Population Pharmacokinetics and Pharmacodynamics of Ciprofloxacin Prophylaxis in Pediatric Acute Lymphoblastic Leukemia Patients.

BACKGROUND: Ciprofloxacin is used as antimicrobial prophylaxis in pediatric acute lymphoblastic leukemia (ALL) to decrease infections with gram-negative bacteria. However, there are no clear guidelines concerning prophylactic dose. AIMS: To determine the pharmacokinetics and pharmacodynamics (PKPD) of ciprofloxacin prophylaxis in a pediatric ALL population. The effect of patient characteristics and antileukemic treatment on ciprofloxacin exposure, the area under the concentration time curve over minimal inhibitory concentration (AUC24/MIC) ratios, and emergence of resistance were studied. METHODS: A total of 615 samples from 129 children (0-18 years) with ALL were collected in a multicenter prospective study. A population pharmacokinetic model was developed. Microbiological cultures were collected prior to and during prophylaxis. An AUC24/MIC of ≥125 was defined as target ratio. RESULTS: A 1-compartment model with zero-order absorption and allometric scaling best described the data. No significant (P < .01) covariates remained after backward elimination and no effect of asparaginase or azoles were found. Ciprofloxacin AUC24 was 16.9 mg*h/L in the prednisone prophase versus 29.3 mg*h/L with concomitant chemotherapy. Overall, 100%, 81%, and 18% of patients at, respectively, MIC of 0.063, 0.125, and 0.25 mg/L achieved AUC24/MIC ≥ 125. In 13% of the patients, resistant bacteria were found during prophylactic treatment. CONCLUSION: Ciprofloxacin exposure shows an almost 2-fold change throughout the treatment of pediatric ALL. Depending on the appropriateness of 125 as target ratio, therapeutic drug monitoring or dose adjustments might be indicated for less susceptible bacteria starting from ≥ 0.125 mg/L to prevent the emergence of resistance and reach required targets for efficacy.

Pyuria is absent during urinary tract infections in neutropenic patients.

To investigate the diagnostic significance of a normal urine sediment in the work-up for fever of unknown origin in neutropenia. Urinary tract infection was defined as ≥10(5) urinary pathogens in the absence of another focus. Pyuria was found in only 1/23 neutropenic episodes compared to 21/31 in controls (P < 0.0001).

Measurable residual disease in pediatric acute myeloid leukemia: a systematic review.

Introduction: A systematic review was performed to assess the prognostic value of Measurable Residual Disease (MRD) during treatment, for relapse and overall survival in pediatric acute myeloid leukemia (AML).Areas covered: A systematic search of available literature was performed to identify original full-text articles concerning MRD as prognostic for relapse and survival in pediatric AML. Thirteen studies were included, and in all studies, MRD positivity during treatment was associated with worse clinical outcome. MRD positivity was significantly associated with a higher probability of relapse in eleven studies. However, MRD negativity does not exclude the possibility of relapse in pediatric AML, while positivity early during therapy does not exclude cure.Expert opinion: MRD positivity during treatment has emerged as the most powerful prognostic factor in pediatric AML concerning relapse and overall survival and is useful for risk-group adapted treatment. Future studies should identify the optimal time-point(s) for MRD measurements and the optimal technique, to further improve its prognostic significance.

Safety and efficacy of a maternal vaccine for the passive protection of broiler chicks against necrotic enteritis.

Necrotic enteritis is a potentially fatal multifactorial disease of chickens, which under commercial conditions is often associated with increased levels of mortality and reduced bird performance. The safety and efficacy of a Clostridium perfringens type A alpha-toxoid (Netvax™) formulated as an oil emulsion was investigated, following maternal immunization of broiler breeder hens, housed under commercial conditions, by the intramuscular route. A total of 11,234 hens were vaccinated across two integrated poultry sites. The vaccine was safe with no systemic reactions or adverse effects on bird performance detected. Vaccination resulted in a significant increase in anti-alpha toxin antibody in the hen that was maintained throughout the study, and subsequently transferred to their progeny throughout the laying period via egg yolk. Chicks hatched from eggs produced from vaccinated hens were shown to have reduced mortality specifically related to progeny flocks where gross gut lesions associated with necrotic enteritis were observed in control chicks. Further, whilst C. perfringens was isolated from control chicks with necrotic enteritis lesions, no such isolations were made at these time points from chicks from vaccinated hens. These results indicate that, under commercial conditions, maternal vaccination with Netvax™ can help to control losses related to necrotic enteritis.

