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Aim: The aim of this study was to describe the characteristics of Posterior Reversible Encephalopathy Syndrome (PRES) in infants and young children (

Background: Pediatric myelodysplastic syndromes (MDSs) are a heterogeneous group of clonal disorders, accounting for less than 5% of childhood hematologic malignancies. Usual indications to HSCT are: MDSs with excess of blasts, MDSs secondary to previously administered chemoradiotherapy and RCC associated with monosomy 7, complex karyotype, severe neutropenia, or erythrocyte/platelet transfusion dependence [Locatelli & Strahm, Blood 2018]. We previously demonstrated that TBdepl-haploHSCT is a suitable option for children with acute leukemia, with outcomes comparable to those reported in studies using either an HLA-identical sibling or an unrelated volunteer as donor. Here we present the results of this approach in children with MDSs. Patients and methods: Between February 2013 and February 2021, 23 children with MDSs other than juvenile myelomonocytic leukemia received TBdepl-haploHSCT from an HLA-partially matched relative at Ospedale Pediatrico Bambino Gesù, Rome, Italy or at IRCCS Fondazione Policlinico San Matteo, Pavia, Italy as part of a prospective study (#NCT01810120). All patients were prepared to the allograft using a fully-myeloablative conditioning regimen including a combination of cytotoxic drugs and/or total body irradiation (TBI). Anti-T-lymphocyte globulin (ATLG) was used before transplantation (12 mg/kg total dose, from days -5 to day -3) to modulate bi-directional donor/recipient alloreactivity. Rituximab (200 mg/sqm) was administered on day -1 to prevent post-transplantation EBV-induced lymphoproliferative disorders (PTLD). No patient received any post-transplant pharmacological GvHD prophylaxis. Results: Characteristics of patients enrolled in the study are shown in Table 1 (which reports also donor and graft characteristics). Median follow-up of surviving patients is 4.2 years (range: 0.5 - 8.5 years). Seventeen children were affected by refractory cytopenia of childhood (RCC) (2 cases occurring in the context of inherited bone marrow failure syndromes: one had GATA2 deficiency and the other SAMD9L mutation), while 1 and 5 were affected by MDS with excess of blasts 1 (EB1) and EB2 (one had GATA2 deficiency), respectively. Median time to neutrophil and platelet recovery was 14 (range 10-19) and 11 (range 9-14) days, respectively, with four patients (3 with RCC and 1 with EB2) experiencing primary graft failure, the cumulative incidence of this complication being 17.3% (95% CI 0.3-31.5). All these 4 patients were rescued with a second TBdepl-haploHSCT from the same or the other parent. Cumulative incidence of grade II-III acute GvHD was 11.4% (95% CI 0-25.2). One patient developed skin and gut GvHD after the second TBdepl-haploHSCT, while for all other patients skin was the sole organ involved; no case of grade IV GvHD was observed. One patient developed moderate chronic GvHD [cumulative incidence 5.2% (95% CI 0-14.8)], which completely resolved with low-dose steroids and ruxolitinib. Notably, no patient died for transplant-related complications. Six patients experienced CMV, 2 HHV-6 and 1 adenoviral infection/reactivation; one patient developed lung aspergillosis, which resolved with specific treatment. One patient affected by EB2, not in remission at time of transplant, relapsed 27 months after HSCT, the 5-year cumulative incidence of relapse being 7.1% (95% CI, 0-19.7); she eventually died after failing a second HSCT. The 5-year probability of overall and event-free survival were 92.3% (95% CI 56.6 -98.9) and 76.3% (95% CI 51.3-89.6) (Figure 1A and B), respectively. Five-year disease-free-survival was 90% (95% CI 47.3-98.5). Because of the low number of events, no prognostic factor related to OS and EFS was found. In particular, the MDS variant did not influence the patient's outcome. The median CD3+ cell count on day +30, +90, +180 and +360 were 113, 171, 558 and 1307/mcl, respectively. Conclusions: These data indicate that TBdepl-haploHSCT is a safe and effective transplant option also in children with MDS. Indeed, the low risk of both non-relapse mortality and acute/chronic GvHD makes this approach particularly attractive in the pediatric setting. Moreover, this haplo strategy compares favorably with T-cell replete approaches [Suo et al., 2020]. Figure 1 Figure 1. Disclosures Merli: JAZZ: Consultancy; SOBI: Consultancy. Locatelli: Miltenyl: Honoraria; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Bellicum: Consultancy, Membership on an entity's Board of Directors or advisory committees; bluebird bio, Inc.: Consultancy; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees.

