The genesis of paediatric haematology in the UK.

Paediatric haematology began to establish itself as a speciality in the UK just over 60 years ago. In that time, clinical trials involving all the specialist centres in the country, and based on scientific advances, have dramatically improved the outlook for children with a range of malignant and non-malignant disorders, but particularly acute leukaemia. As in many specialties, multidisciplinary teams have played a major role in delivering these advances. With these structures in place at a national level, perhaps, of all specialities, paediatric haematology is poised to benefit from the new developments in precision medicine, gene editing and immunotherapy.

Predicting microbiologically defined infection in febrile neutropenic episodes in children: global individual participant data multivariable meta-analysis.

BACKGROUND: Risk-stratified management of fever with neutropenia (FN), allows intensive management of high-risk cases and early discharge of low-risk cases. No single, internationally validated, prediction model of the risk of adverse outcomes exists for children and young people. An individual patient data (IPD) meta-analysis was undertaken to devise one. METHODS: The 'Predicting Infectious Complications in Children with Cancer' (PICNICC) collaboration was formed by parent representatives, international clinical and methodological experts. Univariable and multivariable analyses, using random effects logistic regression, were undertaken to derive and internally validate a risk-prediction model for outcomes of episodes of FN based on clinical and laboratory data at presentation. RESULTS: Data came from 22 different study groups from 15 countries, of 5127 episodes of FN in 3504 patients. There were 1070 episodes in 616 patients from seven studies available for multivariable analysis. Univariable analyses showed associations with microbiologically defined infection (MDI) in many items, including higher temperature, lower white cell counts and acute myeloid leukaemia, but not age. Patients with osteosarcoma/Ewings sarcoma and those with more severe mucositis were associated with a decreased risk of MDI. The predictive model included: malignancy type, temperature, clinically 'severely unwell', haemoglobin, white cell count and absolute monocyte count. It showed moderate discrimination (AUROC 0.723, 95% confidence interval 0.711-0.759) and good calibration (calibration slope 0.95). The model was robust to bootstrap and cross-validation sensitivity analyses. CONCLUSIONS: This new prediction model for risk of MDI appears accurate. It requires prospective studies assessing implementation to assist clinicians and parents/patients in individualised decision making.

Improved outcome with hematopoietic stem cell transplantation in a poor prognostic subgroup of infants with mixed-lineage-leukemia (MLL)-rearranged acute lymphoblastic leukemia: results from the Interfant-99 Study.

To define a role for hematopoietic stem cell transplantation (HSCT) in infants with acute lymphoblastic leukemia and rearrangements of the mixed-lineage-leukemia gene (MLL(+)), we compared the outcome of MLL(+) patients from trial Interfant-99 who either received chemotherapy only or HSCT. Of 376 patients with a known MLL status in the trial, 297 (79%) were MLL(+). Among the 277 of 297 MLL(+) patients (93%) in first remission (CR), there appeared to be a significant difference in disease-free survival (adjusted by waiting time to HSCT) between the 37 (13%) who received HSCT and the 240 (87%) who received chemotherapy only (P = .03). However, the advantage was restricted to a subgroup with 2 additional unfavorable prognostic features: age less than 6 months and either poor response to steroids at day 8 or leukocytes more than or equal to 300 g/L. Ninety-seven of 297 MLL(+) patients (33%) had such high-risk criteria, with 87 achieving CR. In this group, HSCT was associated with a 64% reduction in the risk of failure resulting from relapse or death in CR (hazard ratio = 0.36, 95% confidence interval, 0.15-0.86). In the remaining patients, there was no advantage for HSCT over chemotherapy only. In summary, HSCT seems to be a valuable option for a subgroup of infant MLL(+) acute lymphoblastic leukemia carrying further poor prognostic factors. The trial was registered at www.clinicaltrials.gov as #NCT00015873 and at www.controlled-trials.com as #ISRCTN24251487.

Cytogenetics of childhood acute myeloid leukemia: United Kingdom Medical Research Council Treatment trials AML 10 and 12.

