≥10% seems lower than blasts＜10%, but there was no statistical difference.The 3-year OS rate of patients with chromosomal karyotype complex abnormality was (47.7±11.5) %, and that of patients without complex abnormality was (80±4.2) % which was statistical difference (P＜0.05). Patients with DNMT3A, NRAS, TP53 and GATA2 mutations had shorter OS time compared with patients without mutation of these genes, which shows statistically significant (P＜0.05). COX multivariate analysis showed that age, chromosome karyotype, DNMT3A, TET2, GATA2 and NRAS were the independent factors influencing OS of patients after HSCT, with statistically significant difference. age of patients, donor selection of HSCT, chromosome karyotype, DNMT3A, NRAS, TP53, GATA2 and TET2 gene mutations are all independent factors affecting the OS of patients after HSCT. Therefore, the assessment of the OS of MDS patients with transplantation requires comprehensive consideration.

The impact of pre-existing HLA and red blood cell antibodies on transfusion support and engraftment in sickle cell disease after nonmyeloablative hematopoietic stem cell transplantation from HLA-matched sibling donors: A prospective, single-center, observational study.

BackgroundHematopoietic stem cell transplantation (HSCT) is curative for patients with sickle cell disease (SCD). Prior to HSCT, patients with SCD commonly receive RBC transfusions with some becoming RBC or HLA alloimmunized. This alloimmunization may impact post-HSCT transfusion requirements and donor engraftment.MethodsThe study population included patients with SCD transplanted on a single-center nonmyeloablative, HLA-matched sibling HSCT trial at the National Heart, Lung, and Blood Institute (NHLBI) who had a pre-HSCT sample available for HLA class I antibody testing. We evaluated transfusion requirements and engraftment outcomes comparing patients with and without pre-existing HLA and RBC antibodies.FindingsOf 36 patients studied, 10 (28%) had HLA class I antibodies and 11 (31%) had a history of RBC alloantibodies. Up to day +45 post-HSCT, patients with HLA antibodies received more platelet transfusions (median 2.5 vs 1, p = 0.042) and those with RBC alloantibodies received more RBC units (median 7 vs 4, p = 0.0059) compared to respective non-alloimmunized patients. HLA alloimmunization was not associated with neutrophil engraftment, donor chimerism, or graft rejection. However, RBC alloimmunization correlated with a decreased donor T cell chimerism at 1 year (median 24% vs 55%, p = 0.035).InterpretationPre-existing HLA and RBC alloantibodies are clinically significant for patients undergoing HLA-matched nonmyeloablative HSCT. Testing for both HLA and RBC antibodies is important to help estimate transfusion needs peri‑HSCT. The association of lower donor T cell chimerism and pre-existing RBC alloantibodies needs further investigation.FundingNIH Clinical Center and NHLBI Intramural Research Program (Z99 CL999999, HL006007–11) and the Thrasher Research Fund.

Impact of donor KIRs and recipient KIR/HLA class I combinations on GVHD in patients with acute leukemia after HLA-matched sibling HSCT

In addition to T cells, NK cells can also participate in the outcome of hematopoietic stem cell transplantation (HSCT) mainly through the interaction between donor killer cell immunoglobulin-like receptors (KIRs) and recipient human leukocyte antigen (HLA) class I molecules. There is a risk of GVHD other than leukemia relapse after allogeneic HSCT that activation of donor NK cells in the absence of appropriate inhibitory ligands will be one of the reasons. To investigate the impact of donor KIRs and recipient KIR/HLA class I combinations on GVHD and leukemia relapse in patients with acute leukemia after HSCT, 100 patients with acute leukemia who received HSCT from their HLA-matched siblings were included in this study. Genotypes of 16 KIR genes and two 2DS4 variants (full length and deleted alleles), along with HLA-A/B genotypes, were determined by PCR-SSP. HLA-C genotyping was done with the SSO-Luminex method. Chimerism analysis was done using 16 short tandem repeats (STRs) to detect early leukemia relapse. Acute (a)GVHD occurred in 38 patients, and 16 of them died during the study. None of the recipients showed any sign of leukemia relapse after HSCT. Full donor chimerism was observed in all tested patients during the first year after HSCT. Our results also indicated an increased risk of aGVHD in AA recipients with the C2/Cx, Bw4+ (or A-Bw4+) or HLA-A3−/A11− genotypes who received HSCT from Bx donors. Our results showed that donor selection based on donor-recipient KIR genotypes and recipient HLA class I status can improve the outcome of HSCT.

