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Purpose: NF-κB signaling is critically important for regulation of both innate and adaptive immune responses. While activation of NF-κB has been implicated in malignancies such as leukemia and lymphoma, loss-of-function mutations affecting different NF-κB pathway components have been shown to cause primary immunodeficiency disorders. Recently, haploinsufficiency of NF-κB1 has been described in three families with common variable immunodeficiency (CVID). Methods and Results: We studied a patient with recurrent respiratory infections and bacterial parapharyngeal abscess. Immunological investigations revealed normal total B- cell numbers, but hypogammaglobulinemia, decreased frequencies of class-switched B cells and impaired T-cell proliferation. Targeted next-generation sequencing using a custom-designed panel comprising all known PID genes (IUIS 2014 classification) and novel candidate genes identified a novel heterozygous frameshift mutation in the NFKB1 gene leading to a premature stop codon (c.491delG; p.G165A*31). We could show that the mutation leads to reduced phosphorylation of p105 upon stimulation, resulting in decreased protein levels of p50. The further disease course was mainly characterized by two episodes of severe EBV-associated lymphoproliferative disease responsive to rituximab treatment. Due to disease severity, the patient is considered for allogeneic hematopoietic stem cell transplantation. Interestingly, the father carries the same heterozygous NFKB1 mutation and also shows decreased frequencies of memory B cells but has a much milder clinical phenotype, in line with a considerable phenotypic disease heterogeneity. Conclusions: Deficiency of NF-κB1 leads to immunodeficiency with a wider phenotypic spectrum of disease manifestation than previously appreciated, including EBV lymphoproliferative diseases as a hitherto unrecognized feature of the disease. [ABSTRACT FROM AUTHOR]

Molecular ON-switches in which a chemical compound induces protein-protein interactions can allow cellular function to be controlled with small molecules. ON-switches based on clinically applicable compounds and human proteins would greatly facilitate their therapeutic use. Here, we developed an ON-switch system in which the human retinol binding protein 4 (hRBP4) of the lipocalin family interacts with engineered hRBP4 binders in a small molecule-dependent manner. Two different protein scaffolds were engineered to bind to hRBP4 when loaded with the orally available small molecule A1120. The crystal structure of an assembled ON-switch shows that the engineered binder specifically recognizes the conformational changes induced by A1120 in two loop regions of hRBP4. We demonstrate that this conformation-specific ON-switch is highly dependent on the presence of A1120, as demonstrated by an ~500-fold increase in affinity upon addition of the small molecule drug. Furthermore, the ON-switch successfully regulated the activity of primary human CAR T cells in vitro. We anticipate that lipocalin-based ON-switches have the potential to be broadly applied for the safe pharmacological control of cellular therapeutics. [ABSTRACT FROM AUTHOR]

Veno-occlusive disease (VOD) remains a serious complication after allogeneic hematopoietic stem cell transplantation (HSCT). Prophylactic use of defibrotide (DF) might further reduce VOD rates but has no impact on the incidence of severe VOD or VOD-associated mortality. We investigated the cost-effectiveness of prophylactic DF according to the British Committee for Standards in Haematology/British Society for Blood and Marrow Transplantation guidelines in 348 children who underwent transplantation between 2001 and 2014 in our hospital, 138 of whom were at risk for VOD. The VOD incidence was 7.4% for the total cohort. Patients at risk had a higher incidence of VOD compared with patients without risk factors (15.2% versus 2.4%, P  < .0001). VOD occurred more often in patients after busulfan-based myeloablative conditioning than in patients after total body irradiation (11.2% versus 3.5%, P  = .001). Donor types or the transplantation-related mortality (TRM) risk score did not correlate with VOD incidence. In 81% of patients who responded to therapeutic DF, VOD resolved completely. Overall VOD-associated mortality was .3% for the complete cohort, 3.7% for patients diagnosed with VOD, and 20% for patients with severe VOD. Neither the cumulative incidence of TRM (19% ± 8% versus 17% ± 2%, P  = .706) nor the median length of hospitalization differed between patients with VOD and patients without. The median costs per HSCT in patients with VOD were about one-third higher than the overall median costs per transplantation at our institution. The calculated total costs of prophylactic DF treatment for 138 patients at risk was almost 6 times as high as the incremental costs for patients with VOD. We conclude that prophylactic DF for children at risk for VOD is not cost-effective with respect to TRM and length of hospital stay. [ABSTRACT FROM AUTHOR]

