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Subsequent neoplasms (SNs) compromise long-term survivors after hematopoietic cell transplantion. We performed a retrospective analysis of SNs in a cohort of 10358 recipients of umbilical cord blood transplantation (UCBT) reported to Eurocord/EBMT registries from 1988 to 2018. A total of 233 patients developed SNs. Median age at UCBT was 31 years (y) (0.3-69), and 84 were pediatric patients. Indications for UCBT were malignant hematological diseases in 199 patients (85%). Three groups of SNs were observed. Post-transplant lymphoproliferative disorders (PTLD) were reported in 145 patients in a median of 4 months after UCBT. Of these, 9/145 patients died from relapse, 83/145 from PTLD, and 24/145 from transplant-related causes. At last follow-up, 29/145 were alive; 5y-overall survival (OS) after PTLD diagnosis was 21±3%. Acute leukemia / myelodysplasia (AL/MDS) was diagnosed in 23 patients in a median of 28 months after UCBT and included 3 donor-cell AL. Four of 23 patients died from relapse of primary disease, 8/23 from progression of SNs, and 4/23 from TRM. Seven patients were alive at last follow-up; 5y-OS after AL/MDS diagnosis was 36±10%. Solid tumors (ST) were reported in 65 patients in a median of 54 months after UCBT. Most common tumor sites were lung, thyroid, bone and soft tissue. A total of 33/65 patients died (26 due to ST, 6 to relapse of primary disease, 1 cause missing). At last follow-up, 32/65 patients were alive; 5y-OS after the diagnosis of ST was 51±6%. In conclusion, despite their poor outcomes, SNs that occur after UCBT are extremely rare. Identification of associated risk factors and early detection may help to improve OS.

Four decades ago, Broxmeyer et al. demonstrated that umbilical cord blood (CB) contained hematopoietic stem cells (HSC) and hypothesized that CB could be used as a source of donor HSC for rescue of myeloablated bone marrow. In 1988, Gluckman et al. reported the first successful matched sibling cord blood transplant (CBT) in a child with Fanconi Anemia. In 1991, Rubinstein et al. established an unrelated donor CB bank, and in 1993, the first unrelated CBT used a unit from this bank. Since that time, >40 000 CBTs have been performed worldwide. Early outcomes of CBT were mixed and demonstrated the importance of cell dose from the CB donor. We hypothesized that improvements in CB banking and transplantation favorably impacted outcomes of CBT today and performed a retrospective study combining data from Eurocord and Duke University in 4834 children transplanted with a single unrelated CB unit (CBU) from 1993 to 2019. Changes in standard transplant outcomes (overall survival [OS], disease free survival [DFS], acute and chronic graft-versus-host disease [GvHD], treatment related mortality [TRM], and relapse) over 3 time periods (1: 2010 to 2019) were studied. Increased cell dose and degree of HLA matching were observed over time. OS, times to engraftment, and DFS improved over time. The incidence of TRM and GvHD decreased while the incidence of relapse remained unchanged. Relative contributions of cell dose and HLA matching to transplant outcomes were also assessed and showed that HLA matching was more important than cell dose in this pediatric cohort.

In single unrelated cord blood transplantation (UCBT), an increasing number of HLA allele mismatches (MM) has been associated with inferior overall survival (OS) attributed to higher transplant-related mortali-ty (TRM). Previous studies on the role of allele-level HLA matching following double UCBT (dUCBT) showed conflicting results. Herein we report the impact of allele-level HLA matching on outcomes of a large dUCBT cohort . We included 963 adults with hematologic malignancies, with available allele-level HLA matching at HLA-A, -B, -C and -DRB1, receiving dUCBT between 2006-2019. Assignment of donor-recipient HLA match was performed considering the unit with the highest disparity with the recipient. 392 patients received dUCBT with 0-3 and 571 with ≥4 allele MM. Day-100 and 4-year TRM were 10% and 23%, for recipients of dUCBT with 0-3 MM compared to 16% and 36% for those with ≥4 MM (HR 1.58, p=.002; and HR 1.54, p=.002), respectively. Higher degree of allele MM was also associated with worse neutrophil recovery and lower incidence of relapse; no significant effect on graft-versus-host disease was observed. Patients receiving units with 0-3 MM had a 4-year OS of 54% versus 43% for those with ≥4 MM (HR 1.40, p=.005). The inferior OS associated with higher HLA disparity was only partially mitigated by increased total nucleated cell doses. Our results confirm that allele-level HLA typing is a significant factor for OS following dUCBT and units with ≥4 MM (≤4/8 HLA-matched) should be avoided if possible.

The association between acute lymphoblastic leukaemia (ALL) and acute myeloid leukaemia (AML) and the human leukocyte antigens (HLA) has rarely been studied in terms of diversity of peptide-binding pockets. The objective of this study was to analyse whether motifs of HLA class I and class II peptide-binding pockets and/or their amino acid positions were differentially associated with ALL and AML. We included 849 patients from the Eurocord/European Blood and Marrow Transplant registry. The HLA peptide-binding pockets whose amino acid variability was analysed were B and F for HLA class I, P4, P6, and P9 for HLA-DRB1, and P4 and P9 for HLA-DQB1. The motif RFDRAY in P4 of HLA-DRB1*16:01/02/03/05 alleles and the motif YYVSY in P9 of HLA-DQB1*05:02/04/05 alleles, were statistically associated with ALL (corrected p value [p

We developed a risk score to predict event-free survival (EFS) after allogeneic hematopoietic cell transplantation for sickle cell disease. The study population (n = 1425) was randomly split into training (n = 1070) and validation (n = 355) cohorts. Risk factors were identified and validated via Cox regression models. Two risk factors of 9 evaluated were predictive for EFS: age at transplantation and donor type. On the basis of the training cohort, patients age 12 years or younger with an HLA-matched sibling donor were at the lowest risk with a 3-year EFS of 92% (score, 0). Patients age 13 years or older with an HLA-matched sibling donor or age 12 years or younger with an HLA-matched unrelated donor were at intermediate risk (3-year EFS, 87%; score, 1). All other groups, including patients of any age with a haploidentical relative or HLA-mismatched unrelated donor and patients age 13 years or older with an HLA-matched unrelated donor were high risk (3-year EFS, 57%; score, 2 or 3). These findings were confirmed in the validation cohort. This simple risk score may guide patients with sickle cell disease and hematologists who are considering allogeneic transplantation as a curative treatment relative to other available contemporary treatments.

Background : Graft failure (GF) is a life-threatening complication after allogeneic hematopoietic stem cell transplantation (HCT). In the absence of autologous recovery, a second HCT is necessary to attempt preventing death due to prolonged pancytopenia. Previous studies describing outcomes of second HCT performed after a GF with different types of donor sources report a wide range of overall survival (OS) and transplant related mortality (TRM), however studies including a large number of patients receiving a second transplant with umbilical cord blood as the graft source are scarce. Objective : To describe umbilical cord blood transplants (UCBT) performed after GF following a previous HCT. Study Design : Retrospective registry-based study, using data extracted from Eurocord and the European Society for Blood and Marrow Transplantation (EBMT) databases. The study reports outcomes of 247 umbilical cord blood transplants (UCBT), performed in EBMT transplant centers, after GF following a previous HCT. Data were analyzed separately for patients with malignant (n=141) and non-malignant diseases (n=106). Results : The most frequent HCT that resulted in GF was also UCBT (65.0% and 68.9%), and most GF occurred within 100 days after transplantation (92.3% and 85.9%), for malignant and non-malignant diseases, correspondingly. Median follow-up was 47 for surviving patients with malignant and 38 months for those with non-malignant diseases. We observed a similar cumulative incidence of neutrophil engraftment of 59.1% (95% CI 51.4- 67.9%) and 60.4% (95% CI 51.7- 70.6%), in a median time of 23 and 24 days for malignant and non-malignant diseases, respectively. The 3-year OS was 28.9% (95% CI 21.8- 37.3%) in the malignant disease group and 49.1% (95% CI 39.5- 58.8%) in the non-malignant. In patients with malignancies, TRM was 39.9% (95%CI 32.5 - 49.1%) at 100 days and malignant 57.5% (95%CI 49.4 - 66.8%) at 3 years. In multivariate analyses, none of the characteristics studied were statistically significantly associated with engraftment or overall survival. Conclusion : Although survival for patients requiring a second transplant is not optimal, UCB remains a valid life-saving option for patients with GF.

