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International audience; Objectives: To evaluate the applicability and compliance with guidelines for early initiation of long-term prophylaxis in infants with severe hemophilia A and to identify factors associated with guideline compliance.Study design: This real-world, prospective, multicenter, population-based FranceCoag study included almost all French boys with severe hemophilia A, born between 2000 and 2009 (ie, after guideline implementation).Results: We included 333 boys in the study cohort. The cumulative incidence of long-term prophylaxis use was 61.2% at 3 years of age vs 9.5% in a historical cohort of 39 boys born in 1996 (ie, before guideline implementation). The guidelines were not applicable in 23.1% of patients due to an early intracranial bleeding or inhibitor development. Long-term prophylaxis was delayed in 10.8% of patients. In the multivariate analysis, 2 variables were significantly associated with "timely long-term prophylaxis" as compared with "delayed long-term prophylaxis": hemophilia treating center location in the southern regions of France (OR 23.6, 95% CI 1.9-286.7, P = .013 vs Paris area) and older age at long-term prophylaxis indication (OR 7.2 for each additional year, 95% CI 1.2-43.2, P = .031). Long-term prophylaxis anticipation was observed in 39.0% of patients. Earlier birth year (OR 0.5, 95% CI 0.3-0.8, P = .010 for birth years 2005-2009 vs 2000-2004) and age at first factor replacement (OR 1.9 for each additional year, 95% CI 1.2-3.0, P = .005) were significantly associated with "long-term prophylaxis guideline compliance" vs "long-term prophylaxis anticipation."Conclusions: This study suggests that long-term prophylaxis guidelines are associated with increased long-term prophylaxis use. However, early initiation of long-term prophylaxis remains a challenge.

International audience; Introduction: The value and challenges of long-term prophylaxis (LTP) in adolescents and young adults need further characterisation.Aim: To determine the proportions of adolescents and young adults with severe or moderately severe haemophilia in France under LTP and treatment on demand (OD).Methods: Patients 15 to 25 years old with haemophilia A or B, factor VIII/IX ≤ 2% and no current inhibitor could be included if they had been under factor VIII/IX treatment at least 12 months and kept a treatment and bleeding diary.Results: LTP was administered to 169/212 patients (79.7%) and OD treatment to 40/212 patients (18.9%). The most frequent reasons for initiating LTP were joint bleeding, target joints and frequent bleeds; whereas OD treatment was most often selected on the basis of mild bleeding phenotype or because of constraints on LTP. The mean annual bleed rate (ABR) in the OD group (6.33) was higher than in the LTP group (3.07, p 18–21 and > 21–25 years), but was significantly higher for patients with severe haemophilia (4.02) as compared to those with moderate haemophilia (1.97, p = 0.002). No significant difference was observed in mean ABR for joint bleeds between the LTP and OD groups. Physician reported LTP compliance was good or excellent in 97.0% of patients.Conclusion: LTP is the predominant factor VIII/IX treatment among adolescents and young adults with severe or moderately severe haemophilia in France. LTP was associated with low ABR and high compliance. © 2017 The Authors

BACKGROUND: Intrapartum intravenous zidovudine (ZDV) prophylaxis is a long-standing component of prevention of mother-to-child transmission (MTCT) of human immunodeficiency virus (HIV) in high-resource countries. In some recent guidelines, intravenous ZDV is no longer systematically recommended for mothers receiving combination antiretroviral therapy (cART) with low viral load. We evaluated the impact of intravenous ZDV according to viral load and obstetrical conditions. METHODS: All HIV-1-infected women delivering between 1 January 1997 and 31 December 2010 in the French Perinatal Cohort (ANRS-EPF) were analyzed if they received ART during pregnancy and did not breastfeed. We identified maternal and obstetrical characteristics related to lack of intravenous ZDV and compared its association with MTCT rate and other infant parameters, according to various risk factors. RESULTS: Intravenous ZDV was used in 95.2% of the 11 538 deliveries. Older age, multiparity, and preterm and vaginal delivery were associated with lack of intravenous ZDV (n = 554). In women who delivered with viral load ≥1000 copies/mL, the overall MTCT rate was higher without than with intravenous ZDV (7.5% vs 2.9%; P = .01); however, there was no such difference when the neonate received postnatal intensification therapy. Among them, 77% of women who had viral load

Summary. Background: The development of an inhibitor is the major complication facing patients with hemophilia A treated by administration of factor (F) VIII concentrates. Restoration of tolerance to FVIII can be achieved by prolonged administration of FVIII (immune tolerance induction, ITI). Although ITI has been used for more than 30 years in patients with hemophilia A and inhibitor, its mechanism of action is still poorly understood. Objectives: As administration of high doses of antigen can induce the apoptosis of the T cells recognizing the antigen, a potential mechanism of action of ITI may be the deletion of FVIII-specific T cells. Patients/Methods: We studied the CD4+ T-cell response to FVIII in five (one mild, one moderate and three severe) patients successfully desensitized by administration of FVIII and in control subjects. Results: Following repeated stimulation with autologous dendritic cells loaded with FVIII, FVIII-specific T oligoclonal cell lines were expanded from the blood of one of the successfully desensitized patients. The FVIII-specific T cells produced IL-5, IL-13 and IL-2. By contrast, FVIII-specific T-cell lines could not be derived from three patients with mild hemophilia A without inhibitor or from four normal control subjects. Conclusions: These data represent the first analysis of the cellular mechanisms regulating the induction of tolerance to FVIII. They demonstrate that successful tolerance induction may occur without deletion of FVIII-specific T cells.