Reduced-intensity conditioning has improved survival after hematopoietic stem cell transplantation (HSCT) for hemophagocytic lymphohistiocytosis (HLH) at the cost of more frequent mixed chimerism. The minimum level of donor chimerism (DC) required to prevent HLH reactivation in humans remains to be determined. In a multicenter retrospective study, 103 patients transplanted for hereditary HLH (2000-2013) and DC permanently or transiently <75% (overall, CD31, CD561) were analyzed regarding DC, specific immunologic function, occurrence of systemic reactivations (≥5/8 HLH criteria), partial systemic flares (<5 criteria and HLH-directed treatment), isolated central nervous system reactivations, and management. Recurrence was reported in 18 patients (systemic reactivation n = 11, partial flare n=3, isolated central nervous system reactivation n=4). Ten events occurred during profound immune suppression before day 180 (median DC, 10%; range, 1-100%; CD31 if available, otherwise overall DC), which renders a differentiation between secondary post-HSCT HLH and HLH related to the genetic defect difficult. Eight events occurred between 0.5 and 6.7 years post-HSCT (median DC, 13%; range, 0-30%). In=patients, overall and lineage-specific DC were ≤10% for >6 months (median, 5.1; range, 1.1-10 years) without reactivation. A second HSCT was performed in 18 patients (median, DC 4%; range, 0-19%). Death from reactivation occurred in 4 patients (22% of recurrences). Six patients died of transplant complications following a second HSCT(33% of second HSCT).We conclude that a DC>20%-30% is protective against late reactivation. Lower levels do not, however, inescapably result in recurrences. The decision for or against second HSCT must be based on a thorough risk assessment. [ABSTRACT FROM AUTHOR]

The human parechovirus (HPeV), mainly genotype 3, may cause severe illness in young infants and neonates, including sepsis-like illness and central nervous system (CNS) infection. We lack data concerning the impact and symptoms of HPeV infection in infants in Austria. The aim of the study is to evaluate the spectrum of symptoms and findings in infants with the parechovirus in Vienna and its environs. Patients younger than 3 months of age, with clinically suspected sepsis-like illness or CNS infection and a positive polymerase chain reaction (PCR) for HPeV, were included in the study. Medical records were analyzed retrospectively. Twenty patients were included in the study from 2009 to 2013. The most frequent manifestations were fever and neurological symptoms (89 and 80 %, respectively). Fifty percent of the infants had white blood cell counts out of range. The most notable aspect was cerebral hemorrhage in three neonates, which has not been reported earlier in association with HPeV infection.Conclusion: In Austria, HPeV is a relevant pathogen in sepsis-like disease in infants. The clinical presentation is similar to that described in other studies; cerebral hemorrhage is a new aspect.What Is Known: • Parechovirus infection can cause severe illness in infants. • Symptoms have been described to involve all organs; sepsis-like signs, fever, and irritability are most frequent.What Is New: • Also in Austria, HPeV plays an important role in severe illnesses in infants. • Severe intracranial hemorrhage is described as a new finding. [ABSTRACT FROM AUTHOR]

Primary B-cell disorders comprise a heterogeneous group of inherited immunodeficiencies, often associated with autoimmunity causing significant morbidity. The underlying genetic etiology remains elusive in the majority of patients. In this study, we investigated a patient from a consanguineous family suffering from recurrent infections and severe lupuslike autoimmunity. Immunophenotyping revealed progressive decrease of CD19+ B cells, a defective class switch indicated by low numbers of IgM- and IgG-memory B cells, as well as increased numbers of CD21low B cells. Combined homozygosity mapping and exome sequencing identified a biallelic splice-site mutation in protein C kinase δ (PRKCD), causing the absence of the corresponding protein product. Consequently, phosphorylation of myristoylated alanine-rich C kinase substrate was decreased, and mRNA levels of nuclear factor interleukin (IL)-6 and IL-6 were increased. Our study uncovers human PRKCD deficiency as a novel cause of common variable immunodeficiency-like B-cell deficiency with severe autoimmunity. [ABSTRACT FROM AUTHOR]

