Pieters, R.

Endocrine sequelae and metabolic syndrome in adult long-term survivors of childhood acute myeloid leukemia (Article)

2013-04-01T00:00:00Z

This study focuses on the effect of chemotherapy on endocrinopathies and the metabolic syndrome in adult survivors of childhood acute myeloid leukemia (AML). Endocrine function and metabolic syndrome were evaluated in 12 AML survivors, treated with chemotherapy, and in 9 survivors of myeloid leukemias treated with stem cell transplantation (SCT), after a median follow-up time of 20 years (range 9-31). In survivors treated with chemotherapy, no endocrinopathies or metabolic syndrome were present, although AMH and Inhibin B levels tended to be lower than in controls. In SCT survivors, pituitary deficiencies and metabolic syndrome were more frequent.

Motor performance and functional exercise capacity in survivors of pediatric acute lymphoblastic leukemia (Article)

2013-03-01T00:00:00Z

Background: Impaired motor performance and reduced maximum exercise capacity during and after treatment of acute lymphoblastic leukemia (ALL) has been shown. However, no longitudinal study monitoring motor performance after cessation of treatment has been published. Whether sub-maximal exercise capacity is reduced is unknown. Procedure: Motor performance of pediatric ALL survivors, treated with Dutch Childhood Oncology Group ALL-9 protocol was measured with the movement-ABC at stop treatment and ≥5 years later. At follow-up functional exercise capacity was also investigated using the 6-minute walk test (6MWT). Heart rate and oxygen saturation were measured with a portable pulse oximeter before and after the 6MWT. Results: Nineteen boys and 15 girls, median age 12.3 years (range: 9.0-18.7), median time since completion of chemotherapy 5.2 years (5.0-7.1), participated. Mean height/age and weight/age were within the norm, whereas mean BMI/age was significantly increased (mean SDS 0.38, SEM 0.17, P=0.04). Motor performance had improved significantly (P=0.001). In contrast, functional exercise capacity at follow-up was significantly impaired (mean SDS -2.05, SEM 0.13, P<0.001). Conclusions: At ≥5 years after completion of ALL treatment motor performance had improved significantly, but functional exercise capacity was significantly impaired. The exact underlying cause of this late effect needs further study.

Improved flow cytometric detection of minimal residual disease in childhood acute lymphoblastic leukemia (Article)

2013-03-01T00:00:00Z

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.

Telomere length and telomerase complex mutations in pediatric acute myeloid leukemia (Article)

2013-02-21T00:00:00Z

Treatment factors rather than genetic variation determine metabolic syndrome in childhood cancer survivors (Article)

2013-02-01T00:00:00Z

Background: Genetic variation that regulates insulin resistance, blood pressure and adiposity in the normal population might determine differential vulnerability for metabolic syndrome after treatment for childhood cancer. Objective: To evaluate the contribution of candidate single nucleotide polymorphisms (SNPs) relevant for metabolic syndrome in our single centre cohort of adult long-term childhood cancer survivors. Methods: In this retrospective study 532 survivors were analysed. Median age at diagnosis was 5.7 years (range 0.0-17.8 years), median follow-up time was 17.9 years (range 5.0-48.8) and median age at follow-up was 25.6 years (range 18.0-50.8). JAZF1 gene rs864745, THADA gene rs7578597, IRS1 gene rs2943641, TFAP2B gene rs987237, MSRA gene rs7826222, ATP2B1 gene rs2681472 and rs2681492 were genotyped. The association of genotypes with total cholesterol levels, blood pressure, body mass index, waist circumference and frequency of diabetes were assessed. Results: Metabolic syndrome was more frequent in cranially (23.3%, P = 0.002) and abdominally (23.4%, P = 0.009) irradiated survivors as compared with non-irradiated survivors (10.0%). Association of allelic variants in rs2681472 and rs2681492 with hypertension, rs987237 and rs7826222 with waist circumference and rs864745, rs7578597 and rs2943641 with diabetes were not significant. None of the SNPs was associated with the metabolic syndrome. Adjusting for age, sex, follow-up time, cranial irradiation and abdominal irradiation did not change these results. Conclusions: Treatment factors and not genetic variation determine hypertension, waist circumference, diabetes and metabolic syndrome in adult long-term survivors of childhood cancer.

Peripheral stem cell harvest using regular chemotherapy schedules in childhood cancer (Article)

2012-11-01T00:00:00Z

Prediction of the best moment for the harvest of PBSCs after standard chemotherapy followed by filgrastim in children with cancer is difficult. We retrospectively analyzed the moment of harvesting of 152 procedures in 94 patients. The start of apheresis was guided by WBC count and CD34+ cell measurement in peripheral blood. We defined the first day of filgrastim administration, after completion of mobilizing chemotherapy, as day 1. Median time to harvest in different subgroups is as follows: neuroblastoma 11 days (range, 6-29 days), Ewing's sarcoma nine days (range, 7-15 days), brain tumor 10 days (range, 7-15 days), relapsed Wilms' tumor 16 days (range, 9-20 days), and extracranial GCT seven days (range, 6-14 days). Patients harvested after cyclophosphamide priming (time to harvest within a range of 8-9 days) were analyzed as a separate group. The optimal moment for harvesting in different types of tumors was highly variable, although most consistent in patients diagnosed with Ewing's sarcoma or brain tumors and after cyclophosphamide priming.