Sickle cell disease (SCD) is the most common inherited hemoglobinopathy. Hematopoietic stem cell transplantation (HCT) is the sole curative therapy for SCD, but few patients will have a matched sibling donor. Patients with SCD are mostly of African origin and thus are less likely to find a matched unrelated donor in international registries. Using HaploStats, we estimated HLA haplotypes for 185 patients with SCD (116 from a Brazilian center and 69 from European Society for Blood and Marrow Transplantation [EBMT] centers) and classified the ethnic origin of haplotypes. Then we assessed the probability of finding an HLA-matched unrelated adult donor (MUD), considering loci A, B, and DRB1 (6/6), in international registries. Most haplotypes were African, but Brazilians showed a greater ethnic admixture than EBMT patients. Nevertheless, the chance of finding at least one 6/6 potential allelic donor was 47% for both groups. Most potential allelic donors were from the US National Marrow Donor Program registry and from the Brazilian REDOME donor registry. Although the probability of finding a donor is higher than previously reported, strategies are needed to improve ethnic diversity in registries. Moreover, predicting the likelihood of having an MUD might influence SCD management., SCOPUS: ar.j, info:eu-repo/semantics/published

Sickle cell disease (SCD) affects more than 300,000 children annually worldwide. Despite improved supportive care, long-term prognosis remains poor. Allogeneic hematopoietic cell transplantation (allo-HCT) is the sole validated curative option, resulting in sustained resolution of the clinical phenotype. The medical literature on allo-HCT for SCD is largely limited to children. Recent studies have evaluated allo-HCT efficacy in adults. Here, we conducted a systematic review/meta-analysis to assess the totality of evidence on the efficacy, or lack thereof, of allo-HCT in treating SCD. We performed a comprehensive literature search using PubMed/Medline, Embase, and Cochrane library databases on November 13, 2019. Four authors independently extracted data on clinical outcomes related to benefits (overall survival [OS] and disease-free survival [DFS]) and harms (acute graft-versus-host disease [aGVHD], chronic graft-versus-host disease [cGVHD], nonrelapse mortality [NRM], and graft failure [GF]). Our search identified a total of 1906 references. Only 33 studies (n= 2853 patients) met our inclusion criteria. We also performed a subset analysis by age. Analyses of all-age groups showed pooled rates of 96% for OS, 90% for DFS, 20% for aGVHD, 10% for cGVHD, 4% for NRM, and 5% for GF. In the pediatric population, pooled rates for OS, DFS, aGVHD, cGVHD, NRM, and GF were 97%, 91%, 26%, 11%, 5%, and 3%, respectively. In adults, pooled rates for OS, DFS, aGVHD, cGVHD, NRM, and GF were 98%, 90%, 7%, 1%, 0%, and 14%, respectively. Our data show that allo-HCT is safe and effective, yielding pooled OS rates exceeding 90%. The high GF rate of 14% in adults is concerning and emphasizes the need to evaluate new strategies.

We report the results of an analysis of unrelated allogeneic hematopoietic stem cell transplantations (HSCT) in 71 patients with sickle cell disease (SCD) transplanted in EBMT centers between 2005 and 2017. Median age was 9.3 years; graft type was bone marrow in 79% and peripheral blood in 21%. Recipient-donor HLA match at high resolution typing was 10/10 in 31, 9/10 in 20, and 8/10 in 4 patients; the other patients had intermediate resolution typing. The most frequent conditioning regimens were fludarabine-thiotepa-treosulfan (64%) or busulfan-cyclophosphamide (12%). Cumulative incidence of neutrophil engraftment was 92%; platelet engraftment was 90%. Eleven patients (15%) experienced graft failure. Grade II-IV acute graft-vs.-host disease (GvHD) was 23%; 3-year chronic GvHD was 23%. Three-year overall survival (OS) was 88 +/- 4%. GRFS was 62 +/- 6%. HLA matching was the most significant risk factor for OS: 3-year OS was 96 +/- 4% in 10/10 group vs. 75 +/- 10% in 9-8/10 (p = 0.042); GRFS was 69 +/- 9% vs. 50 +/- 12% (p = 0.114), respectively. In conclusion, unrelated donor HSCT is a valid option for SCD patients who lack an HLA-identical sibling donor, preferably in the context of clinical trials. Using a 10/10 HLA-matched unrelated donor yields better survival indicating that HLA matching is an important donor selection factor in this nonmalignant disease.

Graft failure (GF) is a life-threatening complication after allogeneic hematopoietic stem cell transplantation (HCT). The incidence of GF after HCT using human leukocyte antigen (HLA) matched donors is lower than 5%; however, a higher incidence is observed when considering HLA mismatched donors or alternative donor sources and the use of reduced intensity conditioning (RIC) regimens. In the absence of autologous recovery, a second HCT is necessary to attempt preventing death due to prolonged pancytopenia. Previous studies describing outcomes of second HCT performed after a GF with different types of donor sources report a wide range of overall survival (OS) and transplant related mortality (TRM), however studies including a large number of patients receiving a second transplant with umbilical cord blood (UCB) are scarce. In this retrospective study, using registry data from Eurocord and the European Society for Blood and Marrow Transplantation (EBMT), we report the outcomes of 247 umbilical cord blood transplants (UCBT) performed after GF following a previous HCT. Data were analyzed separately for patients with malignant (n=141) and non-malignant diseases (n=106). UCB was the most frequent stem cell source that resulted in GF (65.0% and 68.9%, for malignant and non-malignant diseases, respectively), and most of the reported GF occurred within 100 days after HCT (92.3% and 85.9%). The most frequent diagnosis in malignant diseases was acute leukemia (64.5%), mainly acute myeloid leukemia (AML). In non-malignant diseases, the diagnoses most often reported were inborn error of metabolism (IEM) (37.7%), followed by bone marrow failure syndrome (BMF) (34.0%) and primary immune deficiency (PID) (21.7%). In malignant and non-malignant diseases, we observed a similar cumulative incidence of neutrophil engraftment of 59.1% (95% CI 51.4- 67.9%) and 60.4% (95% CI 51.7- 70.6%), in a median time of 23 and 24 days, and a 3-year OS of 28.9% (95% CI 21.8- 37.3%) and 49.1% (95% CI 39.5- 58.8%). TRM at 100 days and at 3 years for malignant diseases was 39.9% (95%CI 32.5 - 49.1%), and 57.5% (95%CI 49.4 - 66.8%). In multivariate analyses, none of the characteristics were statistically significantly associated with engraftment or overall survival. Although survival for patients requiring a second transplant is not optimal, UCB remains a valid life-saving option for some patients with GF as it is an immediately available source of stem cells in severely ill patients. The most appropriate stem cell source for salvage transplants remains controversial, as it depends on multiple factors including center choice, donor availability, underlying disease and existing comorbidities. Disclosures Peffault De Latour: Pfizer: Membership on an entity's Board of Directors or advisory committees, Other: Travel support, Research Funding; Amgen: Consultancy, Other, Research Funding; Jazz Pharmaceuticals: Honoraria; Alexion, AstraZeneca Rare Disease: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel support, Research Funding.

Four decades ago, Hal Broxmeyer demonstrated that umbilical cord blood (CB) contained hematopoietic stem cells (HSC) and hypothesized that CB could be used as a source of donor HSC for rescue of myeloablated bone marrow. In 1988, Gluckman et al reported the first successful cord blood transplant (CBT) of a child with Fanconi Anemia using matched sibling CB. This patient survives and 35 years later still has durable hematopoiesis from the CB donor graft. In 1991, Rubinstein et al established an unrelated donor (UD) CB bank and in 1993 the first UD CBT was using a unit from this bank. Since that time, >40,000 CBTs have been performed worldwide. We hypothesized that changes in cord blood banking (increased size, diversity, and quality of banked units enabling selection of units with higher cell doses and closer HLA matching) and in transplantation (less use of steroids, availability of newer therapies for prophylaxis and treatment of graft versus host disease [GVHD], improved antifungal and antiviral detection and therapeutics) have improved outcomes of CBT today. To address this hypothesis, we performed a retrospective study combining data from Eurocord and Duke University in a large cohort of children transplanted with a single UD CB unit (CBU) from 1993-2019. Standard transplant outcomes (overall survival [OS], disease free survival [DFS], acute and chronic GVHD, treatment related mortality [TRM], and relapse) and changes in outcomes over 3 time periods (1:2010, n=1802) were studied. Relative contributions of cell dose and HLA matching to transplant outcomes over time were assessed. A total of 4834 patients (4015 from Eurocord and 819 from Duke) were analyzed. The majority of patients, (59%, n=2839) had malignant diagnoses including 1422 with ALL, 887 with AML and 167 with MDS. Of the 1995 with non-malignant diagnoses, 761 had inborn errors of metabolism, 644 had primary immunodeficiency, 325 had a bone marrow failure syndrome and 206 had a histiocytic disorder. Half of the patients had positive serologies for CMV prior to transplant. The median age of the cohort fell from 5.2 to 3.25 years over time. In patients with malignancies, use of total body irradiation decreased over time. The median total nucleated cell (TNC) and CD34+ cell doses administered were 8.07x10e7 and 6.17x10e5 cells/kg and increased over time. HLA matching and transplantation of patients in earlier disease states also increased over time, p In conclusion, we analyzed the largest cohort of pediatric patients undergoing CBT over the past 3 decades. OS, DFS and engraftment have improved over time accompanied by decreases in TRM and acute GVHD. Relapse and chronic GVHD were stable and remain low. These improvements are explained by the increased availability of high quality banked CBUs enabling selection of closer HLA matching and units with higher cell doses. The numbers of CBTs have decreased in the past decade, but these results support the ongoing use of CBT in children lacking matched related or unrelated donors. Figure 1 Figure 1. Disclosures Kurtzberg: Neurogene: Consultancy; CryoCell: Patents & Royalties: Duke licensed IP, and data and regulatory packages for manufacturing and use of cord blood and cord tissue MSCs in the treatment of patients with hypoxic ischemic encephalopathy, cerebral palsy, autism, acute ischemic stroke, COVID-ARDS, and COVID-MIS-C. ; Sinocell: Patents & Royalties: Duke licensed IP, data, and regulatory packages for use of autologous and sibling cord blood to treat children with cerebral palsy.; Celularity: Current holder of stock options in a privately-held company. Troy: SinoCell: Patents & Royalties; CryoCell: Patents & Royalties; Bristol Myers Squibb: Research Funding; Synthetic Biologics: Honoraria; Gamida Cell: Consultancy; The EMMES Corporation: Consultancy; The Community Data Roundtable: Consultancy; AegisCN: Consultancy.