Abstract: The routine transformation of human CD8pos T cells by Herpesvirus saimiri has so far not been achieved in the case of pre-expanded antigen-specific CTLs. Here we transformed 73% of polyclonal EBV-specific CD8pos T cell cultures using an optimized culture medium supplemented with IL-2, IL-7, IL-12, and TGF-β1. Still, antigen-specific cytotoxicity was frequently lost and analysis of the TCR Vβ-chain repertoire revealed a variable outgrowth of several initially subdominant populations. Limiting dilution cloning of cells in the presence of high titers of HVS did not result in clonal transformation but in the rapid loss of the viral genome in outgrowing clones. In summary, our data suggest that transformation of CD8pos T cells out of bulk cultures can be routinely achieved, while viral transformation itself remains an infrequent event on a per cell basis. The practical use of the improved immortalization of antigen-expanded CD8pos T cell lines, however, is limited by the arbitrary outgrowth of subdominant populations of unpredictable specificity. [Copyright &y& Elsevier]

Abstract: Pro-apoptotic signalling upon toll-like receptor (TLR) stimulation in myeloid cells is normally antagonized by the simultaneous activation of anti-apoptotic pathways. We have previously reported that IFN-α can sensitize human monocytes to apoptosis induction by lipopolysaccharide (LPS). Based on these results we investigated whether similarly apoptosis can be cooperatively induced in myeloid tumor cells. When testing established acute myeloid leukemia (AML) cell lines we found the monocytic cell line THP-1 to be sensitive to IFN-α plus LPS induced apoptosis, which was partially dependent on caspase-8 and was associated with an enhanced expression of Fas/CD95. We extended our study to 29 short term blast lines from patients with AML and observed additive effects of IFN-α and LPS on cell death only with few samples indicating that sensitivity to IFN-α plus LPS inducible apoptosis is present in a fraction of AML samples only with no obvious correlation with certain FAB phenotypes. [Copyright &y& Elsevier]

Background and objective: Busulfan pharmacokinetic studies suggest that an individual dosing strategy may be necessary to optimise systemic exposure in order to decrease toxicity and improve outcome in haematopoietic stem cell transplantation. Therapeutic and toxic effects of the busulfan/cyclophosphamide regimen have been related to the area under the busulfan plasma concentration-time curve. Because of practical limitations in obtaining blood from children, saliva was evaluated as an alternative matrix for therapeutic drug monitoring, offering the advantages of a non-invasive, rapid and easy sampling procedure. Another objective was to evaluate an easy and robust liquid chromatography- tandem mass spectrometry method for plasma and saliva busulfan determination. Methods: An online extraction cartridge with column-switching technique, analytical liquid chromatography over a Chromolith RP 18e column, and tandem mass spectrometry were used to quantify busulfan concentrations in matched plasma and saliva samples. The study population consisted of ten patients, aged 1.3–19 years (median age 11.8 years, seven females, three males), undergoing haematopoietic stem cell transplantation. All patients received busulfan 0.8–1.3 mg/kg orally every 6 hours for a total of 16 doses, followed by two doses of cyclophosphamide (60 mg/kg/day). Results: The lowest limit of detection was 2 µg/L and the lower limit of quantification was 10 µg/L. Only 100µL of plasma/saliva was needed. The mean recoveries (SD) of busulfan were 97.2% (2.7) in plasma and 100.4% (1.3) in saliva. Intra- and inter-assay imprecision was 2–3% and 2–4% for plasma, and 1–2% and 2–4% for saliva (concentration range 30–1500 µg/L). The bias was <4% for both plasma and saliva. The correlation between the busulfan concentration in plasma and saliva was highly significant (r = 0.958; p < 0.0001; saliva/plasma ratio = 1.09 ± 0.04; n = 69 sample pairs). The apparent plasma clearance was slightly higher than the apparent saliva clearance (202 ± 31 mL/h/kg vs 189 ± 28 mL/h/kg; p = 0.001). The mean elimination half-life was found to be 2.31 ± 0.46 hours for plasma and 2.30 ± 0.36 hours for saliva; these were not significantly different (p = 0.83). Conclusion: The present study demonstrated that busulfan analysis in saliva could be a valuable and reliable alternative to plasma analysis. [ABSTRACT FROM AUTHOR]