Different outcome in older children with acute lymphoblastic leukemia with different treatment protocols in the Netherlands (Article)

2012-01-01T00:00:00Z

Background: From 1991 until 2004 children with acute lymphoblastic leukemia (ALL) in the Netherlands were treated according to protocols ALL-8 and ALL-9 which were based on different principles. An earlier study showed that the outcome of adolescents highly differed on these protocols. Procedure: In this retrospective study, we analyzed whether the outcome of older children 10-15 years of age at diagnosis differed between the Berlin-Frankfurt-Münster (BFM)-based ALL-8 regimen and the ALL-9 regimen. Two hundred fifty-four older children who were treated according to protocol ALL-8 (n=82) or ALL-9 (n=172) were included in the analysis. Results: A higher 5-year event-free survival (EFS) rate was found for patients treated according to ALL-8 compared to ALL-9 (79±5% vs. 65±4%, P=0.02). Patient characteristics did not differ except for a slightly higher age in ALL-8. Therefore, additional analyses were done including only patients who were 12-15 years of age. In this age group there was also a difference in the 5-year EFS (82±5% vs. 61±5%, P=0.00) as well as in the 5-year overall survival rate; 89±4% compared to 68±5%, respectively (P=0.01). Major difference between protocols was the use of a consolidation and reinduction/intensification course and higher cumulative doses of asparaginase, methotrexate, and anthracyclines in ALL-8. Conclusions: Children 10-15 years of age have been undertreated with the ALL-9 regimen and benefit by intensive treatment components as used in ALL-8. We recommend using BFM-based protocols for these older children with ALL.

The controversy of varicella vaccination in children with acute lymphoblastic leukemia (Article)

2012-01-01T00:00:00Z

Prospective study on incidence, risk factors, and long-term outcome of osteonecrosis in pediatric acute lymphoblastic leukemia (Article)

2011-11-01T00:00:00Z

Purpose: We studied cumulative incidence, risk factors, therapeutic strategies, and outcome of symptomatic osteonecrosis in pediatric patients with acute lymphoblastic leukemia (ALL). Patients and Methods: Cumulative incidence of osteonecrosis was assessed prospectively in 694 patients treated with the dexamethasone-based Dutch Child Oncology Group-ALL9 protocol. Osteonecrosis was defined by development of symptoms (National Cancer Institute grade 2 to 4) during treatment or within 1 year after treatment discontinuation, confirmed by magnetic resonance imaging. We evaluated risk factors for osteonecrosis using logistic multivariate regression. To describe outcome, we reviewed clinical and radiologic information after antileukemic treatment 1 year or more after osteonecrosis diagnosis. Results: Cumulative incidence of osteonecrosis at 3 years was 6.1%. After adjustment for treatment center, logistic multivariate regression identified age (odds ratio [OR], 1.47; P < .01) and female sex (OR, 2.23; P = .04) as independent risk factors. Median age at diagnosis of ALL in patients with osteonecrosis was 13.5 years, compared with 4.7 years in those without. In 21 (55%) of 38 patients with osteonecrosis, chemotherapy was adjusted. Seven patients (18%) underwent surgery: five joint-preserving procedures and two total-hip arthroplasties. Clinical follow-up of 35 patients was evaluated; median follow-up was 4.9 years. In 14 patients (40%), symptoms completely resolved; 14 (40%) had symptoms interfering with function but not with activities of daily living (ADLs; grade 2); seven (20%) had symptoms interfering with ADLs (grade 3). In 24 patients, radiologic follow-up was available; in six (25%), lesions improved/disappeared; in 13 (54%), lesions remained stable; five (21%) had progressive lesions. Conclusion: Six percent of pediatric patients with ALL developed symptomatic osteonecrosis during or shortly after treatment. Older age and female sex were risk factors. After a median follow-up of 5 years, 60% of patients had persistent symptoms.