Improved survival for children and young adolescents with acute myeloid leukemia: a Dutch study on incidence, survival and mortality.

Variation in survival of pediatric acute myeloid leukemia (pAML) over time and between Western European countries exists. The aim of the current study is to assess the progress made for the Dutch pAML population (0-17 years) during 1990-2015, based on trends in incidence, survival and mortality. Data from the population-based Netherlands Cancer Registry were merged with leukemia-related characteristics and treatment specifics from the Dutch Childhood Leukemia Study Group (Dutch Childhood Oncology Group (DCOG) from 2002 onwards). Mortality data (1980-2016) were obtained from the cause of death registry of Statistics Netherlands. Trend analyses were performed over time and by treatment protocol. Between 1990 and 2015, a total of 635 children aged 0-17 years were diagnosed with AML for an average of 25 patients (range 18-36) per year. There was a slight increase in the incidence at age 1-4 years (average annual percentage change (AAPC) of +2.2% per year (95% CI 0.8-3.5, p < 0.01)). Overall, the 5-year survival significantly improved over the past 26 years and nearly doubled from 40% in the early 1990s to 74% in 2010-2015. Multivariable analysis showed a 49% reduction in risk of death for pAML patients treated according to the latest DB-AML 01 protocol (p = 0.03). The continuing decrease of mortality (AAPC -2.8% per year (95% CI -4.1 to -1.5)) supports the conclusion of true progress against pAML in the Netherlands.

RAS pathway mutations as a predictive biomarker for treatment adaptation in pediatric B-cell precursor acute lymphoblastic leukemia.

RAS pathway mutations have been linked to relapse and chemotherapy resistance in pediatric B-cell precursor acute lymphoblastic leukemia (BCP-ALL). However, comprehensive data on the frequency and prognostic value of subclonal mutations in well-defined subgroups using highly sensitive and quantitative methods are lacking. Targeted deep sequencing of 13 RAS pathway genes was performed in 461 pediatric BCP-ALL cases at initial diagnosis and in 19 diagnosis-relapse pairs. Mutations were present in 44.2% of patients, with 24.1% carrying a clonal mutation. Mutation frequencies were highest in high hyperdiploid, infant t(4;11)-rearranged, BCR-ABL1-like and B-other cases (50-70%), whereas mutations were less frequent in ETV6-RUNX1-rearranged, and rare in TCF3-PBX1- and BCR-ABL1-rearranged cases (27-4%). RAS pathway-mutated cells were more resistant to prednisolone and vincristine ex vivo. Clonal, but not subclonal, mutations were linked to unfavorable outcome in standard- and high-risk-treated patients. At relapse, most RAS pathway mutations were clonal (9 of 10). RAS mutant cells were sensitive to the MEK inhibitor trametinib ex vivo, and trametinib sensitized resistant cells to prednisolone. We conclude that RAS pathway mutations are frequent, and that clonal, but not subclonal, mutations are associated with unfavorable risk parameters in newly diagnosed pediatric BCP-ALL. These mutations may designate patients eligible for MEK inhibitor treatment.

MN1 overexpression is driven by loss of DNMT3B methylation activity in inv(16) pediatric AML.