Dendritic cells (DC) are candidates for antigen-presenting cells that present exogenous antigen on MHC class I molecules to cytotoxic T lymphocytes (CTL), a process referred to as cross-priming. We triggered interleukin (IL)-12 release from DC, which was limited to the first day after maturation induction, by a combination of lipopolysaccharide (LPS) and interferon (IFN)-γ. To stimulate T lymphocytes, we used soluble protein derived from lysis of Epstein-Barr virus (EBV)-transformed lymphoblastoid cell lines (LCL) or ovalbumin loaded onto DC. Co-culture was initiated 2–6 or 48 h after maturation corresponding to “semi-mature” actively IL-12-secreting type 1 DC (sm-DC1) or a “fully mature” DC1 that had lost the ability to release IL-12 (fm-DC1), respectively. IL-12-secreting sm-DC1 but not fm-DC1 efficiently triggered cytolytic activity in autologous T lymphocytes. The combination of IL-1β, IL-6, TNF-α, and prostaglandin E2 generated type 2 DC that did not secrete IL-12 (DC2) and could not prime T-cell cytolytic activity. However, supplementation of cultures using DC2 with IL-12 resulted in CTL activity while the presence of anti-IL-12 monoclonal antibodies in cultures using IL-12 secreting sm-DC1 suppressed CTL activity. Thus, actively IL-12-secreting sm-DC1 are necessary and sufficient for the antigen-specific expansion of CTL in response to exogenously provided soluble antigen. [ABSTRACT FROM AUTHOR]

A response by Ornella Parolini, Gabriele Ressmann, Oskar A. Haas, Johanna Pawlowsky, Helmut Gadner, Walter Knapp, and Wolfgang Holter to a letter to the editor about their article "X-Linked Wiskott-Aldrich Syndrome in a Girl" in the January 29, 1998 issue is presented.

Allogeneic hematopoietic stem cell transplantation (HSCT) is the gold standard curative therapy for infants and children with many inborn errors of immunity (IEI), but adolescents and adults with IEI are rarely referred for transplant. Lack of published HSCT outcome data outside small, single-center studies and perceived high risk of transplant-related mortality have delayed the adoption of HSCT for IEI patients presenting or developing significant organ damage later in life. This large retrospective, multicenter HSCT outcome study reports on 329 IEI patients (age range, 15-62.5 years at HSCT). Patients underwent first HSCT between 2000 and 2019. Primary endpoints were overall survival (OS) and event-free survival (EFS). We also evaluated the influence of IEI-subgroup and IEI-specific risk factors at HSCT, including infections, bronchiectasis, colitis, malignancy, inflammatory lung disease, splenectomy, hepatic dysfunction, and systemic immunosuppression. At a median follow-up of 44.3 months, the estimated OS at 1 and 5 years post-HSCT for all patients was 78% and 71%, and EFS was 65% and 62%, respectively, with low rates of severe acute (8%) or extensive chronic (7%) graft-versus-host disease. On univariate analysis, OS and EFS were inferior in patients with primary antibody deficiency, bronchiectasis, prior splenectomy, hepatic comorbidity, and higher hematopoietic cell transplant comorbidity index scores. On multivariable analysis, EFS was inferior in those with a higher number of IEI-associated complications. Neither age nor donor had a significant effect on OS or EFS. We have identified age-independent risk factors for adverse outcome, providing much needed evidence to identify which patients are most likely to benefit from HSCT., (© 2022 by The American Society of Hematology.)

Data on stem cell transplantation (SCT) for Diamond-Blackfan Anemia (DBA) is limited. We studied patients transplanted for DBA and registered in the EBMT database. Between 1985 and 2016, 106 DBA patients (median age, 6.8 years) underwent hematopoietic stem cell transplantation from matched-sibling donors (57%), unrelated donors (36%), or other related donors (7%), using marrow (68%), peripheral blood stem cells (20%), both marrow and peripheral blood stem cells (1%), or cord blood (11%). The cumulative incidence of engraftment was 86% (80% to 93%), and neutrophil recovery and platelet recovery were achieved on day +18 (range, 16 to 20) and +36 (range, 32 to 43), respectively. Three-year overall survival and event-free survival were 84% (77% to 91%) and 81% (74% to 89%), respectively. Older patients were significantly more likely to die (hazard ratio, 1.4; 95% confidence interval, 1.06 to 1.23; P < .001). Outcomes were similar between sibling compared to unrelated-donor transplants. The incidence of acute grades II to IV of graft-versus-host disease (GVHD) was 30% (21% to 39%), and the incidence of extensive chronic GVHD was 15% (7% to 22%). This study shows that SCT may represent an alternative therapeutic option for transfusion-dependent younger patients., (Copyright © 2021. Published by Elsevier Inc.)