Purpose To identify prognostic factors and to evaluate the outcome of children with acute lymphoblastic leukemia (ALL) failure after induction therapy. Patients and Methods Between June 1993 and December 1999, 1,395 leukemic children were included in the French Acute Lymphoblastic Leukemia 93 study. Results Fifty-three patients (3.8%) had a leukemic induction failure (LIF) after three- or four-drug induction therapy. In univariate analysis, high WBC count (P = .001), mediastinal mass (P = .017), T-cell phenotype (T-ALL; P = .001), t(9;22) translocation (P = .001), and a slow early response (at day 8 and/or on day 21, P = .001) were predictive of LIF. The following three prognostic groups for LIF were identified by multivariate analysis: a low-risk group with B-cell progenitor (BCP) ALL without t(9;22) (odds ratio [OR] = 1), an intermediate-risk group with T-ALL and a mediastinal mass (OR = 7.4, P < .0001), and a high-risk group with BCP-ALL and t(9;22) or T-ALL without a mediastinal mass (OR = 28.4, P < .0001). Complete remission (CR) was subsequently obtained in 43 patients (81%). The 5-year overall survival (OS) rate of the 53 patients was 30% ± 6%. The 5-year OS rate among allogeneic graft recipients, autologous graft recipients, and after chemotherapy were 30.4% ± 9.6% (50% ± 26% after genoidentical transplantation), 50% ± 17.7%, and 41.7% ± 14.2%, respectively (P = .18). Fourteen patients (26%) were still in first CR after a median of 83 months (range, 53 to 117 months). Conclusion Three risk categories for LIF in children with ALL were identified. Approximately one third of patients with LIF can be successfully treated with salvage therapy overall. Subsequent CR after LIF is mandatory for cure.

The properties of saliva led us to hypothesize that the salivary flow increase induced by gum chewing might protect the oral mucosa from lesions due to cancer chemotherapy. We conducted a multicenter randomized trial to evaluate the efficacy of chewing gum in preventing oral mucositis in 145 children receiving chemotherapy regimens expected to induce WHO grade 3-4 oral mucositis in at least 30% of patients. Patients were allocated at random to standard oral care with or without 5 gum pieces per day. No overall reduction in severe oral mucositis occurred in the gum arm (51%) compared with the standard arm (44%). VIDE, COPADM, and multidrug intensive chemotherapy caused severe oral mucositis in 75% of patients in both arms. In patients receiving less toxic regimens, a decrease in WHO grade 1-4 oral mucositis was noted in the gum arm compared with the standard arm (49% vs. 72%, P=0.03). In the multivariate analysis, the risk of oral mucositis was related only to the type of chemotherapy regimen, suggesting that further strategies for preventing oral mucositis could be mainly based on these criteria.

From 1989 to 1998, 341 children were included in the French multicentric LAME (Leucémie Aiguë Myéloblastique Enfant) trials. A total of 309 children were registered in the LAME 89/91 protocol. This intensive regimen included an induction phase (mitoxantrone plus cytarabine), two consolidation courses, one containing timed-sequential high-dose cytarabine, asparaginase and amsacrine; 276 (90%) achieved a CR. The 5-year overall survival (OS) and event-free survival (EFS) were 60+/-4 and 48+/-4%, respectively. From 1997, timed-sequencing of the LAME SP induction chemotherapy led to an unacceptable frequency of consolidation delay; future improvements are unlikely to come from further increases in intensity. The role of allogenic bone-marrow transplantation from an HLA-identical sibling in CR1 was examined. The disease-free survival (DFS) was 52+/-4% for non-allografted patients and 57+/-7% for allografted patients (P=NS); a better OS for allografted patients was shown and could be related either to allo-BMT early in CR1 or to a second allo-BMT in CR2. For the complete responders after consolidation therapy, the 5-year OS was significantly better in patients randomized for no maintenance therapy (MT-) than in patients randomized for MT (77.6+/-8 vs 59+/-8%; P=0.05), while the 5-year DFS was not significantly different. Exposure to low-dose MT might contribute to clinical drug resistance and treatment failure in relapsing patients.

The objective of the present study was to investigate the role of early common infections and perinatal characteristics in the aetiology of childhood common leukaemia. A case-control study was conducted from 1995 to 1998 in France, and included 473 incident cases of acute leukaemia (AL) (408 acute lymphoblastic leukaemia (ALL), 65 acute myeloid leukaemia (AML) age-, sex- and region-matched with 567 population-based controls. Data on the medical history of the child and his/her environment were collected using self-administered questionnaires. Analyses were conducted using nonconditional logistic regression. A slight negative association with early infections was observed (OR=0.8; 95% CI (0.6-1.0)). The association was stronger for early gastrointestinal infections. Early day-care was found to be associated with a decreased risk of AL (OR=0.6; 95% CI (0.4-0.8) and OR=0.8; 95% CI (0.5-1.2) for day-care starting before age 3 months and between 3 and 6 months, respectively). No association with breast-feeding was observed, irrespective of its duration. A birth order of 4 or more was associated with a significantly increased risk of AL (OR=2.0; 95% CI (1.1-3.7) with ALL). A history of asthma was associated with a decreased risk of ALL (OR 0.5; 95% CI (0.3-0.90). Although the results regarding birth order and breast-feeding do not fit with Greaves' hypothesis, the study supports the hypothesis that early common infections may play a protective role in the aetiology of childhood leukaemia, although this effect was not more marked for common ALL.