Integrative analysis of type-I and type-II aberrations underscores the genetic heterogeneity of pediatric acute myeloid leukemia (Article)

2011-10-01T00:00:00Z

Background Several studies of pediatric acute myeloid leukemia have described the various type-I or type- II aberrations and their relationship with clinical outcome. However, there has been no recent comprehensive overview of these genetic berrations in one large pediatric acute myeloid leukemia cohort. Design and Methods We studied the different genetic aberrations, their associations and their impact on prognosis in a large pediatric acute myeloid leukemia series (n=506). Karyotypes were studied, and hotspot regions of NPM1, CEPBA, MLL, WT1, FLT3, N-RAS, K-RAS, PTPN11 and KIT were screened for mutations of available samples. The mutational status of all type-I and type-II aberrations was available in 330 and 263 cases, respectively. Survival analysis was performed in a subset (n=385) treated on consecutive acute myeloid leukemia Berlin-Frankfurt-Munster Study Group and Dutch Childhood Oncology Group treatment protocols. Results Genetic aberrations were associated with specific clinical characteristics, e.g. significantly higher diagnostic white blood cell counts in MLL-rearranged, WT1-mutated and FLT3-ITD-positive acute myeloid leukemia. Furthermore, there was a significant difference in the distribution of these aberrations between children below and above the age of two years. Non-random associations, e.g. KIT mutations with core-binding factor acute myeloid leukemia, and FLT3-ITD with t(15;17)(q22;q21), NPM1- and WT1-mutated acute myeloid leukemia, respectively, were observed. Multivariate analysis revealed a 'favorable karyotype', i.e. t(15;17)(q22;q21), t(8;21)(q22;q22) and inv(16)(p13q22)/t(16;16)(p13;q22). NPM1 and CEBPA double mutations were independent factors for favorable event-free survival. WT1 mutations combined with FLT3-ITD showed the worst outcome for 5-year overall survival (22±14%) and 5-year eventfree survival (20±13%), although it was not an independent factor in multivariate analysis. Conclusions Integrative analysis of type-I and type-II aberrations provides an insight into the frequencies, non-random associations and prognostic impact of the various aberrations, reflecting the heterogeneity of pediatric acute myeloid leukemia. These aberrations are likely to guide the stratification of pediatric acute myeloid leukemia and may direct the development of targeted therapies.

Oncogenic IL7R gain-of-function mutations in childhood T-cell acute lymphoblastic leukemia (Article)

2011-10-01T00:00:00Z

Interleukin 7 (IL-7) and its receptor, formed by IL-7Rα (encoded by IL7R) and γc, are essential for normal T-cell development and homeostasis. Here we show that IL7R is an oncogene mutated in T-cell acute lymphoblastic leukemia (T-ALL). We find that 9% of individuals with T-ALL have somatic gain-of-function IL7R exon 6 mutations. In most cases, these IL7R mutations introduce an unpaired cysteine in the extracellular juxtamembrane-transmembrane region and promote de novo formation of intermolecular disulfide bonds between mutant IL-7Rα subunits, thereby driving constitutive signaling via JAK1 and independently of IL-7, γc or JAK3. IL7R mutations induce a gene expression profile partially resembling that provoked by IL-7 and are enriched in the T-ALL subgroup comprising TLX3 rearranged and HOXA deregulated cases. Notably, IL7R mutations promote cell transformation and tumor formation. Overall, our findings indicate that IL7R mutational activation is involved in human T-cell leukemogenesis, paving the way for therapeutic targeting of IL-7R-mediated signaling in T-ALL.

NUP98/NSD1 characterizes a novel poor prognostic group in acute myeloid leukemia with a distinct HOX gene expression pattern (Article)

2011-09-29T00:00:00Z

Translocations involving nucleoporin 98kD (NUP98) on chromosome 11p15 occur at relatively low frequency in acute myeloid leukemia (AML) but can be missed with routine karyotyping. In this study, high-resolution genome-wide copy number analyses revealed cryptic NUP98/NSD1 translocations in 3 of 92 cytogenetically normal (CN) - AML cases. To determine their exact frequency, we screened > 1000 well-characterized pediatric and adult AML cases using a NUP98/NSD1-specific RT-PCR. Twenty-three cases harbored the NUP98/NSD1 fusion, representing 16.1% of pediatric and 2.3% of adult CN-AML patients. NUP98/NSD1-positive AML cases had significantly higher white blood cell counts (median, 147 × 109/L), more frequent FAB-M4/M5 morphology (in 63%), and more CN-AML (in 78%), FLT3/internal tandem duplication (in 91%) and WT1 mutations (in 45%) than NUP98/NSD1-negative cases. NUP98/NSD1 was mutually exclusive with all recurrent type-II aberrations. Importantly, NUP98/NSD1 was an independent predictor for poor prognosis; 4-year event-free survival was < 10% for both pediatric and adult NUP98/NSD1-positiveAML patients. NUP98/NSD1-positive AML showed a characteristic HOX-gene expression pattern, distinct from, for example, MLLrearranged AML, and the fusion protein was aberrantly localized in nuclear aggregates, providing insight into the leukemogenic pathways of these AMLs. Taken together, NUP98/NSD1 identifies a previously unrecognized group of young AML patients, with distinct characteristics and dismal prognosis, for whom new treatment strategies are urgently needed.