Diamond-Blackfan anemia (DBA) is a rare congenital disorder presenting remarkable phenotypic overlap with other inherited bone marrow failure syndromes, making differential diagnosis challenging and its confirmation often reached with great delay. By whole exome sequencing, we unraveled the presence of pathogenic variants affecting genes already known to be involved in DBA pathogenesis (RPL5 and RPS19) in three patients with otherwise uncertain clinical diagnosis, and provided new insights on DBA genotype-phenotype correlations. Remarkably, the RPL5 c.482del frameshift mutation has never been reported before, whereas the RPS19 c.3G > T missense mutation, although previously described in a 2-month-old DBA patient without malformations and refractory to steroid therapy, was detected here in the mosaic state in different bodily tissues for the first time in DBA patients.

Acute leukemias (AL) is a major cause of cancer death in young age. Extensive research is being conducted to identify novel and innovative approaches for leukemia treatment. Transcriptomic and epigenetic studies might help to discover potential targets, paving the way for molecular-targeted therapies. We analyzed a cohort of 34 AL at diagnosis: 10 pediatric Acute Lymphoblastic Leukemias (8 B-cell, 1 T-cell and 1 bi-phenotypic ALL), 4 young adult ALL (2 B-cell and 2 T-cell phenotype), and 20 pediatric Acute Myeloid Leukemias (AML: 3 M1, 4 M2, 1 M3, 5 M4, 7 M5) with an average blasts population higher than 80% (85+/-11% in ALL; 83+/-13% in AML). Six out of 14 ALL, and 12 out of 20 AML fell under "high risk" category according to clinical standard risk stratification algorithms. On all patients we performed mRNA sequencing (20-million-reads on Illumina Hiseq 4000). Analyses were performed adjusting The expression of a set of 800 microRNAs (miRNAs) was evaluated by means of Nanostring miRNA panel. Expression signatures and associations among the different risk groups were calculated with t-tests and linear regression analyses. Applying stringent FDR statistical measurement, we discovered 3 genes that significantly differentiate the transcriptomic profile of high vsintermediate/standard-risk ALL in mRNAseq. The expression of PGR3 (p53 Responsive Gene) and long-non-coding RNAs (lncRNAs) ENSG00000228737 and ENSG00000253174 were respectively 45.5, 4.2 and 3.9 time downregulated in high-risk ALL. To explore more deeply the apoptosis pathway in ALL, we measured Tp53 expression and found it significantly downregulated in the high-risk vsintermediate-standard-risk ALL (p= Tp53 dysregulation is a known hallmark for tumor progression; Tp53 mutations - ranging from 1-2% to 10% in pediatric and adult ALs - correlate with worse prognosis. However, in our cohorts, this gene signature was found significant only in high-risk ALL, homogeneously distinguishing them from intermediate/standard-risk ALL. Transcriptome clinical variant analyses excluded pathogenetic known variants that could explain such marked difference. Also, it is unlikely that somatic genetic mutations acquired by the tumor would explain such a homogeneous behavior of high-risk ALL. Thus, we analyzed the p53 regulatory pathway. Interestingly we found that miRNAs known to be involved in p53 control were significantly upregulated in high-risk vs intermediate-standard-risk ALL (p We found PRG3 and Tp53 significantly downregulated in high-risk ALL, with PRG3 expression 45 times lower than intermediate/low-risks ALL. Deeper analyses pointed out to an apoptosis control program not generated by a somatic mutation in the tumor, nor a germline clinical patient variant, but by an epigenetic mechanism. We are currently validating these data in a larger cohort, adding also methylome analyses. It will be interesting also to explore the function of the lncRNAs markedly downregulated in our cohort, whose functions are still unknown or partially known. Because of the small numerosity of the ALL high-risk cohort, we were not able to dissect high-risk young adults (4/6) from pediatric ALL (2/6). Although the homogeneity of data suggests a shared apoptosis control mechanism, it will be worthy to explore in a larger cohort whether the general worse prognosis of young adults/adults vs pediatric ALL is at least partially explained by this mechanism. Disclosures No relevant conflicts of interest to declare.