PURPOSE: To determine whether the use of a recombinant human granulocyte colony-stimulating factor ([G-CSF] lenogastrim) can increase the chemotherapy dose-intensity (CDI) delivered during consolidation chemotherapy of childhood acute lymphoblastic leukemia (ALL). PATIENTS AND METHODS: Sixty-seven children with very high-risk ALL were randomized (slow early response to therapy, 55 patients; translocation t(9;22) or t(4;11), 12 patients). Consolidation consisted of six courses of chemotherapy; the first, third, and fifth courses were a combination of high-dose cytarabine, etoposide, and dexamethasone (R3), whereas the second, fourth, and sixth courses included vincristine, prednisone, cyclophosphamide, doxorubicin, and methotrexate (COPADM). G-CSF was given after each course, and the next scheduled course was started as soon as neutrophil count was > 1 × 109/L and platelet count was > 100 × 109/L. CDI was calculated using the interval from day 1 of the first course to hematologic recovery after the fifth course (100% CDI = 105-day interval). RESULTS: CDI was significantly increased in the G-CSF group compared with the non–G-CSF group (mean ± 95% confidence interval, 105 ± 5% v 91 ± 4%; P < .001). This higher intensity was a result of shorter post-R3 intervals in the G-CSF group, whereas the post-COPADM intervals were not statistically reduced. After the R3 courses, the number of days with fever and intravenous antibiotics and duration of hospitalization were significantly decreased by G-CSF, whereas reductions observed after COPADM were not statistically significant. Duration of granulocytopenia was reduced in the G-CSF group, but thrombocytopenia was prolonged, and the number of platelet transfusions was increased. Finally, the 3-year probability of event-free survival was not different between the two groups. CONCLUSION: G-CSF can increase CDI in high-risk childhood ALL. Its effects depend on the chemotherapy regimen given before G-CSF administration. In our study, a higher CDI did not improve disease control.

Among risk factors for developing thromboembolism (VTE) in children with acute lymphoblastic leukemia were Escherichia coli asparaginase, concomitant steroid use, presence of central venous lines, and thrombophilic abnormalities. Developing a predictive model for determining children at increased risk would be beneficial in targeting interventional studies to high-risk groups (HRGs). Predictive variables were incorporated into a risk assessment model, which was evaluated in 456 children and then validated in 339 patients. VTE risk by score was no greater than 2.5 for low-risk group (LRG) and greater than 2.5 for HRG. VTE rates at 3.5 months (validation cohorts) were 2.5% in LRG and 64.7% in HRG. In multivariate analysis adjusted for age, duration of asparaginase administration, enoxaparin prophylaxis, and T-immunophenotype, the HRG was significantly associated with VTE compared with the LRG (hazard/95% confidence interval [CI], 8.22/1.85-36.53). Model specificity was 96.2% and sensitivity was 63.2%. As secondary objective we investigated the use of enoxaparin for VTE prophylaxis in the HRG. HRG patients without enoxaparin prophylaxis showed a significantly reduced thrombosis-free survival compared with children on low-molecular-weight heparin (LMWH). On the basis of the high specificity, the model may identify children with leukemia at risk of VTE. LMWH may help prevent VTE in the HRG; this warrants assessment in larger cooperative clinical trials.

International audience; A case-control study was conducted to investigate the role of a familial history of cancer in the etiology of childhood acute leukemia. The history of cancer in the relatives of 472 cases was compared with that of 567 population-based controls. Recruitment was frequency matched on age, sex and region. The familial history of cancer in each child's relatives was reported by the mother in response to a standardized self-administered questionnaire. A familial history of solid tumor in first or second-degree relatives was associated with an increased risk of acute lymphoblastic leukemia (odds ratio (OR)=1.6 [95% confidence interval, 1.2-2.1]), while a familial history of hematopoietic malignancies in first or second-degree relatives was associated with an increased risk of acute myeloid leukemia (OR=4.3 [1.4-13]). The ORs for the histories of cancer increased with the number of relatives with cancer (OR=1.5 [1.1-2.0] for one relative and OR=2.3 [1.3-3.8] for two relatives or more; Ptrend

Zeta-associated protein 70 (ZAP-70), a member of the Syk family of protein tyrosine kinases, is normally expressed in T and NK cells. While little is known about ZAP-70 expression in normal human B cells, it has been reported that ZAP-70 is expressed in a subset of patients with chronic lymphocytic leukemia (CLL) with a poor prognosis. In this study, we examined the expression and phosphorylation status of ZAP-70 in B-lineage acute lymphoblastic leukemia (Blin-ALL).First, ZAP-70 protein expression was assessed by Western blotting and flow cytometry and ZAP-70 mRNA transcripts were analyzed by reverse transcription polymerase chain reaction (RT-PCR) on human precursor B cell lines. Experiments were then carried out on cells obtained from 18 patients with Blin-ALL and from normal human bone marrow.ZAP-70 was constitutively expressed and phosphorylated on tyr319 in human precursor Blin-ALL cell lines as well as in primary B leukemic cells from all examined Blin-ALL patients with pro-B, pre-B and B phenotypes, but not in malignant myeloid cells. Importantly, analysis of normal human bone marrow revealed expression of ZAP-70 transcripts only in the CD34+ cell fraction (either CD19-CD10- or CD19+CD10+) but not in the CD34- cell fraction (CD19+sIgM- pre-B cells or CD19+sIgM+ immature B cells).ZAP-70 was found to be expressed in the CD34+ normal bone marrow compartment including earlier B-cell progenitors, but not in CD34- pre-B and immature B cells. By contrast, ZAP-70 was consistently expressed and phosphorylated in Blin-ALL cells. Further studies are required to determine whether ZAP-70 may play a pathophysiological role in Blin-ALL.