Discovery of new microRNAs by small RNAome deep sequencing in childhood acute lymphoblastic leukemia (Article)

2011-09-01T00:00:00Z

MicroRNAs (miRNAs) relevant to acute lymphoblastic leukemia (ALL) in children are hypothesized to be largely unknown as most miRNAs have been identified in non-leukemic tissues. In order to discover these miRNAs, we applied high-throughput sequencing to pooled fractions of leukemic cells obtained from 89 pediatric cases covering seven well-defined genetic types of ALL and normal hematopoietic cells. This resulted into 78 million small RNA reads representing 554 known, 28 novel and 431 candidate novel miR genes. In all, 153 known, 16 novel and 170 candidate novel mature miRNAs and miRNA-star strands were only expressed in ALL, whereas 140 known, 2 novel and 82 candidate novel mature miRNAs and miRNA-star strands were unique to normal hematopoietic cells. Stem-loop reverse transcriptase (RT)-quantitative PCR analyses confirmed the differential expression of selected mature miRNAs in ALL types and normal cells. Expression of 14 new miRNAs inversely correlated with expression of predicted target genes (-0.49≤Spearman's correlation coefficients (Rs)≤-0.27, P≤0.05); among others, low levels of novel sol-miR-23 associated with high levels of its predicted (antiapoptotic) target BCL2 (B-cell lymphoma 2) in precursor B-ALL (Rs-0.36, P=0.007). The identification of >1000 miR genes expressed in different types of ALL forms a comprehensive repository for further functional studies that address the role of miRNAs in the biology of ALL.

In vitro efficacy of forodesine and nelarabine (ara-G) in pediatric leukemia (Article)

2011-08-25T00:00:00Z

Forodesine and nelarabine (the pro-drug of ara-G) are 2 nucleoside analogues with promising anti-leukemic activity. To better understand which pediatric patients might benefit from forodesine or nelarabine (ara-G) therapy, we investigated the in vitro sensitivity to these drugs in 96 diagnostic pediatric leukemia patient samples and the mRNA expression levels of different enzymes involved in nucleoside metabolism. Forodesine and ara-G cytotoxicities were higher in T-cell acute lymphoblastic leukemia (T-ALL) samples than in B-cell precursor (BCP)-ALL and acute myeloid leukemia (AML) samples. Resistance to forodesine did not preclude ara-G sensitivity and vice versa, indicating that both drugs rely on different resistance mechanisms. Differences in sensitivity could be partly explained by significantly higher accumulation of intra-cellular dGTP in forodesine-sensitive samples compared with resistant samples, and higher mRNA levels of dGK but not dCK. The mRNA levels of the transporters ENT1 and ENT2 were higher in ara-G-sensitive than -resistant samples. We conclude that especially T-ALL, but also BCP-ALL, pediatric patients may benefit from forodesine or nelarabine (ara-G) treatment.

Frequency and prognostic implications of JAK 1-3 aberrations in Down syndrome acute lymphoblastic and myeloid leukemia (Article)

2011-08-01T00:00:00Z

The heterogeneity of pediatric MLL-rearranged acute myeloid leukemia (Article)

2011-08-01T00:00:00Z

Translocations involving the mixed-lineage leukemia (MLL) gene, localized at 11q23, comprise 15 to 20% of all pediatric acute myeloid leukemia (AML) cases. This review summarizes current knowledge about the etiology, biology, clinical characteristics and differences in outcome in MLL-rearranged pediatric AML. Furthermore, we discuss the role of cooperating events in MLL-rearranged pediatric AML, and future therapeutic strategies to improve outcome. We conclude that MLL-rearranged pediatric AML is a heterogeneous disease, and prognosis depends on various factors, for example, translocation partner, age, WBC and additional cytogenetic aberrations. The relationship of outcome with specific translocation partners requires that they be searched for in the diagnostic work-up of AML. To achieve further improvements in outcome, unraveling the biology of MLL-rearranged pediatric AML is warranted.

CD73 (5′-nucleotidase) expression has no prognostic value in children with acute lymphoblastic leukemia (Article)

2011-08-01T00:00:00Z

Prognostic significance of additional cytogenetic aberrations in 733 de novo pediatric 11q23/MLL-rearranged AML patients: Results of an international study (Article)