Introduction Acute Lymphoblastic Leukemia (ALL) is the most common malignancy in children and represents 75-80% of leukemia cases. Despite its recognized role in the initiation of the disease, the bone marrow (BM) microenvironment is not well understood especially in its non-cellular component. High-throughput protein detection represent an opportunity to unveil new biomarkers for leukemia diagnosis, prognosis, monitoring, and to potentially identify biological targets. Methods In the present exploratory study, we evaluated the soluble compartment of BM and peripheral blood (PB) in 16 ALL pediatric patients by the means of the SOMAscan assay, a highly multiplexed, affinity proteomics platform able to measure 1305 proteins in as little as 65µl of sample using modified protein-binding single-stranded DNA aptamers called SOMAmers, revealed on a high dimension oligo-probe array. The raw hybridization data were normalized using hybridization controls and were log2-transformed. Transformed HybNormalized BM and PB plasma data were compared using GraphPad PRISM 8. Multiple comparisons were set with an FDR threshold of 0.01%. Results The comparative analysis between plasma samples from BM and PB showed that 228 proteins are differentially expressed in BM and PB plasma irrespective of the nature of disease (fig1). A more detailed mix-effect analysis of protein expression in the different patient categories and compartment defined in Table 1 revealed that out of these 228 proteins, 92 showed a significant differential expression when performing a mixed-effect analysis between the different groups. Among the top 20 proteins differentially expressed between BM and PB, 18 show differential expression between patient groups. The patient group effect is dominated by the differences between BM and PB plasma in ALL Common patients , which account for 17 of the observed differences in protein levels. In spite of the limited sample size, the analysis revealed a difference in NOTC2 level between ALL Common PB and ALL Pro-B PB. Conclusion In our study we applied a quantitative large-scale proteomic analysis on BM and PB plasma in children suffering from ALL, identifying differential protein expression between the two compartments. In the last few years, SomaScan has emerged as a very attractive method due to its high throughput capacity, faster discovery mode compared to other proteomic platforms and small sample size required, which make it ideal for the identification of novel biomarkers. In spite of the limiting samples sizes, some differences in protein expression were revealed. We are taking this analysis further by analyzing more patients in different groups and including normal donor samples. Table 1 Table Disclosures No relevant conflicts of interest to declare.

We traced the neoplastic history (from 5 to 11 years of age) of a child with concomitant Fanconi anemia and Li-Fraumeni syndrome. Interestingly, the patient developed a highly malignant T-cell non-Hodgkin lymphoma (NHL), which does not represent the typical tumor type in the two aforementioned syndromes, presumably due to the underlying genomic instability. By using a combination of molecular and immunohistochemical approaches, we characterized the accumulation of multiple genetic alterations in a single patient, with both germline (parentally inherited biallelic FANCA variants and a likely de novo nonsense variant in TP53) and somatic (TP53 loss of heterozygosity and 5q interstitial deletion) contributions. Our findings support the interplay of TP53 and FANC genes in DNA damage response pathways and further highlight the genetic heterogeneity of lymphomas as well as the contribution of genomic instability to lymphomagenesis.

Transient myeloproliferative disorder (TMD) is a spontaneously resolving clonal myeloid proliferation characterized by circulating megakaryoblasts in the peripheral blood that is restricted to neonates with Down syndrome (DS) or those with trisomy 21 mosaicism. Cutaneous manifestations of TMD are observed in only 5% of affected neonates and present as a diffuse eruption of erythematous, crusted papules, papulovesicles, and pustules, often with prominent and initial facial involvement. We describe the case of a male infant with DS and TMD, associated with a vesiculopustular eruption, which appeared on day 36 of life, and review previous cases.