PURPOSE In a previous randomized study, the authors reported that granulocyte colony-stimulating factor (G-CSF) increased the chemotherapy dose-intensity delivered during the consolidation therapy of high-risk childhood acute lymphoblastic leukemia (ALL). The aim of the current study was to perform an economic evaluation in the same cohort. METHODS In this open-label multicenter randomized trial, prophylactic G-CSF was administered after consolidation therapy courses. Economic data were retrospectively quantified for each patient: hospital stays, drugs, and blood products. RESULTS Sixty-seven children were enrolled in the very high-risk branch of the FRALLE 93 protocol. Chemotherapy dose-intensity was significantly increased (105 +/- 5% in the G-CSF group vs. 91 +/- 4% in the non-G-CSF group, P < 0.001). The mean total costs per child were not statistically different: 32,309 dollars in the G-CSF group versus 31,569 dollars in the non-G-CSF group. Further analysis per child and per course (R3 or COPADM) demonstrated that the mean cost of hospitalization and the mean cost of intravenous antibiotics were significantly decreased in the G-CSF group after R3 courses (3,857 dollars vs. 4,993.80 dollars, P < 0.001; 171.40 dollars vs. 306.20 dollars, P = 0.029, respectively), but the cost of platelet transfusion was significantly increased (P = 0.03). Conversely, post-COPADM costs were similar. Finally, mean costs per course in the two randomized groups were not significantly different: 5,848.80 dollars versus 6,181 dollars and 7,388.10 dollars versus 6,475.70 dollars for R3 and COPADM, respectively. The 3-year probability of event-free survival between the two groups was not different. CONCLUSIONS G-CSF can increase chemotherapy dose-intensity in very high-risk ALL without raising costs, but event-free survival was not improved. The cost benefit of prophylactic treatment by G-CSF relies on the chemotherapeutic regimen given prior to G-CSF administration.

Recombinant factor VIIa (rFVIIa; NovoSeven ® , Novo Nordisk, Bagsvaerd, Denmark) appears effective and relatively safe for the treatment of bleeding and for surgical prophylaxis in patients with Glanzmann thrombasthenia as reported to the International Registry on rFVIIa and Congenital Platelet Disorders. One of the shortcomings of the Registry data is the heterogeneity of treatment protocol, including dosage, number of doses used, duration of treatment before declaration of failure, and mode of rFVIIa administration (bolus v continuous infusion). The data are not yet sufficient to define optimal regimens for various indications such as the type of bleeding or the type of procedures. The place of this drug compared to platelet transfusion in the overall management of patients with Glanzmann thrombasthenia will need to be determined in relationship to a number of challenges and unresolved issues in the clinical care of these patients. These issues include: how to improve local measures for patients with mucosal bleeds, optimal management of young women during menarche, optimal platelet transfusion regimens for various indications, the relationship between antiplatelet antibodies detected by monoclonal antibody-specific immobilization of platelet antigens (MAIPA) and effectiveness of platelet transfusion, whether there are other biological tests that may correlate with effectiveness of platelet transfusion, and management of pregnancy and delivery regarding antiplatelet immunization.

This study reports on six cases of deficiency in the human complement regulatory protein Factor H (FH) in the context of an acute renal disease. Five of the cases were observed in children presenting with idiopathic hemolytic uremic syndrome (HUS). Two of the children exhibited a homozygous deficiency characterized by the absence of the 150-kD form of Factor H and the presence, upon immunoblotting, of the 42-kD Factor H-like protein 1 (FHL-1) and other FH-related protein (FHR) bands. Southern blot and PCR analysis of DNA of one patient with homozygous deficiency ruled out the presence of a large deletion of the FH gene as the underlying defect for the deficiency. The other four children presented with heterozygous deficiency and exhibited a normal immunoblotting pattern of proteins of the FH family. Factor H deficiency is the only complement deficiency associated with HUS. These observations suggest a role for FH and/or FH receptors in the pathogenesis of idiopathic HUS.

As a result of steady improvements in chemotherapy over the past 20 years, more than 70% of children with acute lymphoblastic leukemia (ALL) are cured with intensive induction regimens followed by maintenance therapy [1]. However, there is still a 30% therapeutic failure rate manifesting as disease recurrence, generally bone marrow relapses, early in the treatment period. In this context, several centers have reported the safety and efficacy of allogeneic or autologous bone marrow transplantation (BMT) or chemotherapy alone. The better results have been reported in patients who had BMT from a matched sibling donor as compared to those who did not have a matched sibling donor and received chemotherapy alone [2, 3, 4]. There have been few controlled studies comparing results of autologous and allogeneic BMT in similar patients by using the same preparative regimen [5, 6, 7, 8].

Background Clofarabine alone or in combination with cyclophosphamide and etoposide has shown a good efficacy and a tolerable toxicity profile in previous studies of children with relapsed or refractory leukaemia. This report describes a retrospective study of 38 French patients who received clofarabine as a monotherapy or in combination for relapsed or refractory acute lymphoblastic leukaemia (ALL) outside of clinical trials after marketing authorization. Methods We retrospectively analysed data for 38 patients, up to 21 years old, attending 17 French centres. Thirty patients received clofarabine alone or in combination for a bone marrow relapse of acute lymphoblastic leukaemia (ALL) or refractory disease and eight patients for a high level of minimal residual disease (MRD). Survival and response durations were estimated by the Kaplan-Meier method. Results For the 30 patients who received clofarabine for a bone marrow relapse of ALL (number of relapse, 1-3; median, 1), the overall remission rate (ORR) was 37%: eight complete remission (CR) and three complete remission without platelet recovery (CRp). Ten of the 11 responding patients subsequently underwent haematopoietic stem cell transplantation (HSCT). Only four of the eight patients who received clofarabine while in remission for a high level of MRD, showed a moderate improvement of MRD. Seven of these eight patients received HSCT and six of them were alive at the end of the study. One other patient was alive without receiving HSCT. However, clofarabine treatment was associated with a high risk of infection and hepatotoxicity. Febrile neutropenia grade ≥ 3 was reported in 79% of patients and documented infections grade ≥ 3 occurred in nine patients (24%). Hepatotoxicity grade 3 was reported in nine patients (24%). We observed four deaths related to treatment. Conclusion In our experience, the efficacy of clofarabine is poorer than previously reported. Its toxicity is high and can be life threatening. Prospective studies on clofarabine used during earlier phases of the disease may help to define how best this new drug can be exploited for childhood and adolescent ALL.