Genetic polymorphism related to monocyte-macrophage function is associated with graft-versus-host disease

Despite detailed human leukocyte antigen (HLA) matching and modern immunosuppressive therapy, severe graft-versus-host disease (GvHD) remains a major hurdle for successful allogeneic hematopoietic stem cell transplantation (HSCT). As the genetic diversity in GvHD complicates the systematic discovery of associated variants across populations, we studied 122 GvHD-associated single nucleotide polymorphisms (SNPs) in 492 HLA-matched sibling HSCT donor-recipient pairs from Finland and Spain. The association between these candidate SNPs and grade III–IV acute GvHD and extensive chronic GvHD was assessed. The functional effects of the variants were determined using expression and cytokine quantitative trait loci (QTL) database analyses. Clear heterogeneity was observed in the associated markers between the two populations. Interestingly, the majority of markers, such as those annotated to IL1, IL23R, TLR9, TNF, and NOD2 genes, are related to the immunological response by monocytes-macrophages to microbes, a step that precedes GvHD as a result of intestinal lesions. Furthermore, cytokine QTL analysis showed that the GvHD-associated markers regulate IL1β, IFNγ, and IL6 responses. These results support a crucial role for the anti-microbial response in GvHD risk. Furthermore, despite apparent heterogeneity in the genetic markers associated with GvHD, it was possible to identify a biological pathway shared by most markers in both populations.

At Least 20% Donor Myeloid Chimerism Is Necessary to Reverse the Sickle Phenotype after Allogeneic Hematopoietic Stem Cell Transplantation

Hematopoietic stem cell transplantation (HSCT) is the only curative option for patients with sickle cell disease (SCD). HLA-matched sibling HSCT has become increasingly successful, and haploidentical approaches are being explored to increase the donor pool. Advances have also been made in gene therapy such that clinical trials are now underway for the first time in patients with SCD. The question of what percentage of donor chimerism is necessary to cure SCD is critical in identifying the target percentage of the transferred globin gene needed to reverse SCD. One study showed that a whole blood donor chimerism level as low as 11% was sufficient to render patients free of SCD1; however, lineage specific chimerism was not performed. We have previously demonstrated that peripheral blood myeloid chimerism reflects the HSC compartment. We sought to determine the percentage of donor myeloid chimerism (DMC) necessary to reverse the sickle phenotype in our patients who underwent HSCT.In protocols testing both HLA-matched and haploidentical nonmyeloablative HSCT in over 70 patients with SCD, we identified 3 who despite robust initial engraftment experienced declining DMC levels over prolonged follow up eventually associated with return of SCD. All 3 patients’ donors had sickle cell trait with HbS ranging from 36-40%. Percentages of DMC and HbS were monitored at day 100 and at least every 6-12 months post-transplant thereafter. The 3 patients initially achieved nearly 100% DMC; however, their chimerism levels slowly decreased over time (see Figure 1). As long as mean DMC was >20%, mean %HbS remained less than 50% (similar to their sickle cell trait donors) and the patients stayed free of SCD-related symptoms. However, mean DMC at 2, 3, 4, and 5 years post-transplant fell below 20%, were 18, 12, 11, and 10%, and were associated with mean %HbS levels of 52.5, 51.6, 57.8, and 64.2%. Further, 2 of the patients had return of their recurrent painful crises, and the third patient with moyamoya syndrome developed severe anemia requiring re-initiation of chronic transfusion therapy.We subsequently developed a mathematical model to test the hypothesis that the percentage of necessary DMC is determined solely by the differences between donor and recipient red blood cell (RBC) half-lives. The model incorporates donor and recipient stem cell kinetics (self renewal and differentiation), which are assumed to be identical, as well as donor and recipient RBC survival times, which are known to be different. The model assumes first order kinetics for erythropoiesis and RBC senescence/destruction. It predicts the fraction of donor RBC as a function of DMC and donor/recipient RBC lifetimes. Although we did not directly measure RBC lifetimes, we identified 4 patients with SCD in the literature where reticulocyte percentages ranged from 9.0 to 21.0% and corresponding RBC half-lives determined by radioactive sodium chromate were 1 to 11 days2. We assumed that in erythropoietic homeostasis, RBC lifetime must be proportional to the inverse of the reticulocyte fraction; and we confirmed this relationship using data from these 4 patients with a good coefficient of determination (R2= 0.91). Therefore we reasoned that (at stable hemoglobin concentration) the reticulocyte fraction is a good proxy for RBC lifetime. Figure 1 depicts a comparison between the measured and our model predicted HbS fractions in our 3 patients transplanted with sickle cell trait donors using a mean sickle RBC lifetime of 7 days. As observed in our 3 patients, our model predicts that DMC of at least 20% is necessary to reverse the sickle phenotype.Therefore, our model predicts that genetic strategies aimed at SCD must achieve correction levels of at least 20% to reverse the sickle phenotype. Further, the mechanism explaining why a minority of donor myeloid cells is adequate is due soley to the vast differences in RBC survival between donor and recipient cells. Lastly, recipient RBC survival is lowest in patients with the highest reticulocyte counts suggesting that gene therapy may be most efficacious in subjects with high reticulocyte percentages as our model predicts that lower levels of correction would be sufficient in this setting. Further studies are indicated to validate our findings.1. Walters MC et al. Biol Blood Marrow Transplant. 2001;7(12):665-673.2. Weinstein IM et al. Blood. 1954;9(12):1155-1164. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Comparing Outcomes with Bone Marrow or Peripheral Blood Stem Cells as Graft Source for Matched Sibling Transplants in Severe Aplastic Anemia across Different Economic Regions