Background: Acute Chest Syndrome (ACS) is the second cause of hospitalization in Sickle Cell Disease (SCD), burdened by significant morbidity and mortality. The guidelines regarding management of ACS are sometimes difficult to follow in the real world and the prevention and treatment strategies of ACS are often applied in an uneven manner in the various settings (community care, regional hospitals, reference university centers). Moreover, epidemiology, clinical phenotype and outcomes as well as risk factors could vary in different populations according to ethnicity, genotype or health care system organization. Aims and Methods: A retrospective multicenter observational study was conducted to investigate the epidemiology of ACS and to the evaluate the diagnostic and therapeutic pathways of ACS in children with SCD (age 0-18 years) in the 2013-2018 period, after the publication of the Italian Association of Pediatric Hematology Oncology (AIEOP) Guidelines for the Management of SCD in Childhood in Italy in 2012. Results: 126 children were recruited and 122 included in the analysis, with 208 evaluable episodes of ACS (range: 1-6 episodes /patient) from 11 AIEOP Centers. 73 M, 49 F. Mean age was 10.9 years. 85% patients were of African origin, 92% were HbSS/SB°; mean age at diagnosis of SCD of the entire cohort was 25,3 months (range 0-16,8). 44.2% of patients had more than one episode of ACS during the study period; 37% had had a previous episode before 2013. 58% had comorbidities, mostly respiratory (asthma or allergy). 75% of the patients underwent disease modifying treatment during study period (73% hydroxyurea, 2% chronic transfusion). The seasonality of ACS episodes was important in our country: 75% of episodes occured between October and March. 95% of ACS episodes were secondary to a Vaso-Occlusive Crisis. 76% of the admissions occurred in SCD reference centers, 24% in regional hospitals, but 30% later required transfer to reference centers for worsening of clinical conditions or need of exchange transfusion. The mean length of hospitalization was 9.6 days (range 1-46); one patient died of pneumococcal sepsis; 6 episodes required transfer to the Intensive Care Unit, mechanical ventilation was required in one episode. A good adherence to the AIEOP Guidelines was documented for some aspects: 99% of the patients were hospitalized, 98% performed chest X-ray for the diagnosis of ACS and in 99% antibiotic therapy was started. Others aspects were less satisfactory and in need of improvement: incentive spirometry was only performed in 19% of admissions; oxygen therapy was performed only in 75% of patients even if SatO2 was8g/dl, while in 16% an exchange transfusion was performed for severe respiratory distress (of these 71% were performed in patients transfered from regional hospitals); 38% required inhaled bronchodilators, 6% steroids. A preliminay evaluation of risk factors for recurrent ACS showed that in our cohort allergy to inhaled allergens (p 0.02) and enuresis (p 0.01) were associated with increased prevalence of recurrent ACS; patients with asthma/wheezing also presented more recurrent ACS compared to patients wihout them (23% vs 13%) but this data did not reach statistical significance. Conclusion: This study represents the first analysis in Italy of ACS, which is confirmed as a frequent event in our cohort, with a significant proportion of patients who experience recurrent ACS. Steps need to be undertaken to improve management of ACS and adherence to the AIEOP guidelines at a national level: stimulate the application of early preventive measures that are still under-utilized, increase the appropriateness of multidisciplinary specialist approach (transfusion specialist, acute care physicians, pneumologists, hematologists) strengthen the dissemination of information through training events for all the Hospitals of the network. Disclosures Colombatti: AddMedica: Consultancy; Global Blood Therapeutics: Consultancy; Novartis: Consultancy.