Objective To better describe the natural history, mode of inheritance, and the epidemiological and clinical features of isolated congenital asplenia, a rare and poorly understood primary immunodeficiency. Study design A French national retrospective survey was conducted in hospital pediatric departments. A definitive diagnosis of ICA was based on the presence of Howell-Jolly bodies, a lack of detectable spleen, and no detectable cardiovascular malformation. Results The study included 20 patients (12 males and 8 females) from 10 kindreds neither related to each other nor consanguineous. The diagnosis of ICA was certain in 13 cases (65%) and probable in 7 cases (35%). Ten index cases led to diagnosis of 10 additional cases in relatives. Five cases were sporadic and 15 were familial, suggesting autosomal dominant inheritance. Median age was 12 months at first infection (range, 2-516 months), 11 months at diagnosis of asplenia (range, 0-510 months), and 9.9 years at last follow-up (range, 0.7-52 years). Fifteen patients sustained 18 episodes of invasive bacterial infection, caused mainly by Streptococcus pneumoniae (61%). Outcomes were poor, with 9 patients (45%) dying from fulminant infection. Conclusions ICA is more common than was previously thought, with an autosomal dominant inheritance in at least some kindreds. Relatives of cases of ICA should be evaluated for ICA, as should children and young adults with invasive infection.

Abstract 710 Background: Pediatric cohort studies in previously untreated [PUPs] children with hemophilia A [HA] identified potential risk factors for clinical meaningful inhibitor development [IH], among them HA severity < 1%, high risk gene mutations [HRS], high dose FVIII replacement therapy [HD] and use of recombinant FVIII concentrates [rFVIII]. The aim of the present study was to develop a simple score to rate inhibitor development in HA children. Patients and methods: 184 consecutively ascertained PUPs [enrollment between 1982 and 2008] with HA [severity < 5%] followed over a period of 200 exposure days were randomly selected into two groups [G1; G2] of 92 patients each. Three variables [HRS: 2 points], [HD: median single dose > 30 IU/kgbw within 8 to 12 weeks after therapy start] and [rFVIII] 1 point each, were incorporated into a risk assessment model, which was i) evaluated in patient group G1 [pilot cohort: patients not included in the CANAL study] and ii) validated in the second group [G2: confirmation cohort]. The risk discrimination cut-off was calculated using two independent statistical methods [ROC-analysis: MedCalc 11.0.0; CART regression tree: CART 6.0]. In addition, model specific cumulative IH-free survival [Cox regression analysis: hazard ratio (HR) and 95% confidence interval (CI)], negative predictive values [NPV], sensitivity and specificity were calculated. In Cox regression analysis adjustment was performed for i) age at first symptomatic bleed [proxy for early start of prophylaxis], ii) severity of factor VIII-deficiency, and iii) for treatment periods [1982-2008: divided into 3 to 5 year intervals]. Results: The risk discrimination cut-off in the pilot cohort G1 was calculated as “2”. Model sensitivity was 78.8 % and specificity was 49.8% respectively. In the confirmation cohort G2 seventeen children with HA developed a high responding IH [18.5%]: 16 out of 52 in children with a risk score > 2 [30.8%] and one out of 40 with a risk score < 2 [2.5%: p 2 was significantly reduced with an adjusted HR [CI] of 11.2 [1.5-83.5]. The corresponding NPV to rule out a high responding IH development in the confirmation cohort was 92% [80.5-98.5]. Conclusion: The model is able to rule out the development of high responding IH in white children with HA < 5% with good discriminative ability. Thus the model can be instrumental to individualize treatment strategies in HA patients. The score needs to be validated in other populations to test its generalizability. Disclosures: No relevant conflicts of interest to declare.