Bone marrow (BM) is the preferred graft source for hematopoietic stem cell transplantation (HSCT) in severe aplastic anemia (SAA) compared with mobilized peripheral blood stem cells (PBSCs). We hypothesized that this recommendation may not apply to those regions where patients present later in their disease course, with heavier transfusion load and with higher graft failure rates. Patients with SAA who received HSCT from an HLA-matched sibling donor from 1995 to 2009 and reported to the Center for International Blood and Marrow Transplant Research or the Japan Society for Hematopoietic Cell Transplantation were analyzed. The study population was categorized by gross national income per capita and region/countries into 4 groups. Groups analyzed were high-income countries (HIC), which were further divided into United States–Canada (n = 486) and other HIC (n = 1264); upper middle income (UMIC) (n = 482); and combined lower-middle, low-income countries (LM-LIC) (n = 142). In multivariate analysis, overall survival (OS) was highest with BM as graft source in HIC compared with PBSCs in all countries or BM in UMIC or LM-LIC (P < .001). There was no significant difference in OS between BM and PBSCs in UMIC (P = .32) or LM-LIC (P = .23). In LM-LIC the 28-day neutrophil engraftment was higher with PBSCs compared with BM (97% versus 77%, P = .002). Chronic graft-versus-host disease was significantly higher with PBSCs in all groups. Whereas BM should definitely be the preferred graft source for HLA-matched sibling HSCT in SAA, PBSCs may be an acceptable alternative in countries with limited resources when treating patients at high risk of graft failure and infective complications.

Hematopoietic Stem Cell Transplantation from HLA Identical Sibling Forsickle Cell Disease an International Survey on Behalf of Eurocord-Monacord, EBMT Paediatric Disease Working Party and CIBMTR