2011-06-30T00:00:00Z

We previously demonstrated that outcome of pediatric 11q23/MLL-rearranged AML depends on the translocation partner (TP). In this multicenter international study on 733 children with 11q23/MLL-rearranged AML, we further analyzed which additional cytogenetic aberrations (ACA) had prognostic significance. ACAs occurred in 344 (47%) of 733 and were associated with unfavorable outcome (5-year overall survival [OS] 47% vs 62%, P < .001). Trisomy 8, the most frequent specific ACA (n = 130/344, 38%), independently predicted favorable outcome within the ACAs group (OS 61% vs 39%, P = .003; Cox model for OS hazard ratio (HR) 0.54, P = .03), on the basis of reduced relapse rate (26% vs 49%, P < .001). Trisomy 19 (n = 37/344, 11%) independently predicted poor prognosis in ACAs cases, which was partly caused by refractory disease (remission rate 74% vs 89%, P = .04; OS 24% vs 50%, P < .001; HR 1.77, P = .01). Structural ACAs had independent adverse prognostic value for event-free survival (HR 1.36, P = .01). Complex karyotype, defined as ≥ 3 abnormalities, was present in 26% (n = 192/733) and showed worse outcome than those without complex karyotype (OS 45% vs 59%, P = .003) in univariate analysis only. In conclusion, like TP, specific ACAs have independent prognostic significance in pediatric 11q23/MLL-rearranged AML, and the mechanism underlying these prognostic differences should be studied.

MicroRNAs in acute leukemia: from biological players to clinical contributors (Article)

2011-06-24T00:00:00Z

MicroRNAs (miRNAs) are involved in the management of hematopoiesis. As a consequence, miRNA dysregulation causes disruption of the hematopoietic system and leukemia may arise. We here comprehensively discuss miRNAs found discriminative for cytogenetic and molecular subtypes of acute leukemia. These miRNAs are either known miRNAs involved in leukemogenesis with proven tumor suppressor or oncogenic activities or are newly identified by high-throughput sequencing with yet unknown function. Furthermore, forces are outlined that drive aberrant miRNA function, which include genetic abnormalities (for example, deletions, translocations and mutations) and epigenetic aberrations (for example, aberrant DNA methylation or histone modifications). Interestingly, leukemia-silenced miRNAs can be re-expressed upon treatment with de-methylating agents. Targeting miRNA expression may serve a therapeutical role, albeit at present this way of targeted therapy is in its infancy. However, emerging knowledge about the biology of miRNAs in leukemia may result into a role for these miRNAs in the diagnosis and treatment of acute leukemia.Leukemia advance online publication, 24 June 2011; doi:10.1038/leu.2011.151.

The long-term impact of in vitro drug sensitivity on risk stratification and treatment outcome in acute lymphoblastic leukemia of childhood (CoALL 06-97) (Article)

2011-06-01T00:00:00Z

Background In a study of childhood acute lymphoblastic leukemia (CoALL 06-97 study), the in vitro sensitivityof the patients' cells to prednisolone, vincristine and asparaginase was introduced as anew additional risk parameter for treatment stratification. In parallel in vivo treatment responsewas assessed by determining the presence and extent of minimal residual disease in a subset ofpatients (n=224). Here we report the long-term impact of in vitro sensitivity-based risk stratificationaccording to survival and compare the results of in vitro sensitivity with in vivo response. Design and Methods Patients with a sensitive in vitro profile were treated with a reduced intensity protocol (n=167)whereas patients defined as low risk according to conventional parameters but with a resistantin vitro profile were given intensified therapy (n=47). Results At a median follow-up of 6.8 years event-free survival was 0.80±0.03 for patients with a sensitiveprofile, 0.73±0.03 for those with an intermediate profile and 0.67±0.08 for those with aresistant profile (P=0.015). Overall, the treatment results of the cases stratified according to invitro sensitivity were similar to those of the historical control group stratified based on conventionalrisk factors. Minimal residual disease at the end of induction was a strong predictor ofoutcome in B-precursor and T-cell acute lymphoblastic leukemia. There was no correlationbetween in vitro and in vivo treatment response in B-precursor leukemia (Spearman's r=0.13;P=0.15) in contrast to T-cell acute lymphoblastic leukemia (Spearman's r=0.63; P<0.001) Conclusions A moderate reduction in treatment intensity for patients with a sensitive in vitro profile waspossible without jeopardizing treatment outcome. However, in vitro drug testing was affectedby a decrease in risk predictive power over time and was not correlated with in vivo assessmentof minimal residual disease in B-precursor acute lymphoblastic leukemia. It was, therefore,abandoned in favor of the assessment of in vivo response in subsequent CoALL trials.