Efforts to implement family cord blood banking have been developed in the past decades for siblings requiring stem cell transplantation for conditions such as sickle cell disease. However, public banks are faced with challenging decisions about the units to be stored, discarded, or used for other endeavors. We report here 20 years of experience in family cord blood banking for sickle cell disease in two dedicated public banks. Participants were pregnant women who had a previous child diagnosed with homozygous sickle cell disease. Participation was voluntary and free of charge. All mothers underwent mandatory serological screening. Cord blood units were collected in different hospitals, but processed and stored in two public banks. A total of 338 units were stored for 302 families. Median recipient age was six years (11 months-15 years). Median collected volume and total nucleated cell count were 91 mL (range 23–230) and 8.6×108 (range 0.7–75×108), respectively. Microbial contamination was observed in 3.5% (n=12), positive hepatitis B serology in 25% (n=84), and homozygous sickle cell disease in 11% (n=37) of the collections. Forty-four units were HLA-identical to the intended recipient, and 28 units were released for transplantation either alone (n=23) or in combination with the bone marrow from the same donor (n=5), reflecting a utilization rate of 8%. Engraftment rate was 96% with 100% survival. Family cord blood banking yields good quality units for sibling transplantation. More comprehensive banking based on close collaboration among banks, clinical and transplant teams is recommended to optimize the use of these units.

We report the results of an analysis of unrelated allogeneic hematopoietic stem cell transplantations (HSCT) in 71 patients with sickle cell disease (SCD) transplanted in EBMT centers between 2005 and 2017. Median age was 9.3 years; graft type was bone marrow in 79% and peripheral blood in 21%. Recipient-donor HLA match at high resolution typing was 10/10 in 31, 9/10 in 20, and 8/10 in 4 patients; the other patients had intermediate resolution typing. The most frequent conditioning regimens were fludarabine-thiotepa-treosulfan (64%) or busulfan-cyclophosphamide (12%). Cumulative incidence of neutrophil engraftment was 92%; platelet engraftment was 90%. Eleven patients (15%) experienced graft failure. Grade II-IV acute graft-vs.-host disease (GvHD) was 23%; 3-year chronic GvHD was 23%. Three-year overall survival (OS) was 88 ± 4%. GRFS was 62 ± 6%. HLA matching was the most significant risk factor for OS: 3-year OS was 96 ± 4% in 10/10 group vs. 75 ± 10% in 9-8/10 (p = 0.042); GRFS was 69 ± 9% vs. 50 ± 12% (p = 0.114), respectively. In conclusion, unrelated donor HSCT is a valid option for SCD patients who lack an HLA-identical sibling donor, preferably in the context of clinical trials. Using a 10/10 HLA-matched unrelated donor yields better survival indicating that HLA matching is an important donor selection factor in this nonmalignant disease.

Allogeneic hematopoietic stem cell transplantation (HSCT) is, to date, the only curative treatment for sickle cell disease (SCD). Because an human leukocyte antigen (HLA)-matched sibling donor is not always available, alternative stem cell sources such as unrelated or haploidentical related donors have been explored. To date, few series of SCD patients transplanted with an unrelated donor, cord blood, and haploidentical related donor have been reported, but the high rates of rejection and chronic graft versus host disease have limited their widespread application. We describe the outcomes of a retrospective, registry-based, survey on 144 alternative donor HSCT performed for SCD in 30 European Society for Blood and Marrow Transplantation centers between 1999 and 2017. Data on 70 unrelated adult donors (49%), six cord blood (4%), and 68 haploidentical donors (47%; including post-HSCT Cy, ex vivo T-cell depleted, and other haplo-HSCTs) were reported and missing information was updated by the centers. Overall, 16% patients experienced graft failure, Grade II–IV acute GVHD at 100 days was 24%, whereas Grade III–IV was 10%. Chronic GVHD was observed in 24% (limited for 13 patients and extensive for 18 patients). Overall, the 3-year overall survival (OS) was 86% ± 3% and 3-year event-free survival (EFS; considering death and graft failure as events) was 72% ± 4%. We therefore conclude that alternative donor HSCT for SCD can be feasible but efforts in decreasing relapse and GVHD should be promoted to increase its safe and successful utilization. Moreover, a better knowledge of HLA matching and the tailoring of conditioning could help improve EFS and OS.

Despite adequate immunization and penicillin prophylaxis, bacterial infections remain a leading cause of morbidity and mortality in patients with sickle cell disease (SCD). Besides hyposplenism, inflammatory and genetic factors might modulate their susceptibility to bacterial infections. We performed a candidate gene association of single nucleotide polymorphisms (SNPs) located in Toll-like receptor (TLR) genes, encoding prominent molecules for innate immune responses, with the occurrence of bacterial infections in patients with SCD. A cohort followed in centres in Brazil, France and Senegal (n = 430) was divided in two groups: patients who presented at least one episode of bacterial infection (n = 235) and patients who never had bacterial infections (n = 195). There were no differences in gender or age distribution among the groups. The frequency of the TLR2 rs4696480 TA genotype was significantly lower in the infected group (50% vs. 67%, odds ratio [OR] = 0·50, 95% confidence interval [CI] 0·34-0·75, P

Hematopoietic stem-cell transplantation (HSCT) is currently the only curative therapeutic option for the treatment of thalassemia. In spite of the high cure rate, HSCT can lead to life-threatening adverse events in some patients. Busulfan (Bu) is a key component of the conditioning regimen prior to HSCT. Inter-individual differences in Bu pharmacokinetics (PK) are hypothesized to influence Bu efficacy and toxicity. Since Bu is mainly metabolized by glutathione S-transferase (GST), we investigated the relationship of GSTA1 and GSTM1 genotypes with first-dose PK and HSCT outcomes in 44 children with thalassemia intermedia and thalassemia major. All children received a myeloablative conditioning regimen with IV Bu. Association analysis revealed a relationship between GSTA169C>T (or haplotype *A/*B) and first Bu dose PK that was dependent on sex and Pesaro risk classification (PRC). Among female patients and patients with PRC I–II, homozygous individuals for the GSTA1T−69 allele defining haplotype *B, had higher Bu exposure and lower clearance (P⩽0.01). Association with HSCT outcomes showed that patients with the GSTM1 null genotypes had higher occurrence of regimen-related toxicity (P=0.01). These results suggest that GST genotypes could be useful to tailor the first Bu dose accordingly to improve HSCT outcome.