PURPOSE: Karyotype is an independent indicator of prognosis in acute myeloid leukemia (AML) that is widely applied to risk-adapted therapy. Because AML is rare in children, the true prognostic significance of individual chromosomal abnormalities in this age group remains unclear. PATIENTS AND METHODS: This cytogenetic study of 729 childhood patients classified them into 22 subgroups and evaluated their incidence and risk. RESULTS: Rearrangements of 11q23 were the most frequent abnormality found in approximately 16% of patients, with 50% of these in infants. The outcome for all patients with 11q23 abnormalities was intermediate; no difference was observed for those with t(9;11)(p21-22;q23). The core binding factor leukemias with the translocations t(8;21)(q22;q22) and inv(16)(p13q22) occurred at incidences of 14% and 7%, respectively, predominantly in older children, and their prognosis was favorable. An adverse outcome was observed in patients with monosomy 7, abnormalities of 5q, and t(6;9)(p23;q34). Abnormalities of 3q and complex karyotypes, in the absence of favorable-risk features, have been associated with an adverse outcome in adults, but the results were not significant in this childhood series. However, the presence of 12p abnormalities predicted a poor outcome. CONCLUSION: Because the spectrum of chromosomal changes and their risk association seem to differ between children and adults with AML, biologic differences are emerging, which will contribute to the redefinition of risk stratification for different age groups in the future.

Outcome of congenital acute lymphoblastic leukemia treated on the Interfant-99 protocol.

Acute lymphoblastic leukemia (ALL) diagnosed in the first month of life (congenital ALL) is very rare. Although congenital ALL is often assumed to be fatal, no studies have been published on outcome except for case reports. The present study reports the outcome of 30 patients with congenital ALL treated with the uniform Interfant-99 protocol, a hybrid regimen combining ALL treatment with elements designed for treatment of acute myeloid leukemia. Congenital ALL was characterized by a higher white blood cell count and a strong trend for higher incidence of MLL rearrangements and CD10-negative B-lineage ALL compared with older infants. Induction failure rate was 13% and not significantly different from that in older infants (7%, P = .14), but relapse rate was significantly higher in congenital ALL patients (2-year cumulative incidence [SE] was 60.0 [9.3] vs 34.2 [2.3], P < .001). Two-year event-free survival and survival of congenital ALL patients treated with this protocol was 20% (SE 9.1%). Early death in complete remission and treatment delays resulting from toxicity were not different. The survival of 17% after last follow-up, combined with a toxicity profile comparable with that in older infants, justifies treating congenital ALL with curative intent. This trial was registered at www.clinicaltrials.gov as no. NCT 00015873, and at www.controlled-trials.com as no. ISRCTN24251487.

Pharmacokinetics of high-dose methotrexate in infants treated for acute lymphoblastic leukemia.

BACKGROUND: Interfant-99 was an international collaborative treatment protocol for infants with acute lymphoblastic leukemia (ALL). PROCEDURE: We collected data on 103 infants at the time of their first treatment with high-dose methotrexate (HD MTX), 5 g/m(2). Children <6 months of age received two-third of the calculated dose based on body surface area (BSA), children 6-12 months three-fourth of the calculated dose, and children >12 months full dose. RESULTS: The median steady-state MTX concentration at the end of the 24-hr infusion was 57.8 microM (range 9.5-313). The median systemic clearance was 6.22 L/hr/m(2) BSA, and tended to increase with age (P = 0.099). Boys had higher clearance than girls, 6.77 and 5.28 L/hr/m(2) (P = 0.030), and tended to have lower median MTX concentration at 24 hr. Eight infants had MTX levels below 20 microM, a level judged to be sufficient in B-lineage ALL in children >1 year of age. All infants tolerated the dose well enough to receive a second dose of HD MTX without dose reduction. We found no significant effect on disease-free survival for MTX steady-state concentration, MTX clearance, or time to MTX below 0.2 microM. CONCLUSIONS: Our data provide no support for a change in the dosing rules for MTX used in Interfant-99. However, in view of the poor treatment results for infants, one might consider increase in the dose for patients who reach plasma levels below median after the first MTX dose.

A treatment protocol for infants younger than 1 year with acute lymphoblastic leukaemia (Interfant-99): an observational study and a multicentre randomised trial.