Introduction:Hematopoietic stem cell transplantation (HSCT) is, to date, the only curative therapy for sickle cell disease (SCD). However, HSCT is offered to relatively few patients with SCD for a number of reasons including lack of a suitable HLA-matched donor, lack of consensus on indications for HSCT, the potential for trading one chronic condition (i.e., SCD) for another, such as chronic graft-versus-host disease (GVHD), and the mortality associated with the procedure. To-date, most HSCTs for SCD have utilized matched siblings as donors and are performed in children and adolescents. We report outcomes after HLA-matched sibling HSCT of patients reported to the Eurocord-Monacord/European Group for Blood and Marrow Transplantation (EBMT) and Center for International Blood and Marrow Transplant Research (CIBMTR).Material and methods:One thousandpatients with SCD received HLA identical sibling HSCT between 1991 and 2013; n=439 from CIBMTR and n=561 from EBMT centers. HSCTs were performed in 90 centers in 23 countries.Results:Median age at HSCT was 9 years (range 1-54y); 85% of patients were aged <16 years. Approximately half of patients were female and 53% of HSCTs were performed after 2007. Most patients (94%) were homozygotes for hemoglobin S (HBS). The most common indication for HSCT was stroke. Other indications included: central nervous system event lasting longer than 24 hours, elevated cerebral arterial velocity, acute chest syndrome or vaso-occlusive crisis requiring hospitalization. Red blood cell transfusions were given before HSCT to 93% and hydroxyurea to 56% of the evaluable patients (N=510). Most HSCTs (n=872; 87%) used myeloablative-conditioning regimens, mainly based on the combination of busulfan with cyclophosphamide (n=719; 82%) or fludarabine (n=82; 9%). One hundred and twenty six patients (13%) received reduced intensity conditioning regimens; fludarabine with cyclophosphamide was the predominant regimen (n=48; 38%). Most regimens included in vivo T-cell depletion (71%) with anti-thymocyte globulin (n=630) or alemtuzumab (n=76). The predominant GVHD prophylaxis regimens were cyclosporine alone (19%), or combined with methotrexate (56%). The predominant stem cell source was bone marrow (84%); peripheral blood and cord blood were employed in 7% and 9% of patients, respectively. The median follow-up was 45 (1.1-324.6) months.The cumulative incidence (CI) of neutrophil engraftment at day+60 was 98% (96.6% for CB, 98.3% for BM and 95.2% for PB) with a median time to recovery of 19 days, while that for platelet engraftment was 98 % (96±2% for CB, 99±1% for BM and 98±9% for PBSC) with a median time to recovery of 25 days. Twenty-six patients experienced primary and 47 patients secondary graft failure; 67 patients died mainly due to GVH or infection. The 3-year probabilities of overall (OS) and event-free survival (EFS, alive with engraftment) were 94% (95% CI 92-95) and 90% (95% CI 68-82), respectively. According to stem cell source, 3-year OS was 99% after CB, 94% after BM and 80% after PBS (p<0.0001). In multivariate analysis, every year in age increment (HR 1.1, 95% CI 1.07-1.14, p<0.001) and use of peripheral blood (HR 3.43, 95% CI 1.49-7.88, p=0.004) were associated with higher mortality. In univariate analysis, EFS was better in patients receiving myeloablative compared to reduced intensity conditioning (91±1% vs 82 ±1%, respectively; p<0.001). In multivariate analysis, EFS was lower with every year in age increment (HR 1.05, 95% CI 1.02-1.07, p<0.001), peripheral blood grafts (HR 1.83, 95% CI 1.07-3.15, p=0.03) and HSCTs prior to 2000 (HR 0.77, 95% CI 0.64-0.92, p=0.005). CI of acute GVHD grade 2-4 was 14.4% (12.2-16.7) of chronic GVH 13.3 (11-15.8). Risks of acute GVHD were higher with increasing age (HR1.04 95% CI 1.01-1.07, p=0.008). None of the variables tested were associated with chronic GVHD.Conclusion:This large registry based international study shows that HLA identical sibling transplant is successful more than 90% of the patients with severe SCD with limited transplant related complications (rejection, GVHD). Strategies aimed at lowering graft failure and GVHD are desirable to further optimize the observed 3-year event-free survival. Importantly, these data should increase awareness to early referral to HSCT of patients with severe SCD. DisclosuresWalters:ViaCord and AllCells, Inc: Other: Medical director. Bertrand:ERYTECH Pharma: Consultancy. Peters:Medac: Research Funding; Fresenius: Research Funding; Amgen: Research Funding; Jazz: Research Funding; Novartis: Research Funding; Pfizer: Research Funding; Sanovi: Research Funding; Pierre-Fabre: Research Funding.

Efficacies of hematopoietic stem cell transplantation for severe aplastic anemia: a report of 43 patients

To evaluate the efficacies of hematopoietic stem cell transplantation (HSCT) for severe aplastic anemia (SAA). A total of 43 SAA patients (SAA-I, n = 29; SAA-II, n = 14) underwent HSCT from 2002 January to 2013 December. There were 22 males and 21 females with a median age of 31 (12-49) years. And 35 patients received HLA-matched sibling HSCT (Sib-HSCT) while another 8 had unrelated donor HSCT (UD-HSCT). The hematopoietic stem cells were collected from bone marrow (n = 10), peripheral blood (n = 23) and bone marrow & peripheral blood (n = 10). Conditioning regimens were mostly composed of Fludarabine, antihuman thymocyte gloBulin and cyclophosphamide. Cyclosporine and methotrexate (including mycophenolate mofetil for UD) were offered for preventing graft-versus-host disease (GVHD). The median counts of mononuclear cell and CD34(+) stem cell were 8.1 (2.4-13.5) ×10(8)/kg and 3.7 (2.3-14.7) ×10(6)/kg respectively. Hematopoiesis reconstitution was achieved in 42 patients. The median periods for neutrophils to 0.5×10(9)/L and platelets to 20×10(9)/L were +10 (+8-+25) days and +14 (+8-+80) days respectively. The median of survival time was not reached. Overall survival (OS) and failure-free survival (FFS) at 5 years had no differences between UD-HSCT and Sib-HSCT groups (OS: 87.6% vs 84.5%, P = 0.87; FFS: 86.2% vs 79.5%, P = 0.64). The same results also were seen between age ≤ 20 group and > 20 group (OS: 92.0% vs 82.5%, P = 0.39; FFS: 91.0% vs 77.3%, P = 0.38). The median follow-up time was 17.0 (0.4-140.0) months. And 8/43 patients died from thrombotic microangiopathy (TMA) (n = 1), TMA associated with capillary leak syndrome (n = 1), pulmonary infection (n = 3), acute GVHD of grade IV (n = 1) and unknown causes (n = 2). Sib-HSCT is preferred for SAA patients under 40 years. Searching of HLA-matched unrelated donor should be performed soon after diagnosis if HLA-matched sibling is unavailable.