T cells engineered to express chimeric antigen receptors (CAR-T cells) have shown impressive clinical efficacy in the treatment of B cell malignancies. However, the development of CAR-T cell therapies for solid tumors is hampered by the lack of truly tumor-specific antigens and poor control over T cell activity. Here we present an avidity-controlled CAR (AvidCAR) platform with inducible and logic control functions. The key is the combination of (i) an improved CAR design which enables controlled CAR dimerization and (ii) a significant reduction of antigen-binding affinities to introduce dependence on bivalent interaction, i.e. avidity. The potential and versatility of the AvidCAR platform is exemplified by designing ON-switch CARs, which can be regulated with a clinically applied drug, and AND-gate CARs specifically recognizing combinations of two antigens. Thus, we expect that AvidCARs will be a highly valuable platform for the development of controllable CAR therapies with improved tumor specificity.

AML SCT-BFM 2007 was the first hematopoietic stem cell transplantation (HCT) trial in Germany to comply with the European Clinical Trials Directive, and aimed to standardize pediatric HCT for acute myeloid leukemia (AML) across centers in Germany, Austria, and the Czech Republic. Children with high-risk features and a good early response achieving a complete first remission (CR-1) and those in CR-2 after a first relapse were stratified to receive HCT from a matched donor after myeloablative conditioning consisting of busulfan, cyclophosphamide, and melphalan. Four-year EFS and OS were 61 and 70%. Cumulative incidence of relapse (CIR) was 22%. TRM was 15% and correlated with age reaching 9% (SE 3%) in children younger than 12 years and 31% (SE 9%) in older children and adolescents. Children with poorly responding primary disease or relapse were allocated to receive early HCT after a cytoreductive regimen with fludarabine, amsacrine, and cytarabine, followed by reduced intensity conditioning and prophylactic donor lymphocyte infusions. Four-year EFS and OS were 49 and 53%. CIR was 38% and TRM 11%. For patients with primary poor response disease, early use of RIC HCT followed by prophylactic DLI can induce long-term remissions in more than 50% (EFS 46% (SE 9%)).

Langerhans cell histiocytosis (LCH) is a MAPK pathway-driven disease characterized by the accumulation of CD1a + langerin + cells of unknown origin. We have previously reported that the Notch signaling pathway is active in LCH lesions and that the Notch ligand Jagged2 (JAG2) induces CD1a and langerin expression in monocytes in vitro. Here we show that Notch signaling induces monocytes to acquire an LCH gene signature and that Notch inhibition suppresses the LCH phenotype. In contrast, while also CD1c + dendritic cells or IL-4-stimulated CD14 + monocytes acquire CD1a and langerin positivity in culture, their gene expression profiles and surface phenotypes are more different from primary LCH cells. We propose a model where CD14 + monocytes serve as LCH cell precursor and JAG2-mediated activation of the Notch signaling pathway initiates a differentiation of monocytes toward LCH cells in selected niches and thereby contributes to LCH pathogenesis., (©2018 Society for Leukocyte Biology.)

Developmental tumors in children and young adults carry few genetic alterations, yet they have diverse clinical presentation. Focusing on Ewing sarcoma, we sought to establish the prevalence and characteristics of epigenetic heterogeneity in genetically homogeneous cancers. We performed genome-scale DNA methylation sequencing for a large cohort of Ewing sarcoma tumors and analyzed epigenetic heterogeneity on three levels: between cancers, between tumors, and within tumors. We observed consistent DNA hypomethylation at enhancers regulated by the disease-defining EWS-FLI1 fusion protein, thus establishing epigenomic enhancer reprogramming as a ubiquitous and characteristic feature of Ewing sarcoma. DNA methylation differences between tumors identified a continuous disease spectrum underlying Ewing sarcoma, which reflected the strength of an EWS-FLI1 regulatory signature and a continuum between mesenchymal and stem cell signatures. There was substantial epigenetic heterogeneity within tumors, particularly in patients with metastatic disease. In summary, our study provides a comprehensive assessment of epigenetic heterogeneity in Ewing sarcoma and thereby highlights the importance of considering nongenetic aspects of tumor heterogeneity in the context of cancer biology and personalized medicine.