BACKGROUND: Acute lymphoblastic leukaemia in infants younger than 1 year is rare, and infants with the disease have worse outcomes than do older children. We initiated an international study to investigate the effects of a new hybrid treatment protocol with elements designed to treat both acute lymphoblastic leukaemia and acute myeloid leukaemia, and to identify any prognostic factors for outcome in infants. We also did a randomised trial to establish the value of a late intensification course. METHODS: Patients aged 0-12 months were enrolled by 17 study groups in 22 countries between 1999 and 2005. Eligible patients were stratified for risk according to their peripheral blood response to a 7-day prednisone prophase, and then given a hybrid regimen based on the standard protocol for acute lymphoblastic leukaemia, with some elements designed for treatment of acute myeloid leukaemia. Before the maintenance phase, a subset of patients in complete remission were randomly assigned to receive either standard treatment or a more intensive chemotherapy course with high-dose cytarabine and methotrexate. The primary outcomes were event-free survival (EFS) for the initial cohort of patients and disease-free survival (DFS) for the patients randomly assigned to a treatment group. Data were analysed on an intention-to-treat basis. This trial was registered with ClinicalTrials.gov, number NCT 00015873, and at controlled-trials.com, number ISRCTN24251487. FINDINGS: In the 482 enrolled patients who underwent hybrid treatment, 260 (58%) were in complete remission at a median follow-up of 38 (range 1-78) months, and EFS at 4 years was 47.0% (SE 2.6, 95% CI 41.9-52.1). Of 445 patients in complete remission after 5 weeks of induction treatment, 191 were randomised: 95 patients to receive a late intensification course, and 96 to a control group. At a median follow-up of 42 (range 1-73) months, 60 patients in the treatment group and 57 controls were disease-free. DFS at 4 years did not differ between the two groups (60.9% [SE 5.2] for treatment group vs 57.0% [5.5] for controls; p=0.81). During the intensification phase, of 71 patients randomly assigned to the treatment group, and for whom toxicity data were available, 35 (49%) had infections, 21 (30%) patients had mucositis, 22 (31%) patients had toxic effects on the liver, and 2 (3%) had neurotoxicity. All types of rearrangements in the (mixed lineage leukaemia) MLL gene, very high white blood cell count, age of younger than 6 months, and a poor response to the prednisone prophase were independently associated with inferior outcomes. INTERPRETATION: Patients treated with our hybrid protocol, and especially those who responded poorly to prednisone, had higher EFS than most reported outcomes for treatment of infant ALL. Delayed intensification of chemotherapy did not benefit patients.

Adolescents with acute lymphoblastic leukaemia: outcome on UK national paediatric (ALL97) and adult (UKALLXII/E2993) trials.

BACKGROUND: Adolescents with acute lymphoblastic leukaemia (ALL) have languished in the shadow of success of the outcome of therapy in childhood ALL. Their treatment has always been incorporated into either paediatric or adult clinical trials depending on the mode of referral and hence there is a need to address an age and risk specific strategy for improving the outcome of this neglected group of patients. This article has summarised the recent and updated retrospective comparative analysis of adolescents treated on the Medical Research Council (MRC) trials. This analysis adds further emphasis to the treatment approach and the merits and limitations of treatment of adolescents on paediatric and adult trials. METHODS: A retrospective comparative analysis of adolescents aged 15-17 years, treated on either MRC ALL97/revised 99 (n = 61), a randomised paediatric trial or UKALLXII/E2993 (n = 67), an adult trial, between 1997 and 2002 was undertaken. RESULT: Results suggest a trend towards a superior outcome on paediatric trials. The 5-year EFS on ALL97 was 65% (95% CI = 52-78%) and on UKALLXII/E2993 was 49% (95% CI = 37-61%; P = 0.01). Multivariate analysis allowing for age and Ph status, diminished the EFS difference, but confirmed a reduced rate of death in remission in patients managed on the paediatric protocol. CONCLUSIONS: Despite limitations in the methodology, comparative studies including our MRC study suggest a consistent advantage for adolescents managed intensively on paediatric trials. Redefining age limits with risk-based strategy and multi-centre collaboration should be considered to improve the survival of young adults.

Single agent efficacy of rituximab in childhood immunosuppression related lymphoproliferative disease: a United Kingdom Children's Cancer Study Group (UKCCSG) retrospective review.

Survival in childhood lymphoproliferative disease (LPD) remains poor, particularly in non-transplant patients. The anti-CD20 antibody rituximab shows promise but data in children is scant. A retrospective study of 22 (aged 11 months to 18 years) children treated with rituximab is presented. Two had primary immunodeficiency, two had prolonged immunosuppression and 18 had post-transplant LPD (eight bone marrow, five liver, four heart, one kidney). Nine patients had multi-organ involvement and 13 single site disease. Seventeen out of 22 had rituximab alone. In 16, a dose of 375 mg/m2 i.v. weekly was used (less in one patient due to renal dysfunction). Twelve patients received four courses and ten patients received one to three courses. Fever was the main side-effect in four. Eight (47%) had single agent response; four complete and four partial. All had other treatment prior to rituximab. Median follow-up was 35 months (range 22 - 47 months). In childhood LPD unresponsive to standard treatment, rituximab showed single agent response and requires further evaluation.

Two novel activating mutations in the Wiskott-Aldrich syndrome protein result in congenital neutropenia.