In acute myeloid leukemia (AML), specific genomic aberrations induce aberrant methylation, thus directly influencing the transcriptional programing of leukemic cells. Therefore, therapies targeting epigenetic processes are advocated as a promising therapeutic tool for AML treatment. However, to develop new therapies, a comprehensive understanding of the mechanism(s) driving the epigenetic changes as a result of acquired genetic abnormalities is necessary. This understanding is still lacking. In this study, we performed genome-wide CpG-island methylation profiling on pediatric AML samples. Six differentially methylated genomic regions within two genes, discriminating inv(16)(p13;q22) from non-inv(16) pediatric AML samples, were identified. All six regions had a hypomethylated phenotype in inv(16) AML samples, and this was most prominent at the regions encompassing the meningioma (disrupted in balanced translocation) 1 (MN1) oncogene. MN1 expression primarily correlated with the methylation level of the 3' end of the MN1 exon-1 locus. Decitabine treatment of different cell lines showed that induced loss of methylation at the MN1 locus can result in an increase of MN1 expression, indicating that MN1 expression is coregulated by DNA methylation. To investigate this methylation-associated mechanism, we determined the expression of DNA methyltransferases in inv(16) AML. We found that DNMT3B expression was significantly lower in inv(16) samples. Furthermore, DNMT3B expression correlated negatively with MN1 expression in pediatric AML samples. Importantly, depletion of DNMT3B impaired remethylation efficiency of the MN1 exon-1 locus in AML cells after decitabine exposure. These findings identify DNMT3B as an important coregulator of MN1 methylation. Taken together, this study shows that the methylation level of the MN1 exon-1 locus regulates MN1 expression levels in inv(16) pediatric AML. This methylation level is dependent on DNMT3B, thus suggesting a role for DNMT3B in leukemogenesis in inv(16) AML, through MN1 methylation regulation.

miR-139-5p controls translation in myeloid leukemia through EIF4G2.

MicroRNAs (miRNAs) are crucial components of homeostatic and developmental gene regulation. In turn, dysregulation of miRNA expression is a common feature of different types of cancer, which can be harnessed therapeutically. Here we identify miR-139-5p suppression across several cytogenetically defined acute myeloid leukemia (AML) subgroups. The promoter of mir-139 was transcriptionally silenced and could be reactivated by histone deacetylase inhibitors in a dose-dependent manner. Restoration of mir-139 expression in cell lines representing the major AML subgroups (t[8;21], inv[16], mixed lineage leukemia-rearranged and complex karyotype AML) caused cell cycle arrest and apoptosis in vitro and in xenograft mouse models in vivo. During normal hematopoiesis, mir-139 is exclusively expressed in terminally differentiated neutrophils and macrophages. Ectopic expression of mir-139 repressed proliferation of normal CD34(+)-hematopoietic stem and progenitor cells and perturbed myelomonocytic in vitro differentiation. Mechanistically, mir-139 exerts its effects by repressing the translation initiation factor EIF4G2, thereby reducing overall protein synthesis while specifically inducing the translation of cell cycle inhibitor p27(Kip1). Knockdown of EIF4G2 recapitulated the effects of mir-139, whereas restoring EIF4G2 expression rescued the mir-139 phenotype. Moreover, elevated miR-139-5p expression is associated with a favorable outcome in a cohort of 165 pediatric patients with AML. Thus, mir-139 acts as a global tumor suppressor-miR in AML by controlling protein translation. As AML cells are dependent on high protein synthesis rates controlling the expression of mir-139 constitutes a novel path for the treatment of AML.

Quantification of minimal residual disease in children with oligoclonal B-precursor acute lymphoblastic leukemia indicates that the clones that grow out during relapse already have the slowest rate of reduction during induction therapy.

Antigen receptor gene rearrangements are applied for the PCR-based minimal residual disease (MRD) detection in acute lymphoblastic leukemia (ALL). It is known that ongoing rearrangements result in subclone formation, and that the relapsing subclone(s) can contain antigen receptor rearrangement(s) that differ from the rearrangements found in the major clone(s) at diagnosis. However, the mechanism leading to this so-called clonal evolution is not known, particularly at which time point in the disease the relapsing subclone obtains its (relative) therapy resistance. To obtain insight in clonal evolution, we followed the kinetics of several subclones in three oligoclonal ALL patients during induction therapy. Clone-specific nested PCR for immunoglobulin heavy chain or T cell receptor delta gene rearrangements were performed in limiting dilution assays on bone marrow samples taken at diagnosis, at the end of induction therapy and at possible relapse in three children with oligoclonal B-precursor ALL. We demonstrated that in all three patients the subclones were behaving differently in response to therapy. Moreover, in the two patients who relapsed, the clones that grew out during relapse showed the slowest regression or even evoluated during induction therapy and the clones that were not present at relapse showed good response to induction therapy. These results support the hypothesis that at least in some patients already at diagnosis or in the very first weeks, subclones have important differences in respect to resistance. Hence, these data give experimental evidence for the need to develop, during the first months after diagnosis, quantitative PCR assays for at least two different Ig/TCR gene rearrangement targets for every ALL patient.