Unlabelled: Allogeneic hematopoietic stem cell transplantation is required as rescue therapy in about 20% of pediatric patients with acute lymphoblastic leukemia. However, the relapse rates are considerable, and relapse confers a poor outcome. Early assessment of the risk of relapse is therefore of paramount importance for the development of appropriate measures. We used the EuroChimerism approach to investigate the potential impact of lineage-specific chimerism testing for relapse-risk analysis in 162 pediatric patients with acute lymphoblastic leukemia after allogeneic stem cell transplantation in a multicenter study based on standardized transplantation protocols. Within a median observation time of 4.5 years, relapses have occurred in 41/162 patients at a median of 0.6 years after transplantation (range, 0.13-5.7 years). Prospective screening at defined consecutive time points revealed that reappearance of recipient-derived cells within the CD34(+) and CD8(+) cell subsets display the most significant association with the occurrence of relapses with hazard ratios of 5.2 (P=0.003) and 2.8 (P=0.008), respectively. The appearance of recipient cells after a period of pure donor chimerism in the CD34(+) and CD8(+) leukocyte subsets revealed dynamics indicative of a significantly elevated risk of relapse or imminent disease recurrence. Assessment of chimerism within these lineages can therefore provide complementary information for further diagnostic and, potentially, therapeutic purposes aiming at the prevention of overt relapse. This study was registered at clinical., Trials: gov with the number NC01423747., (Copyright© Ferrata Storti Foundation.)

The pharmacokinetics, pharmacodynamics, efficacy and safety of a new recombinant E. coli-asparaginase preparation were evaluated in infants (<1 year of age) with de novo acute lymphoblastic leukemia. Twelve patients were treated according to the INTERFANT-06 protocol and received up to 10,000 U/m(2) recombinant asparaginase as intravenous infusions on days 15, 18, 22, 25, 29 and 33 of remission induction treatment. The asparaginase dose was individually adjusted by protocol to 67% of the calculated dose for infants <6 months, and to 75% of the calculated dose for infants aged 6-12 months. The trough serum asparaginase activities observed were above 20, 50, and 100 U/L in 86%, 71%, and 51% of measured samples, respectively. Looking only at the data assessed 3 days after asparaginase infusion these percentages were 91%, 84%, and 74%, respectively. Asparagine was completely depleted in serum in all but one patient who was the youngest in the study. No anti-asparaginase antibodies were detected during this treatment phase. Observed adverse reactions are known to be possible and are labeled side effects of asparaginase treatment and chemotherapy. We conclude that the asparaginase dose regimen used in infants is safe and provides complete asparagine depletion for the desired time period in nearly all patients. Measured asparaginase trough serum levels justify the higher doses used in infants compared to in older children and show that 3-day intervals are preferred over 4-day intervals. (This trial was registered at www.clinicaltrialsregister.eu as EudraCT number 2008-006300-27).

Current evidence suggests that maturing dendritic cells (DCs) acquire a migratory phenotype to induce T cell responses in lymph nodes or a proinflammatory phenotype to condition the microenvironment at peripheral sites. We show that the interplay of PGE(2) and IFN-gamma generates a more complex pattern of mixed DC phenotypes in response to TLR stimulation. DCs activated by the TLR ligand R-848 in the presence of IFN-gamma and PGE(2) produced high levels of IL-12p70 and IL-23, started migration toward CCL19 within only 10 h, and still continued to secrete IL-12p70 without further restimulation following the migration step. The accelerated onset of migration was a result of PGE(2) and was associated with reduced plastic adherence and lower amounts of activated CD29. In contrast, IFN-gamma by itself enhanced cell adhesion and strongly hindered CCR7-mediated migration in the absence of PGE(2). This suggests a new role for IFN-gamma in the direct regulation of DC migration through enhanced cell adhesion, perhaps to support the development of T cell effector functions at peripheral sites. Together, our data are relevant to the development of DC vaccines, as they demonstrate the existence of dual-functional DCs, which as a result of the simultaneous effects of PGE(2) and IFN-gamma, can migrate rapidly toward lymph node chemokines and carry with them a wave of primary cytokines.

Purpose: To identify results and prognostic factors on long-term survival and local control following treatment of localized Ewing's sarcoma., Patients and Methods: Between 1979 and 2004, a total of 60 children and young adults with Ewing's sarcoma were treated. Patients with distant metastases at presentation (n = 6) and recurrent cases (n = 2) were excluded from this analysis. Patients were exclusively treated within ongoing national and international protocols CESS-81, CESS-86, EICESS-92, EURO-EWING-99. All patients received local irradiation with a total dose of 45-60 Gy; in addition, 41 (79%) of the patients had local surgical procedures, 27 (52%) of them with clear margins., Results: Overall survival rates at 5 and 10 years were 56% and 45%, respectively. Patients