MicroRNA characterize genetic diversity and drug resistance in pediatric acute lymphoblastic leukemia (Article)

2011-05-01T00:00:00Z

Background MicroRNA regulate the activity of protein-coding genes including those involved in hematopoietic cancers. The aim of the current study was to explore which microRNA are unique for seven different subtypes of pediatric acute lymphoblastic leukemia. Design and Methods Expression levels of 397 microRNA (including novel microRNA) were measured by quantitative real-time polymerase chain reaction in 81 cases of pediatric leukemia and 17 normal hematopoietic control cases. Results All major subtypes of acute lymphoblastic leukemia, i.e. T-cell, MLL-rearranged, TEL-AML1- positive, E2A-PBX1-positive and hyperdiploid acute lymphoblastic leukemia, with the exception of BCR-ABL-positive and 'B-other' acute lymphoblastic leukemias (defined as precursor Bcell acute lymphoblastic leukemia not carrying the foregoing cytogenetic aberrations), were found to have unique microRNA-signatures that differed from each other and from those of healthy hematopoietic cells. Strikingly, the microRNA signature of TEL-AML1-positive and hyperdiploid cases partly overlapped, which may suggest a common underlying biology. Moreover, aberrant down-regulation of let-7b (~70-fold) in MLL-rearranged acute lymphoblastic leukemia was linked to up-regulation of oncoprotein c-Myc (PFDR<0.0001). Resistance to vincristine and daunorubicin was characterized by an approximately 20-fold up-regulation of miR- 125b, miR-99a and miR-100 (PFDR≤0.002). No discriminative microRNA were found for prednisolone response and only one microRNA was linked to resistance to L-asparaginase. A combined expression profile based on 14 microRNA that were individually associated with prognosis, was highly predictive of clinical outcome in pediatric acute lymphoblastic leukemia (5- year disease-free survival of 89.4%±7% versus 60.8±12%, P=0.001). Conclusions Genetic subtypes and drug-resistant leukemic cells display characteristic microRNA signatures in pediatric acute lymphoblastic leukemia. Functional studies of discriminative and prognostically important microRNA may provide new insights into the biology of pediatric acute lymphoblastic leukemia.

Integrated Transcript and Genome Analyses Reveal NKX2-1 and MEF2C as Potential Oncogenes in T Cell Acute Lymphoblastic Leukemia (Article)

2011-04-12T00:00:00Z

To identify oncogenic pathways in T cell acute lymphoblastic leukemia (T-ALL), we combined expression profiling of 117 pediatric patient samples and detailed molecular-cytogenetic analyses including the Chromosome Conformation Capture on Chip (4C) method. Two T-ALL subtypes were identified that lacked rearrangements of known oncogenes. One subtype associated with cortical arrest, expression of cell cycle genes, and ectopic NKX2-1 or NKX2-2 expression for which rearrangements were identified. The second subtype associated with immature T cell development and high expression of the MEF2C transcription factor as consequence of rearrangements of MEF2C, transcription factors that target MEF2C, or MEF2C-associated cofactors. We propose NKX2-1, NKX2-2, and MEF2C as T-ALL oncogenes that are activated by various rearrangements.

Erroneous exchange of asparaginase forms in the treatment of acute lymphoblastic leukemia (Article)

2011-04-01T00:00:00Z

For the treatment of children with acute lymphoblastic leukemia (ALL), Dutch pediatric oncologists use the Dutch Childhood Oncology Group ALL 10 protocol. This protocol is complex, as it comprises many different drug regimens. One of the drugs is asparaginase which is available in different forms with different pharmacokinetics: Escherichia coli asparaginase, Erwinia asparaginase, and pegylated E. coli asparaginase [polyethylene glycol (PEG) asparaginase]. Here, we report the case of a 3-year-old patient treated with ALL who was 8 times erroneously treated with E. coli asparaginase instead of PEG asparaginase. As E. coli asparaginase was administered to the patient in the lower dosage regimen of PEG asparaginase, she was undertreated, but at the end of the treatment the patient was in complete remission. This case report describes the actual course of treatment, the reasons why it went wrong, and possible preventive measures.

Germline variation in the MTHFR and MTRR genes determines the nadir of bone density in pediatric acute lymphoblastic leukemia: A prospective study (Article)

2011-03-01T00:00:00Z

Background: This study aims to identify folate-metabolism-related genetic risk factors for low bone mineral density (BMD) during/after pediatric acute lymphoblastic leukemia (ALL) treatment. Patients and methods: We investigated the influence of methylenetetrahydrofolate reductase (MTHFR 677C>T and 1298A>C) and methionine synthase reductase (MTRR 66A>G) single nucleotide polymorphisms (SNPs) on total body BMD (BMDTB) and lumbar spine BMD (BMDLS) in 83 patients. Homocysteine, folate and vitamin B12 were determined. BMD was measured repeatedly using dual-energy X-ray absorptiometry in patients ≥4years (n=68). Results: Carriers of the MTHFR 677 T-allele showed a lower baseline BMDTBthan non-carriers (-0.38 SDS vs. +0.55 SDS, p=0.01) and BMDTBremained lower during/after treatment. MTHFR 677C>T did not influence treatment-related loss of BMDTB(p=0.39). The MTRR 66 G-allele carriers showed a trend towards a lower BMDTBcompared with non-carriers. Combining these two SNPs, patients carrying ≥2 risk alleles had a significantly lower BMDTB(-1.40 SDS) than patients with one (-0.80 SDS) or no risk alleles (-0.31 SDS). Although carriers of the MTHFR 1298A>C had higher homocysteine levels, this SNP was not related to BMDTB. BMDLSof carriers was similar to non-carriers of the investigated SNPs. Conclusions: The MTHFR 677C>T SNP and the MTRR 66A>G SNP were identified as determinants of impaired BMDTBin childhood ALL patients.