Background: Patients (pts) with sickle cell disease (SCD) show a high phenotype variability that is not fully understood. Because inflammation plays a major role in SCD pathophysiology, we hypothesized that single nucleotide polymorphisms (SNPs) in genes of innate and adaptative inflammatory response modulate the occurrence of SCD complications. Objective: to establish associations between SNPs and clinical complications in pts with SCD. Methods: Case-control retrospective study; 500 pts were included, followed at Senegal (n=56), Brazil (n=230) and France (n=214). We analyzed the effect of 20 SNPs in 6 clinical complications: acute chest syndrome (ACS), stroke, leg ulcers, cholelithiasis, osteonecrosis and retinopathy. Using TaqMan 5'-nuclease assay, we genotyped SNPs in genes encoding Toll-like receptor (TLR) 1 (rs4833095), 2 (rs4308099, rs4308100, rs4696480), 6 (rs5743810), 10 (rs11466653, rs11096957), natural killer (NK) group 2 member D (NKG2D) receptor (rs1982536, rs2617160, rs2617169, rs2617170, rs2617171, rs1049174, rs2246809, rs2255336), human leukocyte antigen (HLA)-G (rs9380142), HLA-E (rs2517523), major histocompatibility complex class I polypeptide-related sequence A (MICA, rs1051792) and cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) (rs5742909, rs231775). All SNPs had a call rate >90% and minimum allele frequency >1%. We performed analyses of correspondence for indicating associations between SNPs and number of clinical complications. Logistic regressions were used to identify associations between SNPs and each complication, using geographical origin, SCD genotype, rate of transfusions and gender for modeling adjustment; the significance was adjusted for multiple testing using false discovery rate. Comparisons of genotype frequencies with the population of African descent from 1000 genomes database were done by chi-square. Results: Pts were originally from Brazil (n=228), Sub-Saharan Africa (n=200), French West Indies (n=53), North Africa (n=7) and 12 unknown. SCD genotype (available n=498) was SS (n=402), SC (n=46), SB (n=42), SD or SE (n=4). 280 pts were female; median age was 32 years (range 0-69); 184 pts received at least 20 transfusions. 71 pts presented stroke, 200 had at least 1 episode of ACS, 69 had leg ulcers, 271 had cholelithiasis, 150 had retinopathy and 90 osteonecrosis. 97 pts did not present complications, 135 had 1 type of clinical complication, 134 had 2, 94 had 3, 34 had 4 and 6 had 5. 21 pts underwent hematopoietic stem cell transplantation. 11 pts died during follow-up, mostly from ACS and hemorrhagic stroke. Indication of association with number of complications: TLR2 rs4696480 TA, TLR2rs3804099 CC and HLA-Grs9380142 AA were associated with occurrence of 0-1 clinical complications, MICA rs1051792 AA/AG with up to 2 complications and NKG2D rs2617169 AA with 5 complications. Association between genotypes/haplotypes and each complication: no association was found between SNPs and stroke, ACS, leg ulcers and osteonecrosis. Rs9380142 was the only SNP significantly associated with cholelithiasis in the logistic regression additive model. The G allele increased the risk of cholelithiasis (AG x AA, OR 1.57, 95%CI 1.16-2.15; GGxAA, OR 2.47, 95%CI 1.34-4.64; P=0.02). For retinopathy, in the logistic regression additive model, the presence of the A allele decreased the risk of retinopathy for rs2246809 in pts of same origin (AAxGG: OR 0.22, 95%CI 0.09-0.50; AGxGG: OR 0.47, 95%CI 0.31-0.71; P=0.004), rs2617160 (ATxTT: OR 0.67, 95%CI 0.48-0.92; AAxTT: OR 0.45, 95%CI 0.23-0.84; P=0.04) and rs2617169 in pts of same SCD genotype (AAxTT: OR 0.33,95%CI 0.13-0.82; ATxTT: OR 0.58, 95%CI 0.36-0.91, P=0.049). No haplotype was associated with complications. The genotype distribution of SNPs rs4696480, rs3804099, rs1051792 and rs2617169 differed significantly from the population of African descent from the 1000 genomes database. Discussion: We have previously shown that TLR2 rs4696480 TA decreases occurrence of bacterial infections in SCD; in this study, TA is also associated with less complications. Also, HLA-G rs9380142 AA had less complications and cholelithiasis, and 3 SNPs in NKG2D modulated occurrence of retinopathy. TLR and NKG2D may be activated by heat shock proteins, released in ischemia-reperfusion injury that occurs in SCD. Our findings help to better understand the role of inflammation in phenotype heterogeneity in SCD. Disclosures No relevant conflicts of interest to declare.

Introduction: Allogeneic hematopoietic stem cell transplantation (HSCT) is, to date, the only curative treatment for sickle cell disease (SCD). Because a human leukocyte antigen (HLA) matched sibling donor is not always available, alternative stem cell sources such as unrelated or haploidentical related donors have been explored. The likelihood of finding a 10/10 (HLA-A, B, C, DRB1 and DQB1) matched donor varies among ethnic groups, with the lowest probability among individuals of African descent. To date, few series of SCD patients transplanted with an unrelated donor (UD) have been reported, but the high rates of rejection and chronic graft versus host disease (cGvHD) have limited its widespread application. Patients and methods: We report the results of a retrospective, registry based, survey on 70 UD HSCT performed in patients (pts) with SCD from UD in 22 European Society for Blood and Marrow Transplantation (EBMT) centers between 2005 and 2017. Data were collected from the EBMT database and missing information was updated by the centers. Median follow up was 38 (range 2-154) months. Most pts were HbSS (n=54; 78%), had positive serology for CMV (80%), and a Karnofsky score >80% (98%). Eighteen pts had a major ABO incompatibility. Recurrent vaso-occlusive crisis (n=58), cerebral vasculopathy (n=23) and acute chest syndrome (n=24) were the main indications for HSCT. Red blood cell (RBC) transfusions pre-HSCT were reported in 97% of pts of whom 53% received more than 20 transfusions; 14% of the transfused pts had RBC alloantibodies. Hydroxyurea pre-HSCT was used in 65% of pts. Median age at HSCT was 9.6 years (range 2-43) with 87% of pts being ≤ 16 years. Stem cell source was bone marrow (BM) in 55 pts (79%) and peripheral blood (PBSC) in 15 (21%). The median number of infused TNC /kg was 3.6 x 108 for BM and 7.1 x 108 for PBSC; the median number of infused CD34/kg was 4.4 x 106 for BM and 8.3 x 106 for PBSC. HLA matching at high resolution typing was 10/10 (HLA-A, B, C, DRB1 and DQB1) in 31, 9/10 in 17 and 8/10 in 4 of the patient-donor pairs; intermediate resolution typing was available for 10 (10/10 or 9/10) and the HLA information was missing for the remaining 8 patient-donor pairs. The most frequent conditioning regimens were fludarabine-thiotepa-treosulfan (64%) and busulfan- cyclophosphamide (12%). GvHD prophylaxis was cyclosporine plus methotrexate in 59%. Anti-thymocyte globulin was used in 90% and alemtuzumab in 9% of pts. Results: The cumulative incidence (CI) of neutrophil engraftment at 60 days was 93% (95% CI 76-100), with median time to engraftment of 18 days; platelet engraftment at 180 days was 90% (95% CI 83-98) with a median time of 20 days. Ten pts had graft failure (5 primary and 5 secondary) of whom 6 had a second transplant and were all alive at last FU (median 9.5 months after second HSCT). The CI of grade II-IV aGVHD at 100 days was 23% (95% CI 15-36), and 8 pts (11%) had grade III-IV. Acute GVHD was more frequent in patients who received PBSC (PBSC 42.9%, BM 18.2%, p=0.062). Three-year CI of cGVHD was 23% (95% CI 15-36), 7 pts (10%) had limited and 9 (13%) extensive cGvHD. Three-year overall survival (OS) was 90±4%; three-year event free survival (EFS) (considering death and graft failure as events) was 76±6%; HLA matching between donor and recipient was the most important factor for OS and EFS. Considering only pts-donor pairs with high resolution HLA typing available (n=52), 3-year OS was 96±4% in 10/10 group compared to 77±11% in 9/10 plus 8/10 group (p 0.065), 3-year EFS was 85±7% vs 62±12% (p 0.040), respectively. No significant differences between the groups were observed in CI of neutrophil engraftment, aGVHD and cGVHD. Conclusion: UD HSCT is a valid option for SCD patients who lack an HLA-identical sibling donor. Nevertheless, efforts are still needed to improve outcomes after UD HSCT. Our results indicate that using a 10/10 HLA matched UD improves both OS and EFS compared to donors with 1 or more mismatches; so, when such a matched unrelated donor is not found, using an haplo relative or an unrelated cord blood as donor source should be evaluated. A prospective trial is in preparation to evaluate the use of haploidentical donors for HSCT in SCD (EudraCT number: 2018-002652-33). Disclosures No relevant conflicts of interest to declare.

Background: Hematopoietic stem cell transplantation (HSCT), the only available curative therapy for sickle cell disease (SCD), remains hampered by the lack of histocompatible stem cell donors. Most patients will not have a suitable human leukocyte antigen(HLA) matched sibling donor. In addition, SCD affects ethnic groups that are underrepresented in stem cell donor registries worldwide. Objective: to assess the probability of having a potential allelic HLA matched unrelated donor(MUD) for HLA-A,B and DRB1 loci (6/6) in international donor registries for patients with SCD. Methods: 185 patients with SCD were included, 116 from Brazil, of whom 23 underwent HSCT, and 69 patients who underwent HSCT in centers reporting to the European Society for Blood and Marrow Transplantation (EBMT). All patients had HLA typing available at intermediate or high resolution. For intermediate resolution, using the National Marrow Donor Program (NMDP) code, we assigned alleles based on allele frequencies.We performed HLA haplotype estimation using the HaploStats website, which describes HLA haplotype frequency from the NMDP registry for the following ethnic groups: Caucasian, African-American, Asian, Hispanic and Native American. Because Hispanic is not a primary ethnicity, we did not consider this group in our analyses. Based on haplotype frequency of each ethnic group, we defined the most likely ethnicity for each estimated haplotype; those with frequency >1:1000 in all ethnic groups were named common. Unrelated donor search was done using the World Marrow Donor Association (WMDA) algorithm, which is based on haplotype matching. A potential allelic donor was defined as a full match high resolution 6/6 donor. Because it is described that testing at least 5 potential allelic donors simultaneously increases the chances of having a real donor, we assessed the probability of finding at least 1 and at least 5 potential allelic donors. Patients who received HSCT from MUD were excluded from donor searches (n=10).Comparisons of probabilities of having potential allelic donors between Brazilian and EBMT cohorts were performed by chi-square. Results: In the Brazilian cohort, from 181 HLA haplotypes, 45% were classified as African-American, 18% common, 12% Caucasian, 9% Amerindian and 16% could not be classified. In the EBMT cohort, from 116 HLA haplotypes, 70% were classified as African-American, 8% common, 6% Caucasian, 3% Amerindian and 13% were not classified. Although Brazilians showed greater genetic admixture, chances of finding at least one potential allelic MUD were 47% in both groups (p-value not significant) and chances of having at least 5 potential allelic MUD were 24% for Brazilians and 15% for EBMT (p-value not significant). Overall, most potential allelic MUD were found in the NMDP registry, followed by the Brazilian registry (REDOME) and by the German registry (ZKMD); for the Brazilian cohort, most potential allelic MUD were found in REDOME. Discussion: Migration from Africa to Brazil started at the colonial period, and interethnic admixture have been occurring since then, explaining the higher diversity observed in the Brazilian cohort. Despite differences in ethnic composition, chances of having at least one potential allelic MUD are identical, probably because carrying at least one African or Amerindian haplotype decreases the chances of a full HLA matching. Although we demonstrated higher probabilities of finding a potential allelic MUD in SCD than previous studies, the chances are still low, therefore further strategies are required to increase donor representativeness for SCD. In this setting, alternative sources, such as haploidentical HSCT and cord blood, should be considered. Also, our study might help to predict the probabilities of finding a MUD for patients with SCD. This is important because given that HSCT in SCD has better results if performed at earlier age, knowing which patients are less likely to find a MUD might influence therapy management. Disclosures No relevant conflicts of interest to declare.