From 06/93 to 12/99, 1395 children and adolescents were included in the FRALLE 93 protocol: group A (Very Low Risk): 182, B (Standard risk): 672 and C (High risk including T-ALL): 541. The median FU is 9.5 yr. Inclusion criterias in group B were: age between 1 and 15 yr, WBC< 100,000, BCP phenotype and no poor-prognosis cytogenetic feature (MLL rearrangement or Ph1). Group B pts were randomised in a 2-steps schedule: at inclusion: daunorubicin (DNR: 40 mg/m2 × 2) vs idarubicin (IDA: 8 mg/m2 × 2), and at consolidation: high-dose IV methotrexate (HD-MTX): 4 × 8g/m2 at D1, D15, D29 and D42, vs low-dose po MTX (LD-MTX): 4 × 25 mg/m2; the number of triple IT was the same for both arms (N = 18). Pts with CNS leukemia were not randomized for MTX. During induction, pts were also classified on D21 bone marrow aspiration according to the level of residual blasts: M1 (< 5%): n=555, M2 (6–25%): n=71 and M3 (> 25%): n=41; M2/M3 pts received 1 more antracyclin infusion at D22 and M3 pts were subsequently treated according to the HR group and not randomized for MTX. Both M1 and M2 pts who reached CR were randomized for MTX. The overall group B EFS, DFS and OS were respectively 80±2, 81±2, and 91±1%. Globally there is no statistically significant impact of 1st or 2nd randomization on the outcome of patients. When we look at the outcome according to the D21 status, the EFS are very different: M1: 84±2%, M2: 63±6% and M3: 74±7%. The very poor EFS of M2 pts, who can not actually be classified as SR pts, and the fact that M1 pts represent 90% of pts randomized for MTX led us to analyse separately M1 pts. For M1 pts there is no interaction between 1st and 2nd randomization for survival, EFS and relapse rate (Gail and Simon test, p = 0.51); therefore pts were pooled for 2nd randomization analysis. Among M1 pts, 271 pts were randomized for HD-MTX and 264 for LD-MTX; there is no difference regarding age or leucocytosis between the 2 arms. Results are as follows: 5-yr EFS Relapses CI iBM relapses CI Other relapses CI 5-yr OS (EFS: event-free survival, CI: cumulative incidence, iBM: isolated bone-marrow, OS: overall survival) HD-MTX 87.82±1.99% 0.12±0.04 7.01±0.02 5.17±0.02 95.20±1.30 LD-MTX 79.92±2.47% 0.20±0.06 10.6±0.03 9.10±0.03 88.64±1.95 Test log-rank Gray Gray Gray log-rank p value 0.028 0.037 0.16 0.18 0.07 At the contrary, among M2 pts there is no difference in EFS for pts treated with HD-MTX (61±7%) and LD-MTX (65±9%) Conclusion: HD-MTX seems to benefit to pts with standard-risk BCP-ALL and good early response to chemotherapy, and the benefit is associated with a reduction in the number of relapses. No benefit is demonstrated in M2 pts with SER who have a very poor outcome and do require a more intensive treatment; the better EFS of M3 patients treated with double delayed intensification is in favour of this hypothesis.

This retrospective study was designed to determine the prevalence of inherited prothrombotic risk factors (Factor V Leiden (FV) G1691A and prothrombin G20210A mutations, TT677 genotype of the methylenetetrahydrofolate reductase (MTHFR), protein C, protein S, antithrombin deficiencies) in a population of children with ALL treated according to the FRALLE 2000 study Protocol (High Risk and Standard Risk groups). The study was performed in 5 French Centers including Amiens, Angers, Paris Trousseau, Rouen and Saint-Etienne. From December 2000 to March 2006, 354 children aged 1 to 18 years old were consecutively admitted for ALL and were enrolled in the FRALLE 2000 Protocol. Among them, 281 patients were investigated for hereditary prothrombotic defects at the time of ALL diagnosis. Informed parental consent was required for gene analysis. Abnormal test results for protein S (functional activity and free protein S antigen concentration), protein C and antithrombin were controlled on a second blood sample after induction. In the population studied, the prevalence of one established prothrombotic risk factor was 19,2%: the FV G1691A mutation was diagnosed in 10 patients (3.6%), all heterozygous, 10 patients (3.6%) showed the heterozygous prothrombin G20210A mutation, the TT677 MTHFR genotype was found in 34 children (12.7%), 1 patient showed protein C deficiency (0.4%). No antithrombin deficiency was detected. The prevalence of inherited protein S deficiency could not be evaluated because of missing data in the family medical history. Combined prothrombotic defects were found in 2 patients (0.71%): heterozygous FV G1691A mutation combined with heterozygous prothrombin G20210A mutation in 1 patient and combined with TT677 MTHFR genotype in the second patient. Except for TT677 MTHFR genotype, the prevalence of hereditary prothrombotic risk factors in children with ALL in France were found within the prevalence reported for children treated for ALL (table 1) and comparable to the prevalence in healthy Europeans (Junker et al. 1999, Margaglione et al 2001, Mueller et al. 2005). Comparison of the prevalence of inherited prothrombotic risk factors in children with ALL Country Population FV G1691A +/− ++ PT G20210A +/− +/+ MTHFR TT677 AT PC AT: antithrombin deficiency ; PC: protein C deficiency ; NE : non evaluated NowakGöttl et al 1999 (n=301) Germany ALL children 5.3% 0.3% 2% 0% 7.7% 0.7% 2.3% Mauz-Körholz et al. 2000 (n=108) Germany ALL children 5.6% 0% 2.8% 0% 5.6% 0% 2.7% Mitchell et al. 2002 (n=60) Canada ALL children 3.3% 0% 2% 0% NE NE NE Present study (n=281) France ALL children 3.6% 0% (n=277) 3.6% 0%(n=279) 2.7% (n=268) 0% 0.4%

In patients with hemophilia A and FVIII inhibitor, tolerance to FVIII can frequently be restored by prolonged administration of FVIII (immune tolerance induction). We studied the cellular response to FVIII in two patients successfully desensitized with high doses of FVIII. Each patient had also transiently received glucocorticoids, when the antibody titer increased after an initial decline upon start of the desensitization. On repeated stimulations with FVIII loaded autologous dendritic cells, FVIII-specific T oligoclonal cell lines and T cell clones were derived from one of the two patients up to two years after complete elimination of the inhibitor. The interleukins produced by the T cell lines included IL-2, IL-5 and IL-13. Although ITI is frequently unsuccessful in mild/moderate haemophilia A patients, this patient remained tolerant to FVIII even after a 10 days FVIII continuous infusion, 18 months after complete elimination of the inhibitor. Several microcultures from CD4+ T cells purified after this treatment responded to FVIII specifically demonstrating the long term persistence of FVIII-specific T cells after ITI. By contrast, no FVIII-specific T cell lines could be derived from blood of control individuals without hemophilia A. Thus, although in animal models of tolerance, administration of high doses of antigen result in T cell elimination, deletion of FVIII-specific T cells is not required for successful tolerance induction to FVIII.