Improved Outcomes in Diamond Blackfan Anemia Treated Via Stem Cell Transplantation Since the Year 2000.

Abstract 3202Poster Board III-139 BackgroundDiamond Blackfan Anemia (DBA) is a rare disorder characterized by red cell aplasia, congenital anomalies and a predisposition to cancer. Corticosteroids and red cell transfusions are the mainstays of therapy. Hematopoietic stem cell transplantation (HSCT) is curative in DBA, however outcomes have been variable. Nonetheless, preliminary data suggest improvement in HSCT outcomes since the year 2000. PurposeTo compare the degree of HLA compatibility, pre-transplantation iron burden, and survival before and since the year 2000 in sibling and alternative donor HSCT recipients, in order to evaluate if enhanced HLA matching techniques along with improved control of pre-transplant iron burden contribute to improved outcomes. MethodsThe DBA Registry (DBAR) is a comprehensive database of patients with DBA who are enrolled after informed consent is obtained. The patients, their families, and their physicians complete a detailed questionnaire. A review of medical records and telephone interviews are performed to complete and clarify the information provided. Presence of iron burden prior to transplant was determined based upon fulfillment of any of the following criteria: 1) serum ferritin level > 2000 ng/mL within 2 months prior to HSCT, 2) liver iron concentration greater than 8 micrograms per gram, dry weight within 6 months prior to HSCT (measured by MR or biopsy), 3) liver histology from biopsy or autopsy within 6 months of HSCT demonstrating 3+ or greater hemosiderosis or any level of fibrosis. ResultsAs of August 11, 2009, 583 patients have been enrolled in the DBAR, with 55 having undergone an allogeneic sibling or alternative donor HSCT. The median age at HSCT for all patients was 7 years, 8 months; 7 years, 2 months versus 9 years, 8 months for the 32 sibling and 23 alternative donor HSCT, respectively. There were 8 sibling and 13 alternative donor HSCT done prior to the year 2000 compared to 24 and 10 HSCT performed since the year 2000, respectively.Prior to 2000, 8 of 8 sibling HSCT performed were HLA matched, whereas since the year 2000, there were 19 complete and 1 partial HLA matched sibling HSCT done. HLA reports on 4 sibling HSCT were not available for review. There were 5 full and 7 partial matches amongst the alternative donor HSCT done prior to 2000 and 6 full and 2 partial matches since the year 2000. Prior to the year 2000 HLA typing was done by serologic typing alone or by serologic methods for Class I and molecular methods for Class II. HLA typing was done by molecular techniques after the year 2000.Overall survival at 5 years for all patients is 56.6 ± 8.7%; 77.3 ± 8.3% for sibling HSCT and 31.5 ± 12.7% for alternative donor HSCT (p=0.012). For sibling HSCT done prior to 9 years of age survival is 90.0 ± 9.5% and for those older than 9 years of age is 70.0 ± 11.6% (p=0.007). Survival for all patients prior to 2000 and since 2000 is 47.1 ± 11.0% and 72.4 ± 9.3%, respectively (p=0.184). Survival for alternative donor HSCT is 23.1 ± 11.7% prior to 2000 and 85.7 ± 13.2% since 2000 (p=0.047).From data which are available, there are no reported pre-2000 cases of sibling HSCT where absence of iron is documented. Two cases of sibling HSCT pre-2000 had known significant iron overload. One of these patients is the only fatality amongst sibling donor HSCT pre-2000. Post-2000, there were 3 cases of sibling HSCT with significant iron overload and 7 cases of proven absence of iron burden (data on remaining patients are pending). In the alternative donor HSCT pool prior to 2000, 5 of 13 had proven pre-HSCT iron overload and 1 without (data on remaining 7 patients are pending). Since the year 2000, of the alternative donor HSCT, there were 2 proven cases with iron overload and 2 with proven absence of iron burden (data on 6 remaining patients are pending). Overall survival at 5 years for patients without evidence of iron overload is 53.3 ± 18.6% and for those with iron overload pre-transplant is 38.5 ± 23.3 (p=0.97). ConclusionsThe best outcomes for stem cell transplantation in DBA patients occur when using HLA-matched sibling donors in younger patients. There has been a significant improvement in survival with alternative donor SCT since the year 2000 which may be a result of improved HLA matching techniques. There is a trend to improved survival with lower pre-transplant iron burden. DisclosuresNo relevant conflicts of interest to declare.