Real-time quantitative PCR for the detection of minimal residual disease in acute lymphoblastic leukemia using junctional region specific TaqMan probes.

Analysis of minimal residual disease (MRD) can predict outcome in acute lymphoblastic leukemia (ALL). A large prospective study in childhood ALL has shown that MRD analysis using immunoglobulin (Ig) and T cell receptor (TCR) gene rearrangements as PCR targets can identify good and poor prognosis groups of substantial size that might profit from treatment adaptation. This MRD-based risk group assignment was based on the kinetics of tumor reduction. Consequently, the level of MRD has to be defined precisely in follow-up samples. However, current PCR methods do not allow easy and accurate quantification. We have tested 'real-time' quantitative PCR (RQ-PCR) using the TaqMan technology and compared its sensitivity with two conventional MRD-PCR methods, ie dot-blot and liquid hybridization of PCR amplified Ig/TCR gene rearrangements using clone-specific radioactive probes. In RQ-PCR the generated specific PCR product is measured at each cycle ('real-time') by cleavage of a fluorogenic intrinsic TaqMan probe. The junctional regions of rearranged Ig/TCR genes define the specificity and sensitivity of PCR-based MRD detection in ALL and are generally used to design a patient-specific probe. In the TaqMan technology we have chosen for the same approach with the design of patient-specific TaqMan probes at the position of the junctional regions. We developed primers/probe combinations for RQ-PCR analysis of a total of three IGH, two TCRD, two TCRG and three IGK gene rearrangements in four randomly chosen precursor-B-ALL. In one patient, 12 bone marrow follow-up samples were analyzed for the presence of MRD using an IGK PCR target. The sensitivity of the RQ-PCR technique appeared to be comparable to the dot-blot method, but less sensitive than liquid hybridization. Although it still is a relatively expensive method, RQ-PCR allows sensitive, reproducible and quantitative MRD detection with a high throughput of samples providing possibilities for semi-automation. We consider this novel technique as an important step forward towards routinely performed diagnostic MRD studies.

Despite differential gene expression profiles pediatric MDS derived mesenchymal stromal cells display functionality in vitro.

Pediatric myelodysplastic syndrome (MDS) is a heterogeneous disease covering a spectrum ranging from aplasia (RCC) to myeloproliferation (RAEB(t)). In adult-type MDS there is increasing evidence for abnormal function of the bone-marrow microenvironment. Here, we extensively studied the mesenchymal stromal cells (MSCs) derived from children with MDS. MSCs were expanded from the bone-marrow of 17 MDS patients (RCC: n=10 and advanced MDS: n=7) and pediatric controls (n=10). No differences were observed with respect to phenotype, differentiation capacity, immunomodulatory capacity or hematopoietic support. mRNA expression analysis by Deep-SAGE revealed increased IL-6 expression in RCC- and RAEB(t)-MDS. RCC-MDS MSC expressed increased levels of DKK3, a protein associated with decreased apoptosis. RAEB(t)-MDS revealed increased CRLF1 and decreased DAPK1 expressions. This pattern has been associated with transformation in hematopoietic malignancies. Genes reported to be differentially expressed in adult MDS-MSC did not differ between MSC of pediatric MDS and controls. An altered mRNA expression profile, associated with cell survival and malignant transformation, of MSC derived from children with MDS strengthens the hypothesis that the micro-environment is of importance in this disease. Our data support the understanding that pediatric and adult MDS are two different diseases. Further evaluation of the pathways involved might reveal additional therapy targets.

Progesterone therapy in mares with abnormal oestrous cycles.