Hematopoietic stem cell transplant (HSCT) from an HLA identical sibling is a well-established curative therapy for sickle cell disease (SCD). HSCT from an unrelated donor is a treatment option, but the likelihood of finding a donor varies according to ethnicity and results are still limited. HLA haploidentical relatives can be alternatively used but, to date, only small series of patients have been described. We report outcomes of patients (pts) transplanted with related haploidentical (Haplo) or unrelated (UD) donors grafts and reported to EBMT/EUROCORD databases. Sixty four pts transplanted in 22 EBMT centers between 1991 and 2017 were retrospectively analyzed. Pts were described according to the donor type: haploidentical (n=40) and unrelated (n=24) [adult UD n=19; cord blood (CB) n=5]. The objective of the study was to describe alternative donor transplants for SCD in Europe without performing comparison analyses due to the size and heterogeneity of the groups. Primary endpoint was 3-year overall survival (OS). Median follow-up (FU) was 28 months (range: 1.6-156) [29.5 months (range: 2.1 - 133.5) for Haplo and 24.6 (range: 1.6 - 156) for UD]. Median age at HSCT was 14.2 years (range: 3-31.7) in Haplo and 11.8 (range: 2.1-42.8) in UD, with a predominance of children ( In both groups, vaso-occlusive crisis and cerebral vasculopathy were the most frequent SCD complications and the main indications for HSCT. Other complications were acute chest syndrome (44%), liver disease (31%) and infection (23%). In Haplo, median year of transplant was 2014 (range: 1991-2017) and in UD 2011 (range: 2004-2015). In Haplo, two major protocols were used: (1) post -transplant cyclophosphamide (PTCY) with G-CSF primed bone marrow (BM) and a fludarabine+ cyclophosphamide+thiotepa+2Gy TBI conditioning regimen [16 pts and 2 centers performing most (n=13) of the transplants]; (2) a protocol (performed in 2 centers) consisting in the use of G-CSF mobilized peripheral blood stem cells (PBSC) with ex-vivo B and T cell depletion (BT depleted) (15 pts) and a fludarabine+thiotepa+ treosulfan conditioning regimen (14/15 pts). Haplo donors were most frequently the parents [mother (50%), father (29%), brother (14%) and cousin (7%)]. ATG was used in 95% of transplants and the most frequent combination for graft versus host disease (GvHD) prophylaxis was mycophenolate mofetil (MMF)+sirolimus in PTCY and MMF+ cyclosporine A (CSA) in BT depleted. In UD, graft source distribution was 14 BM, 5 PBSC and 5 CB. Conditioning regimens were mainly myeloablative (83%) with fludarabine+thiotepa+ treosulfan in 54% of HSCT. ATG was used in 87% and campath in 9% of transplants; GvHD prophylaxis was CSA and methotrexate in 50%. Neutrophil engraftment at 60 days was 95±4% in Haplo and 84±8% in adult UD, after a median engraftment time of 18 and 22 days, respectively. In Haplo, 7 pts experienced graft failure (3 primary and 4 late), of those 3 had a second allogeneic transplant and were alive at last FU, at 16, 16 and 63 months respectively; 1 patient died after rescue with autologous transplant and 3 were alive after autologous reconstitution. In adult UD, 3 pts had a primary and 1 a late graft failure, none of them had a second transplant and were all alive at last FU, at 2, 13, 28, 118 months respectively. Grade II-IV acute GvHD at 100 days was 25±7% in Haplo and 21±9% in adult UD; acute GvHD grade III-IV was observed in 3 pts in Haplo (none in BT depleted) and 2 pts in adult UD. Chronic GvHD was observed in 10 pts in Haplo (5 extensive, 3 of these in PTCY) and 3 pts in adult UD (2 extensive). OS at 3 years was 88±4%; being 89±5% in Haplo (88±8% for PTCY, 92±8% for BT depleted) and 94±5% in adult UD. 3-year event free survival was 58±7%; in detail, 60±9% in Haplo (56±12% for PTCY, 68±13% for BT depleted) and 60±12% in adult UD. Overall, 8 pts died (5 Haplo and 3 UD) due to infections or GVHD. Among the 5 pts receiving CB transplant 3 are alive (1 of which after graft failure and a second allogeneic transplant). Conclusion: This preliminary analysis shows that, despite an acceptable OS, rejection and chronic GvHD are still of concern; therefore alternative donor transplants for SCD should be performed in experienced centers with prospective clinical trials. Disclosures Pondarré: Blue Bird Bio: Honoraria; Novartis: Honoraria; Addmedica: Membership on an entity's Board of Directors or advisory committees. Zecca:Chimerix: Honoraria. Locatelli:Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Miltenyi: Honoraria; Bellicum: Consultancy, Membership on an entity's Board of Directors or advisory committees; bluebird bio: Consultancy. Bader:Medac: Patents & Royalties, Research Funding; Riemser: Research Funding; Neovii: Research Funding; Cellgene: Consultancy; Novartis: Consultancy, Speakers Bureau. Bernaudin:AddMedica: Honoraria; Pierre fabre: Research Funding; BlueBirdBio: Consultancy; Cordons de Vie: Research Funding.

Background: Sickle cell disease (SCD) patients (pts) are at increased risk of infection. This susceptibility is partially explained by the loss of spleen function, but other factors such as inflammatory status might also play a role. Despite adequate immunization and antimicrobial prophylaxis, bacterial infection remains a major cause of morbidity and mortality in SCD pts. Toll-like receptors (TLR), transmembrane proteins that recognize a wide range of pathogen molecular patterns, are key receptors in innate immune response. The association among TLR and infections in SCD has scarcely been explored. Objective: To analyze if functional polymorphisms in TLR genes can modulate the susceptibility to bacterial infections in SCD pts. Methods: Case-control retrospective study; 430 SCD pts followed in university hospitals in Brazil (n=196), France (n=180, mainly from Subsaharan Africa and French West Indies) and Senegal (n=54) were included. Sites of bacterial infection were lower respiratory system, abdominal, upper urinary system, bone/joints, central nervous system, blood stream, or any site for tuberculosis. For pts who underwent hematopoietic stem cell transplant (HSCT), only bacterial infections before HSCT were considered. All pts were tested for 7 single nucleotide polymorphisms (SNP) in TLR-1(rs4833095), TLR-2 (rs4696480, rs3804099, rs3804100), TLR-6 (rs5743810) and TLR-10 (rs11466653, rs11096957) using TaqMan 5'-nuclease assay. Univariate analysis was performed with chi-square or Mann-Whitney for categorical and continuous variables respectively. Associations were measured by odds ratio (OR). For SNP associations, significant P-value corrected by Bonferroni (cP) was 0.007; for genetic models, cP was 0.01. Results: Median age of cohort was 30 years (range: 2-70) and 87% were ≥18 years; 57% of pts were female. SCD genotype was SS in 81%, SC in 11%, SB in 7% and other in 1%. Fifty-eight percent received hydroxyurea and 79% received red blood cell transfusions; seven pts underwent HSCT. Eleven pts died during follow-up, mostly from acute chest syndrome and hemorrhagic stroke. All pts had access to immunization against encapsulated bacteria and to penicillin prophylaxis. One hundred ninety-five pts (45%) did not present any bacterial infection, whereas 235 (55%) pts presented at least one episode of bacterial infection, of which 20% had ≥ 3 episodes. Bacterial infections were mainly respiratory (51%), bone/joints (23%) and urinary (20%). In 106 cases, etiological agents were identified; most common agents were Escherichia coli, Streptococcuspneumoniae, Mycobacterium tuberculosis and Salmonella spp. In univariate analysis, there was no association among occurrence of infections and gender, origin and age distribution. Gene analysis: all tested SNPs had a typing rate >90% and minimum allele frequency >1%. The TLR-2 SNP rs4696480 (n=425) showed significant association with infection (p Discussion: In SCD pts, TLR-2 rs4696480 TA genotype might be protective against bacterial infections, whereas TT genotype might increase risk of such infections. Previous reports demonstrated higher secretion of inflammatory factors in cells from AA individuals, lower occurrence and severity of immune diseases in T carriers. TA genotype might stand between deleterious effects of over inflammatory response (AA genotype) and under response (TT genotype) to infectious agents. Disclosures No relevant conflicts of interest to declare.