From December 2000 to May 2004, 433 children with SR-BCP-ALL (age: 1–9, WBC 1%). RESULTS: 1. Efficacy : no leukemic induction failure and a low incidence of patients with D21 M2/M3 marrow have been observed (5 % versus 13% in the FRALLE 93 for the same population). Only 4 out 319 evaluable pts (1.3%) have a high EOI MRD (≥ 1%) at the end of induction therapy. Five relapses have only been observed yet even if MFU is still short (31 months). 2.Toxicity: Seven cases of non leukemic deaths have been observed. Main grade III-IV toxicities were attributable to L-Aspa (CNS thrombosis: 5pts, pancreatitis: 8 cases, allergic reaction during DI °2: 35 pts). One case of secondary AML was observed (UPN98: no DNR) (25 months of CR) with translocation t(9;11) and MLL rearrangement. 3.Randomization: at this FU no difference between the DNR+ and DNR- arms is obvious neither in terms of toxicity nor efficacy. CONCLUSIONS: Most of the initial unusual toxicities mentioned where related to asparaginase even if potentially increased by the use of DEX during induction. Since most of the pancreatitis or CNS thrombosis episodes have been reported before July 2002, a potential hypothesis on the variation in the batches of the E.Coli asparaginase used in France can be made beside the classical ones (learning effect ....). Efficacy results, although preliminary, are very encouraging since at the same MFU 4 times more relapses (20 out of 410 pts) were observed in the FRALLE 93 for a comparable population.

The most frequent oncogenic activation events characterized in childhood T acute lymphoblastic leukemia (T-ALL) result in the transcriptional activation of genes coding for transcription factors. The main genes are TAL1/SCL, a member of the basic region helix-loop-helix gene family, HOX11L2 a member of the homeobox-containing protein family. Conflicting results have been reported concerning molecular epidemiology and prognostic values of these markers (Cavé Blood2003:103442–445, Ferrando Cancer Cell 2002, Dicciani Blood 2003 abs 67) we therefore analysed retrospectively 200 pts treated in the French protocol FRALLE 93 for T-ALL between 11/93 and 12/99. Pts were stratified according to prednisone response at D8 ( good or poor : GPR or PPR) and bone marrow at D21. Pts with D8 PPR or M3 received an intensified treatment with genoidentical or autologous transplant in CR1. Molecular analysis was possible for 79/200 T-ALL samples. Clinical caracteristics were not significantly different between population with or without molecular analysis male (n=121) (69% vs 70%), median age 8.4y (range1.1–19.5) vs 9.2y, median leucocytosis 140.109 (0.6–736) vs 171.915 109 (100 n=48), mediastinal involvement 72% vs 71% ,CNS+ 4% vs 5%, CD 10 neg 54% vs 52%. Steroid response PPR n=37/73, GPR n=36/73 and D21 bone marrow status (M3 n=10, M2 n=11) were similar. CR was obtained in 72/79 pts (91%) after first induction therapy and 2 deaths occurs during induction treatment. With a median follow-up of 63 months (2–123), 5 y OS , EFS and DFS is 62 %± 9 and 54% ±10. SIL-TAL1/SCL fusion was detected in 20/79 (25%) pts; expression of HOX11L2 was observed in 14/79 (17%) pts. These activations are mutually exclusive and they allow the subclassification of 42 % of the patients. Median leucocytosis was significantly higher in SIL-TAL1/SCL pts (p=0.01) but other significant features ( ie median age, D8 response, D21 status) were not significantly different between each group. OS and EFS for TAL1/SCL , HOX11L2+ and none of these were respectively 81%±10, 43%±13, 62%± 7 and 80%±10 , 42%± 13, 55%± 7. OS and EFS D8 PPR/GPR were 47%± 9 vs 78% ±7 (p=0.009) and 41%±8 vs 71%±7 (p= 0.008); OS and EFS M2M3 vs M1 D21 bone marrow status were 45%± 15 vs 70% ±6 (p=0.05) and 36%±10 vs 66%± 6 (p= 0.018). In multivariate Cox model analysis, D8 steroid response and HOX11L2 expression were significantly associated with adverse event (failure or relapse) Risk Ratio :3 and 2.6 - p=0.008 and 0.03 and a higher risk of death Risk Ratio : 4.1 and 3.4 -p=0.061 and 0.05. Finally HOX11L2 expression and D8 PPR are independently associated with poor outcome in FRALLE 93 protocol analysis which confirms our previous report. Discrepancies among series may be explained by confounding factors such as differences in median leucocytes value(140.109 in our study) and treatment.