Hematopoietic stem cell transplantation (HSCT) is the only therapy for a subset of patients with malignant and nonmalignant diseases. Central nervous system (CNS) complications continue to be an important cause of morbidity and significantly contribute to mortality after HSCT. These complications include infections, cerebrovascular lesions, therapy-induced diseases, metabolic disturbances, and post-HSCT carcinogenesis. Following HSCT, three phases can be identified on the basis of the patient's immune status: the pre-engraftment period (30 days after HSCT), the early postengraftment period (30-100 days after HSCT), and the late postengraftment period (100 days after HSCT). There is a distinct relationship between the patient's degree of immunodeficiency after HSCT and the incidence of various complications that may occur. Early diagnosis of CNS complications is crucial for successful management and a good prognosis, and computed tomography and magnetic resonance imaging play an important role in achieving these goals. The global increase in the use of HSCT requires radiologists to be familiar with CNS complications, their relationship to the patient's immune status, and their imaging appearances. This article describes the clinical background of HSCT; reviews the incidence, causes, and timeline of brain complications in children who underwent allogenic HSCT; and identifies the characteristic imaging findings of these disorders.

Relapse remains the leading cause of treatment failure in children with acute lymphoblastic leukaemia (ALL) undergoing allogeneic haematopoietic stem cell transplantation (HSCT). We retrospectively investigated the prognostic role of minimal residual disease (MRD) before and after HSCT in 119 children transplanted in complete remission (CR). MRD was measured by polymerase chain reaction in bone marrow samples collected pre-HSCT and during the first and third trimesters after HSCT (post-HSCT1 and post-HSCT3). The overall event-free survival (EFS) was 50%. The cumulative incidence of relapse and non-relapse mortality was 41% and 9%. Any degree of detectable pre-HSCT MRD was associated with poor outcome: EFS was 39% and 18% in patients with MRD positivity

Although allogeneic haematopoietic stem cell transplantation (HSCT) still represents the only consolidated possibility of cure for sickle cell disease (SCD) patients, its use has been limited by the risk of morbidity and mortality associated with conventional myeloablative therapy. The introduction of treosulfan to replace busulfan in conditioning regimens has recently been explored by virtue of its lower toxicity profile. We report our experience with a treosulfan/thiotepa/fludarabine conditioning for human leucocyte antigen (HLA)-matched sibling or unrelated donor-HSCT in 15 children with SCD, and compare patient outcomes with those of a historical cohort (15 patients) given a busulfan-based regimen. Engraftment was achieved in 28 out of 30 patients (93%), with one case of graft failure in either group. The conditioning regimen was well tolerated in both groups, with no cases of grade III-IV regimen-related toxicity. The 7-year overall survival (OS) and disease-free survival (DFS) for the whole cohort were 100% and 93%, respectively, with a 93% DFS in both busulfan and treosulfan groups. No SCD-related adverse events occurred after engraftment in patients with complete or mixed donor chimerism. This retrospective analysis suggests that a treosulfan-based conditioning regimen is able to ensure engraftment with excellent OS/DFS and low regimen-related toxicity in patients with SCD.

Objective To compare uterine and ovarian volumes and uterine artery (UA) Doppler blood flow among women who were treated with antineoplastic regimens when pediatric aged versus healthy controls. Design Case-control study. Setting Tertiary obstetric and gynecologic center. Patient(s) One hundred twenty-seven women who were treated for childhood cancer with bone marrow transplantation (BMT) and∖or chemotherapy and total body irradiation (TBI) and 64 age-matched healthy controls. Intervention(s) Ultrasonographic and clinical evaluations. Main Outcome Measure(s) Uterine and ovarian volume, detection of follicles, and UA pulsatility index (PI). Result(s) Median uterus and ovarian volumes were reduced by 64% (95% CI, 56.6–70.6) and 83.6% (95% CI, 79.6–86.7), respectively, among cases compared with controls. Median UA PI among cases was increased by 30.3% (95% CI, 19.6–40.8) compared with controls. Ovarian follicles were identified in 24 (18.9%) of 127 cases and 25 (39%) of 64 controls. Uterine volume was reduced after TBI (percent reduction 81.9%; 95% CI, 71.8–87.8) or busulfan (percentage reduction 67.4%; 95% CI, 58.5–75.6) compared with those who had not received a conditioning regimen (percentage reduction 24.4%; 95% CI, 7.6–38.2). The only factors independently associated with reduced uterine and ovarian volumes compared with controls were TBI, busulfan, and BMT. The worst effect on UA PI resulted from BMT and a diagnosis of hematologic disease. Conclusion(s) Bone marrow transplantation as main treatment and TBI and busulfan as conditioning regimens had the worst effect on uterine and ovarian sizes compared with controls. These data should be considered in counseling families on preserving future fertility in children undergoing BMT.