Severe congenital neutropenia (SCN) is characterized by neutropenia, recurrent bacterial infections, and maturation arrest in the bone marrow. Although many cases have mutations in the ELA2 gene encoding neutrophil elastase, a significant proportion remain undefined at a molecular level. A mutation (Leu270Pro) in the gene encoding the Wiskott-Aldrich syndrome protein (WASp) resulting in an X-linked SCN kindred has been reported. We therefore screened the WAS gene in 14 young SCN males with wild-type ELA2 and identified 2 with novel mutations, one who presented with myelodysplasia (Ile294Thr) and the other with classic SCN (Ser270Pro). Both patients had defects of immunologic function including a generalized reduction of lymphoid and natural killer cell numbers, reduced lymphocyte proliferation, and abrogated phagocyte activity. In vitro culture of bone marrow progenitors demonstrated a profound reduction in neutrophil production and increased levels of apoptosis, consistent with an intrinsic disturbance of normal myeloid differentiation as the cause of the neutropenia. Both mutations resulted in increased WASp activity and produced marked abnormalities of cytoskeletal structure and dynamics. Furthermore, these results also suggest a novel cause of myelodysplasia and that male children with myelodysplasia and disturbance of immunologic function should be screened for such mutations.

Complex genomic alterations and gene expression in acute lymphoblastic leukemia with intrachromosomal amplification of chromosome 21.

We have previously identified a unique subtype of acute lymphoblastic leukemia (ALL) associated with a poor outcome and characterized by intrachromosomal amplification of chromosome 21 including the RUNX1 gene (iAMP21). In this study, array-based comparative genomic hybridization (aCGH) (n = 10) detected a common region of amplification (CRA) between 33.192 and 39.796 Mb and a common region of deletion (CRD) between 43.7 and 47 Mb in 100% and 70% of iAMP21 patients, respectively. High-resolution genotypic analysis (n = 3) identified allelic imbalances in the CRA. Supervised gene expression analysis showed a distinct signature for eight patients with iAMP21, with 10% of overexpressed genes located within the CRA. The mean expression of these genes was significantly higher in iAMP21 when compared to other ALL samples (n = 45). Although genomic copy number correlated with overall gene expression levels within areas of loss or gain, there was considerable individual variation. A unique subset of differentially expressed genes, outside the CRA and CRD, were identified when gene expression signatures of iAMP21 were compared to ALL samples with ETV6-RUNX1 fusion (n = 21) or high hyperdiploidy with additional chromosomes 21 (n = 23). From this analysis, LGMN was shown to be overexpressed in patients with iAMP21 (P = 0.0012). Genomic and expression data has further characterized this ALL subtype, demonstrating high levels of 21q instability in these patients leading to proposals for mechanisms underlying this clinical phenotype and plausible alternative treatments.

Treatment for myeloid leukaemia of Down syndrome: population-based experience in the UK and results from the Medical Research Council AML 10 and AML 12 trials.

Down syndrome (DS) children are at an increased risk of developing myelodysplasia and acute myeloid leukaemia (AML). We retrospectively analysed the population-based data on 81 children with myeloid leukaemia of Down syndrome (ML-DS) from the UK National Registry of Childhood Tumours and experience in the Medical Research Council (MRC) AML 10 and AML 12 trials, which enrolled 46 children with ML-DS from 1988 to 2002. Eight per cent of UK children with AML had DS, but DS children comprised only 5% of children registered in MRC trials. The unique clinical characteristics of ML-DS were confirmed. Overall survival (OS) of ML-DS at 5 years increased from 47% in UK children diagnosed from 1988 to 1995 to 75% in children diagnosed from 1996 to 2002. OS for DS children registered in AML 10 and AML 12 was 74% in 5 years and improved from AML 10 to AML 12 (56% vs. 83%) There was no significant difference in OS between DS and non-DS children (OS: 74% vs. 62%, P = 0.4) in the trials, but this result masked a significant increase in early death amongst DS children, with a significant reduction in mortality later on. Relapse was significantly reduced (3% vs. 39%, P = 0.0003), leading to the improved disease-free survival (83% vs. 56%, P = 0.02). Given the increased number of early treatment-related deaths, future treatment protocols should aim to reduce chemotherapy dosage or intensity whilst maintaining low rates of resistant and recurrent disease.

Adolescents with acute lymphoblastic leukaemia: emerging from the shadow of paediatric and adult treatment protocols.