TEL-AML1 fusion transcripts are found in 25% of children with B-cell precursor ALL (BCP-ALL). From June 1993 to December 1999, 1195 children with BCP-ALL were included in the FRALLE 93 protocol. Out of these, 792 were evaluated for TEL-AML1 transcript expression. There is no difference in terms of initial features, DFS, EFS, survival between evaluated (792) and non evaluated (403) patients. Out of the 792 pts, 191 (24%) expressed TEL-AML1 transcripts at diagnosis. To assess the potential prognostic value of TEL-AML1 transcripts quantification, we have retrospectively analysed follow up marrow samples using Europe Against Cancer procedures for real time quantitative RT-PCR assay, on ABI PRISM 7700 (2 reference labs) and Light Cycler apparatus (1 reference lab). Out of the 191 TEL-AML1+ve pts, 83 were evaluated for MRD at different time points after induction therapy (median = D41 (34–55) (53 evaluable pts), at D111 (62–158) (62 pts), at D216 (159–325) (33 pts) and at D838 (365–1287) (49 pts). According to normalized Ct values, samples were attributed to 4 MRD level ranging from 0 to 3 and defined as follows: 0: Ct>40 ; 1 : 36 Distribution of pts according to MRD level at different time points after induction treatment are summarized in the following table. Seventeen relapses have occurred at a median time of 41 months (17–73)(bone marrow: 7, BM + other: 5, testis: 3, CNS: 2). A level 2 positivity at the end of induction was associated with an increased risk of relapse of 3.31(95%CI:1.02 – 10.76, p =.047) while level 3 positivity was associated with a relative risk of 9.52 (95%CI: 2.91 – 31.08, p =.0002). Positivity at D111 was associated with an increased risk of relapse of 8.6 (2.0 – 38.5, p = 0.0042), whatever the level. Combination of data obtained at D41 and D111 allows to distinguish 3 subsets of pts with decreasing relapse-free survival: from 97.5% (95%CI: 85–100%) in pts with no positivity at D111 whatever the D41 result, to 75% (95%CI: 58–92%) in pts with MRD +ve at D111 with low level at D41 and 42% (95%CI: 14–69%) in pts with MRD +ve at D111 with level 2 or 3 at D41 (p Conclusion: RQ-PCR-based MRD detection is a powerful prognostic tool in TEL-AML1+ve leukemia. Combination of two time points allows a relevant stratification of pts according to the risk of relapse, compatible with clinical decision making towards intensification or deescalation in the setting of controlled trials FU time point Number of pts in MRD classes (number of relapses) 0 1 2 3 Not evaluated D41 27 (3) 11 (2) 10 (4) 5 (4) 30 (4) D111 40 (2) 14 (6) 7 (2) 1 (1) 21 (6) D216 29 (2) 2 (1) 1 (0) 1 (1) 50 (13) D838 47 (8) 1 (0) 1 (1) 0 34 (8)

Zeta-associated protein 70 (ZAP-70), a member of the Syk family of protein tyrosine kinases is normally expressed in T and NK cells. ZAP-70 plays a key role in the signaling pathway of the TCR. It has been recently shown that this protein is also expressed in mouse B lineage cells. While little is known about ZAP-70 expression in normal human B cells, it has been reported that ZAP-70 is expressed in a subset of chronic lymphocytic leukemia (CLL), with a poor prognosis, that have nonmutated immunoglobulin variable region genes. In this study, we examined the expression and phosphorylation status of ZAP-70 and Syk in B-lineage acute lymphoblastic leukemia (B-ALL) cells. ZAP-70 and Syk were constitutively expressed and phosphorylated (tyr319 for ZAP and tyr352 for Syk) in human precursor B-ALL cell lines as well as in B leukemic cells from all examined B-ALL patients (n=18). The expression of ZAP-70 was specific to these B-ALL cells as it was not detected in the K562 cell line or malignant myeloid cells. This expression was independant of maturation state of leukemic cells (pro-B or pre-B) and caryotypic status. The expression and phosphorylation of the T-cell “specific” ZAP-70 kinase in all B-ALL cells raises important questions concerning the contribution of this protein to the leukemogenesis process resulting in B-ALL.

Abstract 135 From December 2000 to June 2010, 1201 children with SR-BCP-ALL (age: 1–9 years, WBC Results: 1. Overall efficacy: No leukemic induction failure was observed; 4 induction deaths (0.3%) occurred leading to a 99.7% CR rate. Only 30 out 1097 evaluable pts (2.7%) had a decisional EOI MRD ≥ 1%. Eighty-two relapses have been observed (BM: 47, CNS+: 24, BM+CNS: 9, testis: 2). For the whole population 5-year EFS is 91.5% (95%CI: 89.8–93.3), 5-year OS is 97.4% (95%CI: 96.4–98.4). 5-yr EFS and OS for the A1 and A3 groups are 93.1% vs. 61.2% (p 2. Overall toxicity: 5 non leukemic deaths have been observed after CR (0.4%). Other main toxicities were attributable to ASPA (CNS thrombosis: 1.5%, grade 3–4 pancreatitis: 0.8%, allergic reaction-all grades- during DI °2: 70%). Six cases of secondary neoplasms were observed (AML: 5 pts, astrocytoma: 1 pt). 3. Randomization: 5y EFS and OS from randomization was 92.9% and 97.2% for D21 M1 pts DNR(+) versus 93.3% and 98.2% for D21 M1 DNR(-) pts (p= 0.89 and p= 0.85, respectively). Interestingly EOI MRD levels in the two arms were not different at the sensitivity cut-off of 10−2 (p=0.93) or 10−3(p= 0.74). 4. Prognostic factors: For the whole population, age < 6 years, WBC < 20.000/mm3, ETV6-RUNX1 positivity, as well as D21 marrow M1 and EOI MRD levels of 10−2 and 10−3 were all associated to EFS in univariate analysis. In multivariate analysis remain only WBC < 20.000/mm3 (p=.001), ETV6-RUNX1 positivity (p=.02), EOI MRD levels ≤ 10−3 (p=1.4×10−5). Considering only randomized pts (D21M1 pts) age Conclusions: Very good results were obtained with this VCR-DEX-ASPA oriented protocol. SR-BCP ALL pts which represents around 55 % of all children with ALL can be cured with a modest dose of anthracyclin (75 mg/m2). A further decrease in therapy based on the identified prognostic factors could be proposed. Intensification of the chemotherapy for the rare SR pts with a very high MRD has salvaged half of these pts but the addition of new drugs and/or HSCT is now to be evaluated. Disclosures: No relevant conflicts of interest to declare.