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.

Introduction. Hematopoietic stem cell transplantation (HSCT) exposes the recipient to a high risk of developing viral reactivations especially during the first weeks after HSCT. Neutropenia and lymphopenia, together with the breakdown of physical barriers expose patients not only to bacterial and fungal infections but also to viral infection/reactivations. HHV-6 is usually acquired during infancy, and seroprevalence in the adult population may reach 90%. Like other herpesviruses, HHV-6 persists latently within permissive cells and it can reactivate during immunosuppression. A high incidence of HHV-6 reactivation is frequently detected after HSCT, but its clinical relevance is still debated. The objectives of this retrospective study were to estimate the incidence of HHV6 reactivation after pediatric HSCT, analyze the possible risk factors and evaluate the impact on outcome. Patients and methods. We analyzed 234 pediatric subjects given HSCT from a matched family donor (MFD, 17.5%), a matched unrelated donor (MUD, 43.6%) or a partially matched family donor (PMFD, 38.9%) for malignant (55.2%) or non-malignant hematologic diseases (25.6%) between 2010 and 2017. Data have been analyzed according to patient, donor and transplant characteristics. Risk factors analysis was computed throughout the comparison of the cumulative incidence (CI) of HHV-6 reactivation according to descriptive variables. Kaplan Mayer curves were used to illustrate survival results (OS and EFS), and Log-rank test was used to compare groups. The analysis of outcome was completed by analyzing the CI of neutrophils and platelets engraftment, rejection, acute and chronic GvHD, NRM according to presence or absence of HHV-6 reactivation. Gray-test was used to compare different CI curves. The quantification of HHV-6 exposure was calculated using the area under the curve (AUC) of viremia level over time. AUC expresses the kinetics of the DNA viral load in a time-concentration chart. The stratification of patients according to HHV-6 AUC quartiles (AUCq = each of 4 equal groups into which the HHV6 positive population was divided according to distribution of the value of AUC) allowed to describe the viral burden and the intensity of viral exposure. Risk factors analysis and survival analysis were then performed using the parameter AUC and AUC quartiles. Results. The cumulative incidence of HHV-6 reactivation was 58%, with onset usually within the first month after transplantation (median 19 days). The peak viral load was detected at an average time of 55 days after onset (median 25 days) with a median of 5850 cp/mL (range 50 - 219^106 cp/mL). In univariate analysis, a diagnosis of malignant disease (p=0.021), transplant from PMFD (p=0.010), the use of a busulfan/TBI-based vs Treo-based conditioning regimen (p=0.020), the use of ex vivo T cell-depletion (p Conclusions. Based on the results obtained, HHV-6 infection/reactivation, measured as categorical variable or as viral exposure, does not seem to have an impact on HSCT outcomes. Therefore, routine viral monitoring may not give added benefit in the pediatric HSCT setting. However, in selected populations, early restricted monitoring may identify HSCT recipients at risk of HHV6-related disease. Disclosures Zecca: Chimerix: Honoraria.