Characterization of CEBPA mutations and promoter hypermethylation in pediatric acute myeloid leukemia (Article)

2011-03-01T00:00:00Z

Background Dysfunctioning of CCAAT/enhancer binding protein α (C/EBP α) in acute myeloid leukemia can be caused, amongst others, by mutations in the encoding gene (CEBPA) and by promoter hypermethylation. CEBPA-mutated acute myeloid leukemia is associated with a favorable outcome, but this may be restricted to the case of double mutations in CEBPA in adult acute myeloid leukemia. In pediatric acute myeloid leukemia, data on the impact of these mutations are limited to one series, and data on promoter hypermethylation are lacking. Our objective was to investigate the characteristics, gene expression profiles and prognostic impact of the different CEBPA aberrations in pediatric acute myeloid leukemia. Design and Methods We screened a large pediatric cohort (n=252) for CEBPA single and double mutations by direct sequencing, and for promoter hypermethylation by methylation-specific polymerase chain reaction. Furthermore, we determined the gene-expression profiles (Affymetrix HGU133 plus 2.0 arrays) of this cohort (n=237). Results Thirty-four mutations were identified in 20 out of the 252 cases (7.9%), including 14 doublemutant and 6 single-mutant cases. CEBPA double mutations conferred a significantly better 5-year overall survival compared with single mutations (79% versus 25%, respectively; P=0.04), and compared with CEBPA wild-type acute myeloid leukemia excluding core-binding factor cases (47%; P=0.07). Multivariate analysis confirmed that the double mutations were an independent favorable prognostic factor for survival (hazard ratio 0.23, P=0.04). The combination of screening for promoter hypermethylation and gene expression profiling identified five patients with silenced CEBPA, of whom four cases relapsed. All cases characteristically expressed T-lymphoid markers. Moreover, unsupervised clustering of gene expression profiles showed a clustering of CEBPA double-mutant and silenced cases, pointing towards a common hallmark of abrogated C/EBPα-functioning in these acute myeloid leukemias. Conclusions We showed the independent favorable outcome of patients with CEBPA double-mutant acute myeloid leukemia in a large pediatric series. This molecular marker may, therefore, improve risk-group stratification in pediatric acute myeloid leukemia. For the first time, CEBPA-silenced cases are suggested to confer a poor outcome in pediatric acute myeloid leukemia, indicating that further investigation of this aberration is needed. Furthermore, clustering of gene expression profiles provided insight into the biological similarities and diversities of the different aberrations in CEBPA in pediatric acute myeloid leukemia.

Posterior reversible encephalopathy syndrome in childhood cancer (Article)

2011-02-02T00:00:00Z

Background: Posterior reversible encephalopathy syndrome (PRES) is characterized by seizures, headaches, altered mental status, cortical blindness and typical transient lesions on magnetic resonance imaging. Patients and methods: We describe seven childhood cancer patients with clinical and radiological symptoms of PRES, and reviewed all well-documented PRES cases reported during childhood cancer treatment. Results: Fifty-six children with PRES, including our 7 cases, were identified in the literature. Mean age at onset was 9 (range: 2-17) years. Primary diagnoses were acute lymphoblastic leukemia (n = 31), acute myeloid leukemia (n = 5), non-Hodgkin lymphoma (n = 7) and solid tumors (n = 13). PRES patients presented with seizures (n = 50), altered mental status (n = 20), visual disturbances (n = 24) and/or headaches (n = 17). PRES was associated with hypertension in 49 patients. About 86% of the patients had both clinical and radiological reversible symptoms. Four patients developed epilepsy, in one patient ataxia remained and one patient had a persistent mydriasis. Conclusion: Although PRES has predominantly been described in leukemia patients, it occurs in children with solid tumors as well. Hypertension seems to be the most important trigger for the occurrence of PRES during childhood cancer treatment. Seizures are the most common accompanying sign. Symptoms and radiological findings normalize in ~90% of the cases, but in 10% neurological symptoms remain.