Adolescents and young adults (AYA) with acute lymphoblastic leukaemia (ALL) constitute a distinct population from children and older adults. Based on patterns of referral, they may be treated by either paediatric or adult oncologists. As a group, AYA with ALL have a worse survival and event-free survival (EFS) compared to that achieved by younger children. A systematic review of all published clinical trials, which provide data on treatment and outcome of adolescents with ALL, has been summarised in an effort to determine whether they should be treated on paediatric or adult type protocols. Adolescents appear to have a consistent survival advantage when treated on paediatric regimens.

Prospective gene expression analysis accurately subtypes acute leukaemia in children and establishes a commonality between hyperdiploidy and t(12;21) in acute lymphoblastic leukaemia.

We have prospectively analysed and correlated the gene expression profiles of children presenting with acute leukaemia to the Royal London and Great Ormond Street Hospitals with morphological diagnosis, immunophenotype and karyotype. Total RNA extracted from freshly sorted blast cells was obtained from 84 lymphoblastic [acute lymphoblastic leukaemia (ALL)], 20 myeloid [acute myeloid leukaemia (AML)] and three unclassified acute leukaemias and hybridised to the high density Affymetrix U133A oligonucleotide array. Analysis of variance and significance analysis of microarrays was used to identify discriminatory genes. A novel 50-gene set accurately identified all patients with ALL and AML and predicted for a diagnosis of AML in three patients with unclassified acute leukaemia. A unique gene set was derived for each of eight subtypes of acute leukaemia within our data set. A common profile for children with ALL with an ETV6-RUNX1 fusion, amplification or deletion of ETV6, amplification of RUNX1 or hyperdiploidy with an additional chromosome 21 was identified. This suggests that these rearrangements share a commonality in biological pathways that maintains the leukaemic state. The gene TERF2 was most highly expressed in this group of patients. Our analyses demonstrate that not only is microarray analysis the single most effective tool for the diagnosis of acute leukaemias of childhood but it has the ability to identify unique biological pathways. To further evaluate its prognostic value it needs to be incorporated into the routine diagnostic analysis for large-scale clinical trials in childhood acute leukaemias.

Outcome after first relapse in childhood acute lymphoblastic leukaemia - lessons from the United Kingdom R2 trial.

A retrospective analysis of children with first relapse of acute lymphoblastic leukaemia (ALL), treated on the UKALL R2 protocol at four different hospitals, between June 1995 and December 2002 was performed. Of the 150 children 139 (93%) achieved a second complete remission. The overall survival (OS) and event-free survival (EFS) for the whole group was 56% and 47% respectively. The duration of first complete remission and immunophenotype, but not sites of relapse, were predictive for survival. Using the Berlin-Frankfürt-Münster risk stratification for relapsed ALL, the OS and EFS for standard, intermediate (IR) and high risk (HR) groups were 92% and 92%, 64% and 51%, and 14% and 15%, respectively; P < 0.0001 for both OS and EFS. In the IR group, those with a very early isolated central nervous system relapse also had a significantly worse outcome (P = 0.0001). Given the poor outcome of a second relapse, clear strategies are required to identify those in the IR group who will most benefit from stem cell transplantation (SCT). A higher proportion (16%) of induction failures in the HR group suggest the need for novel agents during this phase of treatment, but SCT was associated with a lower relapse rate and better outcome than those treated with chemotherapy alone.

Red cell pyruvate kinase deficiency: 17 new mutations of the PK-LR gene.

The PK-LR gene was studied in 23 patients with congenital haemolytic anaemia associated with erythrocyte pyruvate kinase (PK) deficiency. Twenty-seven different mutations were detected among the 42 mutated alleles identified: 19 missense mutations, four splice site mutations and one nonsense, one single base deletion and two large deletions. Seventeen of them (107G, 278T, 403T, 409A, 661A, 859C, 958A, 1094T, 1190T, 1209A, 1232C, 1369G, 507A, IVS9 -1c, IVS9 +43c [corrected] del C224, del 5006bp IVS3--> nt 1431) were new. Although all the exons, the flanking regions and the promoter were sequenced in all cases, we failed to detect the second expected mutation in four subjects. To correlate genotype to phenotype, the molecular results were related to the biochemical properties of the mutant enzymes by an analysis of the three-dimensional structure of erythrocyte PK. The new mutant 409A, found in association with the large deletion of 5006 bp in a newborn baby who died soon after birth, was functionally characterized by mutagenesis and in vitro expression of the protein to investigate its contribution in the severity of the clinical pattern. However, the biochemical data obtained for the mutant enzyme cannot explain the severe anaemia found in the PK-deficient patient hemizygous for this mutation.