In two studies in Holland and France a total of 41 mares found either to be in anoestrus or showing continuous oestrous behaviour during the early part of the covering season were treated with a progesterone releasing intravaginal device. Of the 32 mares in the first study, 24 ovulated after treatment and 11 became pregnant to a covering at the induced oestrus. In the second study all but one of the nine mares showed oestrus, seven ovulated and six became pregnant at the post treatment oestrus. By the end of the covering season 15 of the 21 mares treated for anoestrus and seven of the 11 mares treated for persistent oestrus were found to be in foal.

Treatment of keratoconjunctivitis sicca in dogs with cyclosporine ophthalmic ointment: a European clinical field trial.

The results are reported of a six-week clinical trial of the efficacy of 0.2 per cent cyclosporine ophthalmic ointment for the treatment of chronic idiopathic keratoconjunctivitis sicca in dogs in the United Kingdom, Germany and France. The 87 dogs were referral cases with a history of chronic unresponsive keratoconjunctivitis sicca of which the aetiology was unknown, and they had to meet stringent criteria before being included in the trial. The clinical response to the therapy was monitored after seven, 21 and 42 days and the results for the right and left eyes were analysed separately. There was a statistically significant increase in lacrimal secretion throughout the study, with most of the increase occurring during the first week of treatment. The percentage of eyes with improved lacrimal secretion was higher in the dogs with initial Schirmer tear test values > or = 2 mm/min than in those with initial values of 0 or 1 mm/min. The observed steady improvement in conjunctival health was not always correlated with an improvement in lacrimal secretion. The incidence of blepharospasm, other signs of discomfort and corneal oedema decreased significantly during the study. No improvement in corneal vascularisation or pigmentation was observed during the six-week trial. Overall, 76 per cent of the left eyes and 87 per cent of the right eyes were considered to have improved at the end of the treatment period. No serious adverse reactions were observed and only mild irritation was noticed by the owners immediately after the application of the ointment. This irritation resulted in the recording of an adverse reaction at the scheduled observations in only three cases.

Treatment of naturally occurring bovine respiratory disease in juvenile calves with a single administration of a florfenicol plus flunixin meglumine formulation.

The efficacy and safety of a florfenicol plus flunixin meglumine formulation in the treatment of respiratory disease was evaluated in calves less than six weeks of age, compared with a positive control group treated with a well-established florfenicol formulation. A total of 210 calves, selected from nine sites in Belgium, France and Spain, showing severe signs of respiratory disease, were randomly assigned to treatment with either florfenicol plus flunixin meglumine (Resflor; MSD Animal Health) or florfenicol (Nuflor; MSD Animal Health), both administered subcutaneously once. Animals were clinically observed daily for 10 days following treatment initiation. The predominant respiratory pathogens were Pasteurella multocida, Mycoplasma bovis, Mannheimia haemolytica and Histophilus somni. All isolates were subject to in vitro sensitivity testing and found susceptible to florfenicol. In both groups, rectal temperature dropped and clinical index (depression and respiratory signs) significantly improved after treatment. Specifically, for the change in rectal temperature from pretreatment to six hours post-treatment, the florfenicol-flunixin formulation was found significantly superior to florfenicol. Moreover, the florfenicol-flunixin formulation alleviated the clinical signs of disease more rapidly, and was demonstrated to be non-inferior to florfenicol on days 4 and 10. The use of the product combining florfenicol and flunixin in calves is safe and efficacious in the treatment of outbreaks of bovine respiratory disease.

Insights in dynamic kinome reprogramming as a consequence of MEK inhibition in MLL-rearranged AML.