Clinical “cytokine storm” as revealed by monocyte intracellular flow cytometry: correlation of tumor necrosis factor α with severe gut graft-versus-host disease

Background & Aims: Gut graft-versus-host disease (GVHD) contributes significantly to lethality after allogeneic hematopoietic stem-cell transplantation (HSCT). In murine models, macrophage secretion of interleukin 1α (IL-1α) and tumor necrosis factor α (TNF-α) contributes to gut GVHD pathogenesis. To help characterize whether human gut GVHD has similar biological characteristics, monocyte IL-1α and TNF-α production were evaluated after HSCT. Methods: Patients with refractory hematologic malignancy (n = 17) underwent reduced-intensity conditioning, HLA-matched sibling HSCT, and cyclosporine A GVHD prophylaxis. After HSCT, monocyte IL-1α and TNF-α levels were measured using intracellular flow cytometry (IC-FCM), and results were correlated with clinical GVHD. Results: Incidences of acute GVHD were none (n = 3), grades I-II (n = 9), or grades III-IV (n = 5; each case with stage 2–3 gut GVHD). Posttransplantation monocyte IL-1α production (percentage of CD14 +IL-1 + cells) increased significantly from 8.7% ± 3.7% (week 2) to 40.3% ± 7.3% (week 4; P = 0.0065) and was not associated with GVHD severity ( P = 1.00). Conversely, increases in monocyte TNF-α were quantitatively reduced and temporally delayed, from 0.6% ± 0.2% (week 2) to 3.6% ± 1.4% (week 6; P = 0.076). Most importantly, elevation of monocyte TNF-α level correlated with increased gut GVHD severity ( P = 0.0041); increases in monocyte TNF-α levels typically preceded the onset of gut GVHD symptoms. Conclusions: Human gut GVHD after reduced-intensity allogeneic HSCT is associated with monocyte cytokine secretion initially involving IL-1α, followed by TNF-α. Serial measurement of monocyte cytokines, in particular, TNF-α, by IC-FCM may represent a noninvasive method for GVHD monitoring, potentially allowing the identification of patients appropriate for early-intervention strategies.

The impact of donor KIR and patient HLA-C genotypes on outcome following HLA-identical sibling hematopoietic stem cell transplantation for myeloid leukemia

AbstractKiller immunoglobulin–like receptors (KIRs) regulate cell activity of natural killer (NK) cells and some T cells. The predominant ligand for inhibitory KIRs is HLA-C, which subdivides into 2 groups based on the specificity of inhibitory KIRs. The ligands for activatory KIRs are unknown. Following hematopoietic stem cell transplantation (HSCT), recipient tissues may not express a ligand for KIRs present within the graft, and the combination of donor KIR and recipient HLA-C types could influence outcome. HLA and KIR genotypes were determined in 220 donor-recipient pairs from HLA-matched sibling HSCTs performed for myeloid (n = 112) and lymphoid (n = 108) diseases. In HSCTs performed for myeloid disease, overall survival was worse in patients homozygous for group 2 HLA-C (C2) than in patients who carried a group 1 HLA-C (C1) allele (P < .005). Moreover, this effect is seen only when the donor additionally carries the activating KIR gene KIR2DS2 (P = .045). No effect was seen in patients with lymphoid disease. Thus, in HLA-matched sibling HSCT for myeloid leukemia, patients homozygous for C2 alleles receiving a graft from a donor carrying the KIR gene KIR2DS2 have a significantly reduced chance of survival.