Objective: Multipotent mesenchymal stromal cells (MSCs) are endowed with multilineage differentiative potential and immunomodulatory properties. It is still a matter of debate whether donor MSCs have sustained engraftment potential in host bone marrow (BM) after allogeneic hematopoietic stem cell transplantation (allo-HSCT). The aim of this study was to analyze the donor/recipient origin of MSCs in children receiving allogeneic either BM or cord blood (CB) transplantation. Methods: Thirty-seven pediatric patients undergoing allo-HSCT for either a malignant or a nonmalignant disorder were enrolled in the study; 19 received CB and 18 BM transplantation. Results were compared with those obtained in 14 adults given BM transplantation for either malignant or nonmalignant disorders. MSCs were grown from BM aspirates obtained 1-17 and 2-192 months after allo-HSCT in pediatric and adult patients, respectively. MSC samples at the third-fourth passage were phenotypically characterized. Donor/recipient origin of MSCs was assessed by amelogenin assay and microsatellite analysis. Results: MSCs could be grown from 30 of 37 children; at the third-fourth passage MSCs resulted positive (≥98%) for CD73, CD105, CD106, CD29, CD13, CD44 and negative (≤1%) for CD34, CD45, CD14. Mixed chimerism with donor cells was observed in 4 BM and 5 CB transplantation recipients, respectively; full recipient chimerism was detected in the remaining children. Full recipient MSC chimerism was observed also in all assessable (12/14) adult patients. Conclusions: BM of pediatric patients might be a more favorable milieu than that of adults for sustained engraftment of transplanted MSCs. MSCs able to engraft in the host can be transferred with cryopreserved CB units. © 2006 International Society for Experimental Hematology.

Human leucocyte antigen (HLA)-C molecules regulate the function of natural killer cells and may be subdivided into two groups, C(1) and C(2), based on their specificity for inhibitory killer immunoglobulin-like receptors. We analysed the impact of the HLA-C genotype on outcome of HLA-C-matched unrelated donor haematopoietic stem cell transplantation (URD-HSCT) recipients. HLA-C(2) homozygous patients (n = 18) had lower probability of overall survival (P = 0.01) and disease-free survival (P = 0.02), resulting from increased relapse rate (P = 0.02) when compared with both HLA-C(1) homozygous (n = 43) and HLA-C(1),C(2) heterozygous (n = 50) subgroups. Patients lacking HLA-C(1) should, therefore, be considered at increased risk of relapse following HLA-C-matched URD-HSCT.