In the last decade, gene therapy for adenosine deaminase deficiency has been developed as a successful alternative strategy to allogeneic bone marrow transplant and enzyme replacement therapy. Infusion of autologous hematopoietic stem cells, corrected ex vivo by retroviral vectors and combined to low-intensity conditioning regimen, has resulted in immunologic improvement, metabolic correction, and long-term clinical benefits. These findings have opened the way to applications of gene therapy in other primary immune deficiencies using novel vector technology.

Reactivation of HBV is a well known complication in patients undergoing HSCT. Lamivudine treatment appears to prevent hepatitis B virus reactivation and to decrease the mortality in at risk HSCT patients. We describe HBV reactivation occurred in three allogeneic HSCT pediatric patients coming from Eastern Europe, one of whom was successfully treated with lamivudine. Our experience confirms that HBV-DNA may persist as intra-hepatic infection or in extra-hepatic sites and that HBV reactivation may appear during immunodepression. Careful and complete screening for HBV markers is mandatory before HSCT, especially in children coming from countries at risk for HBV. Furthermore, a treatment with lamivudine could also represent an efficacious prophylaxis in pediatric patients to avoid HBV reactivation and to decrease the development of severe hepatic disease.

CEP is a rare inborn error of porphyrin-heme synthesis. Clinical manifestations can range from mild to severe and include erythrodontia, reddish-colored urine, and hemolytic anemia that can be mild or severe and may result in splenomegaly. Completely avoiding exposure to the sun is crucial. Attempts to reduce erythropoiesis and to lower circulating porphyrin levels by means of erythrocyte transfusions have been successful in reducing the expression of the disease. However, the complications of a chronic transfusion regimen are potentially severe. Successful bone marrow transplantation has been reported in CEP. We report a case of successful bone marrow transplantation and prolonged follow-up in an adolescent CEP patient.

We describe a case of Cryptosporidium infection occurring in a child after allogeneic SCT for acute non-lymphoblastic leukemia. This patient presented an intestinal, biliar, and pancreatic Cryptosporidium disease associated with an intestinal aGvHD. The increase in CD3+/CD4+ cells secondary to the reduction of steroid therapy associated with the improvement of aGvHD and the use of antiparasitic treatments (especially nitazoxanide) improved the infection-related symptoms and led to a complete clearance of the Cryptosporidium.

Allogeneic haematopoietic stem cell transplantation (allo-HSCT) is the only cure for juvenile myelomonocytic leukaemia (JMML), but relapse remains the major cause of failure. A second transplant may be considered a way to induce the graft vs. leukaemia effect in patients who relapse after their first HSCT. We describe a 7-month-old girl with JMML who relapsed after a first, related allo-HSCT, and who again relapsed 8 months after the second transplant, despite discontinuation of immusuppressive therapy. She underwent a third allogeneic transplant from another related donor. At the time of this report the patient is in complete remission 26 months after the third transplant. We suggest that a third allo-HSCT may be taken into consideration for JMML patients who experience relapse, even after two previous transplants.

Cytomegalovirus (CMV) is a well-known cause of disease occurring after bone marrow transplantation (BMT). The manifestations of CMV range from asymptomatic infection, defined as active CMV replication in the blood in the absence of clinical manifestations or organ failure abnormalities, to CMV disease, characterized by CMV infection with clinical symptoms or organ function abnormalities. Diagnostic procedures to assess the viral load have improved greatly with the increased use of antigenemia, CMV DNA, and immediate early-messenger RNA. Many conditions concur in determining the risk of developing CMV reactivation or disease after bone marrow transplant with serologic status of donor and recipient, type of bone marrow transplant, presence of graft-versus-host disease being the most studied. However, time and quality of immune reconstitution seems to be the pivotal factors. Pneumonia and gastrointestinal involvement are the most frequently documented clinical pictures with late-onset CMV reactivation or disease representing a new challenge. CMV prophylaxis or pre-emptive therapy adopted during the last few years in allogeneic BMT recipients has changed the natural history of the disease, reducing the risk of CMV disease, CMV-associated death, transplant-related mortality, and has prolonged the period at risk. Specific studies on children are lacking, however, the clinical pictures and features seems to be similar both in children and adults.

Mesial temporal sclerosis (MTS) is a common finding in patients with intractable temporal lobe epilepsy (TLE). In this report, we retrospectively reviewed the neuroimaging results of four children who underwent allogeneic hematopoietic stem cell transplantation (allo-HSCT), and who developed recurrent, partial, intractable seizures following a first event caused by cyclosporine-A (CSA) neurotoxicity. Neuroradiologic findings of MTS were demonstrated in all these patients. We suggest that MTS may be a consequence of CSA neurotoxicity, which induces repeated seizures, associated with other predisposing conditions, as well as being a consequence of the underlying disease and its treatment, and of severe graft-versus-host disease (GvHD).

To investigate the relationship between clinical response and modification of BK viremia, we assessed retrospectively 32 cases of hemorrhagic cystitis (HC) after allogeneic hematopoietic SCT that were treated with i.v. cidofovir (CDV). They were 22 men (69%) and 10 women (31%) with a median age of 24 years, range 3-62. The median number of CDV doses was 3, range 1-8, and the treatment lasted for a median of 3 weeks, range 1-10. Clinical improvement of HC was observed in 27 patients (84%). In 12 of 32 episodes (37.5%), BK viremia was determined before every CDV administration and a complete clinical response was observed in 10 of 12 patients (83%), the reduction of BK viremia load being >= 1 log by 2 weeks after starting CDV. Nephrotoxicity related to CDV was observed in nine patients. Among 26 patients with 100-day follow-up, 4 of 4 patients who had a complete clinical response by 30 days were alive vs 16 of 22 (73%) who did not have the resolution of HC in this time frame. We conclude that in patients with HC, the response to CDV treatment is usually associated with a significant reduction of BK viremia load.

beta-Thalassemia is a severe inherited anemia caused by insufficient production of beta-globin chains. Allogeneic hematopoietic stem cell (HSC) transplantation is currently the only cure, and is limited by donor availability and regimen-related toxicity and mortality. Gene therapy is a promising therapeutic tool for all thalassemic patients lacking a compatible donor and potentially provides transfusion independence in the absence of transplant-related complications, such as graft rejection and graft-versus-host disease. The issue of HSC procurement is critical in this setting because of the specific features of thalassemic syndromes, which include bone marrow (BM) expansion, ineffective erythropoiesis, and splenomegaly. Little is known about the efficiency of CD34(+) cell yield from steady-state BM harvests from thalassemic patients. We have collected data on safety and cell yield from 20 pediatric patients with beta-thalassemia who underwent autologous BM harvest before allogeneic HSC transplantation, and from 49 age-matched sibling donors who also underwent BM harvest. The procedure was safe, as no significant adverse events occurred. In terms of cell yield, no difference was found between patients and normal donors in the number of CD34(+) cells and total nucleated cells harvested. Most importantly, no difference was found in the proportion of myeloid and erythroid progenitors, suggesting a similar repopulating capacity. On the basis of these results, we conclude that steady-state BM can be used as a safe and efficient source of HSC for gene therapy of b-thalassemia. "beta-Thalassemia is a severe inherited anemia caused by insufficient production of beta-globin chains. Allogeneic hematopoietic stem cell (HSC) transplantation is currently the only cure, and is limited by donor availability and regimen-related toxicity and mortality. Gene therapy is a promising therapeutic tool for all thalassemic patients lacking a compatible donor and potentially provides transfusion independence in the absence of transplant-related complications, such as graft rejection and graft-versus-host disease. The issue of HSC procurement is critical in this setting because of the specific features of thalassemic syndromes, which include bone marrow (BM) expansion, ineffective erythropoiesis, and splenomegaly. Little is known about the efficiency of CD34(+) cell yield from steady-state BM harvests from thalassemic patients. We have collected data on safety and cell yield from 20 pediatric patients with beta-thalassemia who underwent autologous BM harvest before allogeneic HSC transplantation, and from 49 age-matched sibling donors who also underwent BM harvest. The procedure was safe, as no significant adverse events occurred. In terms of cell yield, no difference was found between patients and normal donors in the number of CD34(+) cells and total nucleated cells harvested. Most importantly, no difference was found in the proportion of myeloid and erythroid progenitors, suggesting a similar repopulating capacity. On the basis of these results, we conclude that steady-state BM can be used as a safe and efficient source of HSC for gene therapy of b-thalassemia."