Single kinase-targeted cancer therapies often failed prolonged responses because cancer cells bypass through alternative routes. In this study, high-throughput kinomic and proteomic approaches enabled to identify aberrant activity profiles in mixed lineage leukemia (MLL)-rearranged acute myeloid leukemia (AML) that defined druggable targets. This approach revealed impaired activity of proteins belonging to the mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K) pathway. Pharmacological druggable MAPK pathway targets tested in primary MLL-rearranged AML included MAPKK1/2 (MEK), cyclic AMP-responsive element-binding protein (CREB) and MAPK8/9 (JNK). MEK inhibition showed to severely decrease MLL-rearranged AML cell survival without showing cytotoxicity in normal controls, whereas inhibition of CREB and JNK failed to exhibit MLL selectivity. Exploring the working mechanism of MEK inhibition, we assessed proteome activity in response to MEK inhibition in THP-1. MAPK1/3 (Erk) phosphorylation was instantly decreased in concurrence with a sustained Akt/mammalian target of rapamycin (mTOR) phosphorylation that enabled a subpopulation of cells to survive MEK inhibition. After exhaustion of MEK inhibition the AML cells recovered via increased activity of vascular endothelial growth factor receptor-2 (VEGFR-2) and Erk proteins to resume their proliferative state. Combined MEK and VEGFR-2 inhibition strengthened the reduction in MLL-rearranged AML cell survival by blocking the Akt/mTOR and MAPK pathways simultaneously. The generation of insights in cancerous altered activity profiles and alternative escape mechanisms upon targeted therapy allows the rational design of novel combination strategies.

miR-9 is a tumor suppressor in pediatric AML with t(8;21).

MicroRNAs (miRNAs) play a pivotal role in the regulation of hematopoiesis and development of leukemia. Great interest emerged in modulating miRNA expression for therapeutic purposes. In order to identify miRNAs, which specifically suppress leukemic growth of acute myeloid leukemia (AML) with t(8;21), inv(16) or mixed lineage leukemia (MLL) rearrangement by inducing differentiation, we conducted a miRNA expression profiling in a cohort of 90 cytogenetically characterized, de novo pediatric AML cases. Four miRNAs, specifically downregulated in MLL-rearranged, t(8;21) or inv(16) AMLs, were characterized by their tumor-suppressive properties in cell lines representing those respective cytogenetic groups. Among those, forced expression of miR-9 reduced leukemic growth and induced monocytic differentiation of t(8;21) AML cell lines in vitro and in vivo. The tumor-suppressive functions of miR-9 were specifically restricted to AML cell lines and primary leukemic blasts with t(8;21). On the other hand, these functions were not evident in AML blasts from patients with MLL rearrangements. We showed that miR-9 exerts its effects through the cooperation with let-7 to repress the oncogenic LIN28B/HMGA2 axis. Thus, miR-9 is a tumor suppressor-miR which acts in a stringent cell context-dependent manner.

Improved flow cytometric detection of minimal residual disease in childhood acute lymphoblastic leukemia.

Most current treatment protocols for acute lymphoblastic leukemia (ALL) include minimal residual disease (MRD) diagnostics, generally based on PCR analysis of rearranged antigen receptor genes. Although flow cytometry (FCM) can be used for MRD detection as well, discordant FCM and PCR results are obtained in 5-20% of samples. We evaluated whether 6-color FCM, including additional markers and new marker combinations, improved the results. Bone marrow samples were obtained from 363 ALL patients at day 15, 33 and 78 and MRD was analyzed using 6-color (218 patients) or 4-color (145 patients) FCM in parallel to routine PCR-based MRD diagnostics. Compared with 4-color FCM, 6-color FCM significantly improved the concordance with PCR-based MRD data (88% versus 96%); particularly the specificity of the MRD analysis improved. However, PCR remained more sensitive at levels <0.01%. MRD-based risk groups were similar between 6-color FCM and PCR in 68% of patients, most discrepancies being medium risk by PCR and standard risk by FCM. Alternative interpretation of the PCR data, aimed at prevention of false-positive MRD results, changed the risk group to standard risk in half (52%) of these discordant cases. In conclusion, 6-color FCM significantly improves MRD analysis in ALL but remains less sensitive than PCR-based MRD-diagnostics.

Aurora kinases in childhood acute leukemia: the promise of aurora B as therapeutic target.