Leukemia recurrence remains the main cause of treatment failure in children with ALL. The prognosis of children with relapsed ALL is strongly influenced by the site of relapse, time elapsing between diagnosis and recurrence and by blast cell immune-phenotype. Recently, several groups reported probabilities of disease-free survival (DFS) in the order of 60% for children with low/intermediate-risk ALL in 1st relapse using different approaches. Despite this remarkable progress, the best re-induction and consolidation treatment remains to be defined. We, thus, conducted from 10/2003 to 06/2012 a multicenter randomized trial aimed at comparatively evaluating the efficacy of 2 alternative approaches in centers affiliated to the Italian AIEOP network. Included in the study were patients below the age of 18 experiencing late (i.e. more than 6 months from treatment discontinuation) isolated extramedullary (IEM) relapse (S1 patients, #22) or children with B-cell precursor (BCP)-ALL who had early (i.e. between 18 months from diagnosis and 6 months after completion of front-line treatment) or late combined bone marrow (BM) relapse, late isolated BM relapse of BCP–ALL, and very early (i.e. within 18 months from diagnosis) or early IEM of either BCP-ALL or T-cell ALL (S2 patients, # 255). S2 children were randomized to receive induction therapy consisting of either two multiagent chemotherapy courses, F1 and F2 (ARM-A), according to the ALL-REZ BFM P95/96 trial, or a classical 4-drug continuous reinduction therapy (protocol I-A-IDA, ARM-B). After induction, ARM-A patients were given a continuous treatment element including idarubicin (Protocol II-IDA), followed by 5 additional alternating R1/R2 courses and maintenance therapy (24 months). After induction therapy, ARM-B patients received 8 additional alternating R1/R2 courses and maintenance therapy (24 months, see Figure 1). S1 patients received ARM-A treatment with the exception of only 3 alternating R1/R2 consolidation courses and shorter maintenance therapy (12 months). All S2 children with an available HLA-identical sibling were candidate to receive allogeneic hematopoietic stem cell transplantation (allo-HSCT), while those relapsing within 48 months from diagnosis were eligible to receive the allograft also from an unrelated donor (UD) of BM cells or a suitable UD cord blood unit. For S1 patients, both autologous (auto) and allo-HSCT were considered acceptable post-remissional options; auto-HSCT was also employed in a minority of S2 children relapsing more than 48 months from diagnosis. Of the whole cohort of children, 143 were males and 134 were females; median age at diagnosis and at time of relapse was 5 (range 0.2-17) and 9 (range 1.1-17.9) years, respectively. Among the 255 S2 patients, 127 were allocated to ARM-A and 128 to ARM-B. All S1 children obtained a 2nd complete remission (CR), while the probability of reaching 2nd CR of S2 patients was similar in the 2 randomization arms (95% vs 96%). The 6-year DFS for the 22 S1 children was 75% (confidence interval, CI, 55-94); 13 of these children were given either auto-HSCT (#7) or allo-HSCT (#6), while 9 were treated with chemotherapy only. The 6-year DFS for the 255 S2 children was 65% (CI 57-72); the DFS for ARM-A and ARM-B patients was 60% (CI 48-71) and 69% (CI 60-79), respectively (p=ns). Among S2 patients, 179 received allo-HSCT either from an HLA-identical sibling (#51) or a BM/CB UD (#105) or an HLA-partially matched relative (#23) after T-cell depletion of the graft (TCD haplo-HSCT). Of the remaining 76 children, 12 were given auto-HSCT, while 64 received chemotherapy only. The 6-year DFS for S2 children who did or did not receive allo-HSCT was 66% (CI 58-74) and 58% (CI 38-77), respectively (p=ns). Among allo-HSCT recipients, the 6-year DFS was 59% (CI 42-75), 72% (CI 62-82) and 62% (CI 41-83) for children given HLA-identical sibling, UD or TCD haplo-HSCT, respectively (p=ns). Altogether, these results confirm that a high proportion of children with low-/intermediate-risk relapsed ALL can be rescued by second line therapy, including transplantation. A re-induction course similar to that employed in first-line therapy is as effective as that proposed in the ALL-REZ BFM P95/96 trial. UD-HSCT and TCD haplo-HSCT are suitable and effective alternatives for children candidate to an allograft but lacking an HLA-identical sibling. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Evidences have been reported that bone marrow (BM)-derived endothelial progenitor cells (EPCs) circulate in peripheral blood (PB) of healthy subjects. These EPCs seem to play an important role in maintaining homeostasis of vessel walls, participating in both neo-angiogenetic processes and re-endothelization of the wall of injured vessels. The aim of this study was to assess the number and origin of circulating EPCs in PB and BM of children who underwent allogeneic BMT for malignancies. Thirty-five patients with acute lymphoid leukemia (n=18), acute myeloid leukemia (n=13), non Hodgkin lymphoma (n=1), myelodysplastic syndrome (n=1), chronic myeloid leukemia (n=1), and rabdomyosarcoma (n=1) were enrolled in this study. We evaluated PB samples at 21 days, and either PB or BM samples at 45, 60, 90, 120, 180, and 365 days after transplantation. The number of EPCs was evaluated as CD34+VEGFR-2+ or CD34+CD133+VEGFR-2+ cells by cytofluorimetric analysis, and by in vitro culture. PB (n=10) and BM (n=10) samples from age-matched BM donors were analyzed as controls. Donor or recipient origin of EPCs was assessed, by micro-satellite analysis, on at least 10 individually-picked endothelial colonies. The percentage of circulating CD34+VEGFR-2+ cells was significantly lower (p=0.02) in patients tested 21 days after transplant (median 0.01%, 0–1.0) than in PB from controls (median 0.06%, 0–1.3). At the same time point, the percentage of CD34+ co-expressing the CD133 and VEGFR-2 antigens, representing a restricted subset of immature EPCs, was lower, although still not-statistically significant, in patients (median 0.0%, 0.0–4.6) than in controls (median 0.6%, 0.0–15.1); the number of EPC-derived colonies was also significantly lower (p