Background: Gene therapy (GT) with hematopoietic stem cells is a promising treatment for inherited immunodeficiencies. Objectives: Limited information is available on the relative contribution of de novo thymopoiesis and peripheral expansion to T-cell reconstitution after GT as well as on the potential effects of gene transfer on hematopoietic stem cells and lymphocyte replicative lifespan. We studied these issues in patients affected by adenosine deaminase severe combined immune deficiency after low-intensity conditioning and reinfusion of retrovirally transduced autologous CD34(+) cells. Methods: Immunophenotype, proliferative status, telomere length, and T-cell receptor excision circles were investigated at early and late time points (up to 9 years) after GT treatment. Control groups consisted of pediatric healthy donors and patients undergoing allogeneic bone marrow transplantation (BMT). Results: We observed no telomere shortening in the bone marrow compartment and in granulocytes, whereas peripheral blood naive T cells from both GT and BMT patients showed a significant reduction in telomere length compared with healthy controls. This was in agreement with the presence of a high fraction of actively cycling naive and memory T cells and lower T-cell receptor excision circles. Conclusion: These data indicate that T-cell homeostatic expansion contributes substantially to immune reconstitution, like BMT, and is not associated with senescence in the stem cell compartment. (J Allergy Clin Immunol 2011;127:1368-75.) BACKGROUND:Gene therapy (GT) with hematopoietic stem cells is a promising treatment for inherited immunodeficiencies.OBJECTIVES:Limited information is available on the relative contribution of de novo thymopoiesis and peripheral expansion to T-cell reconstitution after GT as well as on the potential effects of gene transfer on hematopoietic stem cells and lymphocyte replicative lifespan. We studied these issues in patients affected by adenosine deaminase severe combined immune deficiency after low-intensity conditioning and reinfusion of retrovirally transduced autologous CD34(+) cells.METHODS:Immunophenotype, proliferative status, telomere length, and T-cell receptor excision circles were investigated at early and late time points (up to 9 years) after GT treatment. Control groups consisted of pediatric healthy donors and patients undergoing allogeneic bone marrow transplantation (BMT).RESULTS:We observed no telomere shortening in the bone marrow compartment and in granulocytes, whereas peripheral blood naive T cells from both GT and BMT patients showed a significant reduction in telomere length compared with healthy controls. This was in agreement with the presence of a high fraction of actively cycling naive and memory T cells and lower T-cell receptor excision circles.CONCLUSION:These data indicate that T-cell homeostatic expansion contributes substantially to immune reconstitution, like BMT, and is not associated with senescence in the stem cell compartment.

Recently vancomycin-resistant and sporadically linezolid-resistant Enterococcus species have been described in adults. We report what we believe to be the first case of a child with prolonged bone marrow aplasia following haematopoietic stem cell transplantation developing a fatal sepsis caused by Enterococcus faecium resistant to glycopeptides and linezolid. Recently vancomycin-resistant and sporadically linezolid-resistant Enterococcus species have been described in adults. We report what we believe to be the first case of a child with prolonged bone marrow aplasia following haematopoietic stem cell transplantation developing a fatal sepsis caused by Enterococcus faecium resistant to glycopeptides and linezolid.

Immune-mediated refractoriness to platelet transfusion is a major problem in patients undergoing HSCT. In a cohort of 50 pediatric patients affected by beta thalassemia coming from Middle East countries, we experienced a high incidence of refractoriness because of anti-HLA antibodies during post-HSCT aplasia. In a risk factors analysis, factors predicting a negative transfusion outcome were presence of spleen and the number of anti-HLA antibodies. We adopted a policy to select platelet donors by avoiding HLA antigens against which the patient had specific antibodies. Transfusion of dedicated units resulted in 26% refractoriness compared to 74% to random units (p < 0.0001). When dedicated transfusions were used, the presence of spleen did not influence transfusion outcome. Analyzing transfusion outcome depending on the degree of HLA match and ABO compatibility, 76% successful transfusions were obtained with HLA-matched- ABO compatible followed by 67% in HLA-1mismatch- ABO compatible or HLA-matched- ABO incompatible and by 46% in HLA-1mismatch- ABO incompatible. In conclusion, we provide evidence that the selection of platelet donors according to patient characteristics, anti-HLA antibodies and ABO matching, is successful in reducing platelet refractoriness in heavily alloimmunized thalassemia patients undergoing transplantation. Immune-mediated refractoriness to platelet transfusion is a major problem in patients undergoing HSCT. In a cohort of 50 pediatric patients affected by beta thalassemia coming from Middle East countries, we experienced a high incidence of refractoriness because of anti-HLA antibodies during post-HSCT aplasia. In a risk factors analysis, factors predicting a negative transfusion outcome were presence of spleen and the number of anti-HLA antibodies. We adopted a policy to select platelet donors by avoiding HLA antigens against which the patient had specific antibodies. Transfusion of dedicated units resulted in 26% refractoriness compared to 74% to random units (p < 0.0001). When dedicated transfusions were used, the presence of spleen did not influence transfusion outcome. Analyzing transfusion outcome depending on the degree of HLA match and ABO compatibility, 76% successful transfusions were obtained with HLA-matched- ABO compatible followed by 67% in HLA-1mismatch- ABO compatible or HLA-matched- ABO incompatible and by 46% in HLA-1mismatch- ABO incompatible. In conclusion, we provide evidence that the selection of platelet donors according to patient characteristics, anti-HLA antibodies and ABO matching, is successful in reducing platelet refractoriness in heavily alloimmunized thalassemia patients undergoing transplantation.

Pseudomonas aeruginosa is one leading gram-negative organism associated with nosocomial infections. Bacteremia is life-threatening in the immunocompromised host. Increasing frequency of multi-drug-resistant (MDRPA) strains is concerning. We started a retrospective survey in the pediatric hematology oncology Italian network. Between 2000 and 2008, 127 patients with Pseudomonas aeruginosa bacteremia were reported from 12 centers; 31.4% of isolates were MDRPA. Death within 30 days of a positive blood culture occurred in 19.6% (25/127) of total patients; in patients with MDRPA infection it occurred in 35.8% (14/39). In the multivariate analysis, only MDRPA had significant association with infection-related death. This is the largest series of Pseudomonas aeruginosa bacteremia cases from pediatric hematology oncology centers. Monitoring local bacterial isolates epidemiology is mandatory and will allow empiric antibiotic therapy to be tailored to reduce fatalities.

Background: Neurotoxicity is a recognized complication of cyclosporine A (CSA) treatment. The incidence of severe CSA-related neurological complications following hematopoietic stem cell transplantation (HSCT) is 4-11%. Methods: We describe 6 cases of CSA related neurotoxicity out of 67 matched related HSCT performed in paediatric Middle East patients affected by haemoglobinopaties (5 beta thalassemia major, 1 sickle cell disease-SCD). Conditioning regimen consisted of iv busulphan, cyclophosphamide and graft-versus-host-disease (GvHD) prophylaxis with CSA, methylprednisolone, methotrexate and ATG. Results: All 6 patients presented prodromes such as arterial hypertension, headache, visual disturbances and vomiting, one to two days before overt CSA neurotoxicity. CSA neurotoxicity consisted of generalized seizures, signs of endocranial hypertension and visual disturbances at a median day of onset of 11 days after HSCT (range +1 to +40). Brain magnetic resonance imaging (MRI) performed in all subjects showed reversible leukoencephalopathy predominantly in the posterior regions of the brain (PRES) in 5/6 patients. EEG performed in 5/6 patients was always abnormal. Neurotoxicity was not explainable by high CSA blood levels, as all patients had CSA in the therapeutic range with a median of 178 ng/ml (range 69-250). CSA was promptly stopped and switched to tacrolimus with disappearance of clinical and radiological findings. All patients are symptoms-free at a median follow up of 882 days (range 60-1065). Conclusions: Our experience suggests that paediatric patients with haemoglobinopaties have a high incidence of CSA related neurological events with no correlation between serum CSA levels and neurotoxicity. Prognosis is good following CSA removal. Specific prodromes such as arterial hypertension, headache or visual disturbances occurring in the early post-transplant period should be carefully evaluated with electrophysiological and MRI-based imaging in order to intervene promptly and avoid irreversible sequels. Background: Neurotoxicity is a recognized complication of cyclosporine A (CSA) treatment. The incidence of severe CSA-related neurological complications following hematopoietic stem cell transplantation (HSCT) is 4-11%. Methods: We describe 6 cases of CSA related neurotoxicity out of 67 matched related HSCT performed in paediatric Middle East patients affected by haemoglobinopaties (5 beta thalassemia major, 1 sickle cell disease-SCD). Conditioning regimen consisted of iv busulphan, cyclophosphamide and graft-versus-host-disease (GvHD) prophylaxis with CSA, methylprednisolone, methotrexate and ATG. Results: All 6 patients presented prodromes such as arterial hypertension, headache, visual disturbances and vomiting, one to two days before overt CSA neurotoxicity. CSA neurotoxicity consisted of generalized seizures, signs of endocranial hypertension and visual disturbances at a median day of onset of 11 days after HSCT (range +1 to +40). Brain magnetic resonance imaging (MRI) performed in all subjects showed reversible leukoencephalopathy predominantly in the posterior regions of the brain (PRES) in 5/6 patients. EEG performed in 5/6 patients was always abnormal. Neurotoxicity was not explainable by high CSA blood levels, as all patients had CSA in the therapeutic range with a median of 178 ng/ml (range 69-250). CSA was promptly stopped and switched to tacrolimus with disappearance of clinical and radiological findings. All patients are symptoms-free at a median follow up of 882 days (range 60-1065). Conclusions: Our experience suggests that paediatric patients with haemoglobinopaties have a high incidence of CSA related neurological events with no correlation between serum CSA levels and neurotoxicity. Prognosis is good following CSA removal. Specific prodromes such as arterial hypertension, headache or visual disturbances occurring in the early post-transplant period should be carefully evaluated with electrophysiological and MRI-based imaging in order to intervene promptly and avoid irreversible sequels.