We investigated the effects of targeting the mitotic regulators aurora kinase A and B in pediatric acute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML). Aurora protein expression levels in pediatric ALL and AML patient samples were determined by western blot and reverse phase protein array. Both kinases were overexpressed in ALL and AML patients (P<0.0002), especially in E2A-PBX1-translocated ALL cases (P<0.002), compared with normal bone-marrow mononuclear cells. Aurora kinase expression was silenced in leukemic cell lines using short hairpin RNAs and locked nucleic acid-based mRNA antagonists. Aurora B knockdown resulted in proliferation arrest and apoptosis, whereas aurora A knockdown caused no or only minor growth delay. Most tested cell lines were highly sensitive to the AURKB-selective inhibitor barasertib-hydroxyquinazoline-pyrazol-anilide (AZD1152-HQPA) in the nanomolar range, as tested with an MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) assay. But most importantly, primary ALL cells with a high aurora B protein expression, especially E2A-PBX1-positive cases, were sensitive as well. In adult AML early clinical trials, clear responses are observed with barasertib. Here we show that inhibition of aurora B, more than aurora A, has an antiproliferative and pro-apoptotic effect on acute leukemia cells, indicating that particularly targeting aurora B may offer a new strategy to treat pediatric ALL and AML.

High-risk childhood acute lymphoblastic leukemia in first remission treated with novel intensive chemotherapy and allogeneic transplantation.

Children with acute lymphoblastic leukemia (ALL) and high minimal residual disease (MRD) levels after initial chemotherapy have a poor clinical outcome. In this prospective, single arm, Phase 2 trial, 111 Dutch and Australian children aged 1-18 years with newly diagnosed, t(9;22)-negative ALL, were identified among 1041 consecutively enrolled patients as high risk (HR) based on clinical features or high MRD. The HR cohort received the AIEOP-BFM (Associazione Italiana di Ematologia ed Oncologia Pediatrica (Italy)-Berlin-Frankfurt-Münster ALL Study Group) 2000 ALL Protocol I, then three novel HR chemotherapy blocks, followed by allogeneic transplant or chemotherapy. Of the 111 HR patients, 91 began HR treatment blocks, while 79 completed the protocol. There were 3 remission failures, 12 relapses, 7 toxic deaths in remission and 10 patients who changed protocol due to toxicity or clinician/parent preference. For the 111 HR patients, 5-year event-free survival (EFS) was 66.8% (±5.5) and overall survival (OS) was 75.6% (±4.3). The 30 patients treated as HR solely on the basis of high MRD levels had a 5-year EFS of 63% (±9.4%). All patients experienced grade 3 or 4 toxicities during HR block therapy. Although cure rates were improved compared with previous studies, high treatment toxicity suggested that novel agents are needed to achieve further improvement.

Outcome in children with Down's syndrome and acute lymphoblastic leukemia: role of IKZF1 deletions and CRLF2 aberrations.

Children with Down's syndrome (DS) have an increased risk of developing acute lymphoblastic leukemia (ALL) and have a low frequency of established genetic aberrations. We aimed to determine which genetic abnormalities are involved in DS ALL. We studied the frequency and prognostic value of deletions in B-cell development genes and aberrations of janus kinase 2 (JAK2) and cytokine receptor-like factor 2 (CRLF2) using array-comparative genomic hybridization, and multiplex ligation-dependent probe amplification in a population-based cohort of 34 Dutch Childhood Oncology Group DS ALL samples. A population-based cohort of 88 DS samples from the UK trials was used to validate survival estimates for IKZF1 and CRLF2 abnormalities. In total, 50% of DS ALL patients had ≥1 deletion in the B-cell development genes: PAX5 (12%), VPREB1 (18%) and IKZF1 (35%). JAK2 was mutated in 15% of patients, genomic CRLF2 rearrangements in 62%. Outcome was significantly worse in patients with IKZF1 deletions (6-year event-free survival (EFS) 45 ± 16% vs 95 ± 4%; P=0.002), which was confirmed in the validation cohort (6-year EFS 21 ± 12% vs 58 ± 11%; P=0.002). This IKZF1 deletion was a strong independent predictor for outcome (hazard ratio EFS 3.05; P=0.001). Neither CRLF2 nor JAK2 were predictors for worse prognosis. If confirmed in prospective series, IKZF1 deletions may be used for risk-group stratification in DS ALL.