beta-thalassemia is a major health problem worldwide, and stem cell transplantation (SCT) is the only curative option. Oral Busulfan (Bu) based conditioning is widely used in this setting. Due to the variability of Bu systemic exposure, intravenous (i.v.) Bu has been proposed as a standard of care, with no need for drug monitoring and dose adjustment. Patients with beta-thalassemia from countries with limited resources might be at higher risk of erratic Bu metabolism because of liver dysfunction, severe iron overload, and specific ethnic/genetic features. We studied Bu pharmacokinetics in 53 children with advanced beta-thalassemia from Middle Eastern countries who underwent a total of 57 matched related donor SCTs. Forty-two percent of the children required dose adjustment because they did not achieve the therapeutic window after the first dose. With a Bu dose-adjustment policy, regimen-related toxicity was limited. At a median follow-up of 564 days, the probabilities of 2-year survival, current thalassemia-free survival, rejection, and treatment-related mortality were 96%, 88%, 21%, and 4%, respectively. Conditioning with i.v. Bu and dose adjustment is feasible and well tolerated, although recurrence of thalassemia remains an unsolved problem in children with advanced disease. beta-thalassemia is a major health problem worldwide, and stem cell transplantation (SCT) is the only curative option. Oral Busulfan (Bu) based conditioning is widely used in this setting. Due to the variability of Bu systemic exposure, intravenous (i.v.) Bu has been proposed as a standard of care, with no need for drug monitoring and dose adjustment. Patients with beta-thalassemia from countries with limited resources might be at higher risk of erratic Bu metabolism because of liver dysfunction, severe iron overload, and specific ethnic/genetic features. We studied Bu pharmacokinetics in 53 children with advanced beta-thalassemia from Middle Eastern countries who underwent a total of 57 matched related donor SCTs. Forty-two percent of the children required dose adjustment because they did not achieve the therapeutic window after the first dose. With a Bu dose-adjustment policy, regimen-related toxicity was limited. At a median follow-up of 564 days, the probabilities of 2-year survival, current thalassemia-free survival, rejection, and treatment-related mortality were 96%, 88%, 21%, and 4%, respectively. Conditioning with i.v. Bu and dose adjustment is feasible and well tolerated, although recurrence of thalassemia remains an unsolved problem in children with advanced disease. Abstract beta-thalassemia is a major health problem worldwide, and stem cell transplantation (SCT) is the only curative option. Oral Busulfan (Bu) based conditioning is widely used in this setting. Due to the variability of Bu systemic exposure, intravenous (i.v.) Bu has been proposed as a standard of care, with no need for drug monitoring and dose adjustment. Patients with beta-thalassemia from countries with limited resources might be at higher risk of erratic Bu metabolism because of liver dysfunction, severe iron overload, and specific ethnic/genetic features. We studied Bu pharmacokinetics in 53 children with advanced beta-thalassemia from Middle Eastern countries who underwent a total of 57 matched related donor SCTs. Forty-two percent of the children required dose adjustment because they did not achieve the therapeutic window after the first dose. With a Bu dose-adjustment policy, regimen-related toxicity was limited. At a median follow-up of 564 days, the probabilities of 2-year survival, current thalassemia-free survival, rejection, and treatment-related mortality were 96%, 88%, 21%, and 4%, respectively. Conditioning with i.v. Bu and dose adjustment is feasible and well tolerated, although recurrence of thalassemia remains an unsolved problem in children with advanced disease.

Gene therapy is a highly attractive strategy for many types of inherited disorders of the immune system. Adenosine deaminase (ADA) deficient-severe combined immunodeficiency (SCID) has been the target of several clinical trials based on the use of hematopoietic stem/progenitor cells engineered with retroviral vectors. The introduction of a low intensity conditioning regimen has been a crucial factor in achieving stable engrafment of hematopoietic stem cells and therapeutic levels of ADA-expressing cells. Recent studies have demonstrated that gene therapy for ADA-SCID has favorable safety profile and is effective in restoring normal purine metabolism and immune functions. Stem cell gene therapy combined with appropriate conditioning regimens might be extended to other genetic disorders of the hematopoietic system.

Allogeneic BMT represents the only chance of cure for beta-thalassemia. Occasionally, two affected individuals from the same family share a matched healthy sibling. Moreover, a high incidence of transplant rejection is still observed in Pesaro class III patients, requiring a second BMT procedure. In these settings, one option is to perform a second BM harvest from the same donor. Although BM harvest is a safe procedure in children, ethical issues concerning this invasive practice still arise. Here, we describe our series of seven pediatric, healthy donors, who donated BM more than once in favor of their beta-thalassemic HLA-identical siblings between June 2005 and January 2008. Three donors donated BM twice to two affected siblings and four donors donated twice for the same sibling following graft rejection of the first BMT. All donors tolerated the procedures well and no relevant side effects occurred. There was no significant difference between the two harvests concerning cell yield and time to engraftment. Our experience shows that for pediatric donors, a second BM donation is safe and feasible and good cellularity can be obtained. We suggest that a second harvest of a pediatric donor can be performed when a strong indication for BMT exists. Allogeneic BMT represents the only chance of cure for beta-thalassemia. Occasionally, two affected individuals from the same family share a matched healthy sibling. Moreover, a high incidence of transplant rejection is still observed in Pesaro class III patients, requiring a second BMT procedure. In these settings, one option is to perform a second BM harvest from the same donor. Although BM harvest is a safe procedure in children, ethical issues concerning this invasive practice still arise. Here, we describe our series of seven pediatric, healthy donors, who donated BM more than once in favor of their beta-thalassemic HLA-identical siblings between June 2005 and January 2008. Three donors donated BM twice to two affected siblings and four donors donated twice for the same sibling following graft rejection of the first BMT. All donors tolerated the procedures well and no relevant side effects occurred. There was no significant difference between the two harvests concerning cell yield and time to engraftment. Our experience shows that for pediatric donors, a second BM donation is safe and feasible and good cellularity can be obtained. We suggest that a second harvest of a pediatric donor can be performed when a strong indication for BMT exists.

Caspofungin is an antifungal drug active as a rescue treatment for invasive aspergillosis in cancer patients (4), with a MIC level of 0.5 mg/liter (2) and a minimum target concentration of 1 mg/liter (5, 6). In adults, these levels are achieved after a loading dose of 70 mg on the first day, followed by 50 mg once a day in the following days (5), while in children, a dose of 50 mg/m2 without a loading dose is required (6). We observed an 8-year-old girl with a previous pulmonary aspergillosis following allogeneic hemopoietic stem cell transplant for Pearson's syndrome that relapsed concomitant with the development of posttransplant acute myeloblastic leukemia (1). The infection relapsed during liposomal amphotericin B secondary prophylaxis; voriconazole treatment was not feasible because of concomitant barbiturate therapy (3), and therefore, caspofungin at 50 mg/m2 (5) (i.e., 30 mg/day) was administered. The patient was in poor clinical and psychological condition and refused any hospital stay; therefore, after 8 days of in-hospital treatment, the patient was discharged for the weekend. No antifungal treatment was administered for two days, and then the patient was admitted again to restart therapy. Blood samples for the determination of caspofungin blood concentrations were taken on the last day of treatment (Friday) and the first day of restarting treatment (Monday), before (through) and 30 min after the end (peak) of caspofungin infusion. The samples were stored at −80°C and sent to the Department of Pharmacology of the University of Florence for the determination of caspofungin blood levels. Caspofungin concentrations were determined by high-pressure liquid chromatography (lowest sensitivity limit, 0.125 mg/liter). A Chromsep C8 4.6-by-250-mm 5-μm column preceded by an adequate precolumn was used for the analysis; a flow rate of 1.3 ml/min was used. The fluorescence detector excitation and emission wavelengths were set at 220 and 304 nm, respectively. The drug was isolated from the plasma by solid-phase extraction (5). The caspofungin levels that resulted were 6.7 mg/liter through and 11.5 mg/liter peak on the last day of treatment and 1.9 mg/liter through and 5.6 mg/liter peak on the first day of resumption of therapy. Even after a 48-h interruption, the caspofungin blood concentration was above the required target concentration for the treatment of invasive aspergillosis (2, 5, 6). Even if further studies are needed to confirm this observation, our experience suggests that the administration of caspofungin for 5 days/week, after an adequate period (at least 1 week) of daily therapy, may be a possible approach for children with invasive aspergillosis who cannot receive other antifungal drugs and may not be hospitalized for some reason.