Impact of reduced chemotherapy treatment for good risk childhood acute lymphoblastic leukaemia on infectious morbidity (Article)

2011-02-01T00:00:00Z

Reducing infectious morbidity is an important goal to improve childhood acute lymphoblastic leukaemia (ALL) survival. To explore the impact of chemotherapy reduction on infectious morbidity, we compared outpatient and inpatient infectious morbidity of reduced versus intensive (conventional) chemotherapy. One hundred and seventy-one children newly diagnosed with ALL between 2004 and 2007 and treated according to the Dutch Childhood Oncology Group ALL 10 protocol were prospectively followed during the 2-year treatment course. Stratified by minimal residual disease, 54 patients received reduced (standard risk; SR) and 117 patients received intensive (medium risk; MR) intensification/maintenance treatment. SR outpatients had a median of 1 febrile episode versus 4 in MR outpatients (P=0·002). SR patients had fewer hospitalizations for fever. They were admitted a median of 0 times, with a median of 0days of hospitalization, median 0days of fever, median 0 times chemotherapy interruption and median 0 times intravenous antibiotics. MR patients were admitted for fever median 2 times (P<0·001) with 10days of hospitalization (P<0·001), 2days of fever (P<0·001), one chemotherapy interruption (P<0·001) and two intravenous antibiotics administration (P<0·001). These data indicate that reduced intensification/maintenance compared to conventional intensive intensification/maintenance chemotherapy for good risk childhood ALL resulted in major decrease of infectious morbidity.

Evaluation of gene expression signatures predictive of cytogenetic and molecular subtypes of pediatric acute myeloid leukemia (Article)

2011-02-01T00:00:00Z

Background Pediatric acute myeloid leukemia is a heterogeneous disease characterized by non-random genetic aberrations related to outcome. The genetic subtype is currently detected by different diagnostic procedures which differ in success rate and/or specificity. Design and Methods We examined the potential of gene expression profiles to classify pediatric acute myeloid leukemia. Gene expression microarray data of 237 children with acute myeloid leukemia were collected and a double-loop cross validation approach was used to generate a subtype-predictive gene expression profile in the discovery cohort (n=157) which was then tested for its true predictive value in the independent validation cohort (n=80). The classifier consisted of 75 probe sets, representing the top 15 discriminating probe sets for MLL-rearranged, t(8;21)(q22;q22), inv(16)(p13q22), t(15;17)(q21;q22) and t(7;12)(q36;p13)-positive acute myeloid leukemia. Results These cytogenetic subtypes represent approximately 40% of cases of pediatric acute myeloid leukemia and were predicted with 92% and 99% accuracy in the discovery and independent validation cohort, respectively. However, for NPM1, CEBPA, MLL(-PTD), FLT3(-ITD), KIT, PTPN11 and N/K-RAS gene expression signatures had limited predictive value. This may be caused by a limited frequency of these mutations and by underlying cytogenetics. This latter is exemplified by the fact that different gene expression signatures were discovered for FLT3-ITD in patients with normal cytogenetics and in those with t(15;17)(q21;q22)-positive acute myeloid leukemia, which pointed to HOXB-upregulation being specific for FLT3-ITD+ cytogenetically normal acute myeloid leukemia. Conclusions In conclusion, gene expression profiling correctly predicted the most prevalent cytogenetic subtypes of pediatric acute myeloid leukemia with high accuracy. In clinical practice, this gene expression signature may replace multiple diagnostic tests for approximately 40% of pediatric acute myeloid leukemia cases whereas only for the remaining cases (predicted as 'acute myeloid leukemia-other') are additional tests indicated. Moreover, the discriminative genes reveal new insights into the biology of acute myeloid leukemia subtypes that warrants followup as potential targets for new therapies.

High IGSF4 expression in pediatric M5 acute myeloid leukemia with t(9;11)(p22;q23) (Article)

2011-01-20T00:00:00Z

Pediatric mixed-lineage leukemia (MLL) - rearranged acute monoblastic leukemia with t(9;11)(p22;q23) has a favorable outcome compared with other MLL-rearranged AML. The biologic background for this difference remains unknown. Therefore, we compared gene expression profiles (GEPs; Affymetrix HGU133 + 2.0) of 26 t(9; 11)(p22;q23) patients with 42 other MLL-rearranged AML patients to identify differentially expressed genes. IGSF4, a cellcell adhesion molecule, was found to be highly expressed in t(9;11)(p22;q23) patients, which was confirmed by real-time quantitative polymerase chain reaction and Western blot. IGSF4 expression within t(9;11)(p22;q23) patients was 4.9 times greater in French-American-British morphology classification (FAB) - M5 versus other FAB-types (P = .001). Methylation status investigation showed that high IGSF4-expressing t(9;11)(p22;q23) patients with FAB-M5 have no promoter hypermethylation, whereas all other cases do. Cell-line incubation with demethylating agent decitabine resulted in promoter demethylation and increased expression of IGSF4. Down-regulation of IGSF4 by siRNA did not affect proliferation or drug sensitivity. In a cohort of 79 MLL-rearranged AML cases, we show significant better overall survival for cases with high IGSF4 expression (5-year overall survival 0.70 vs 0.37, P = .03) In conclusion, we identified IGSF4 overexpression to be discriminative for t(9;11)(p22;q23) patients with FABM5, regulated partially by promoter methylation and resulting in survival benefit.

L-asparaginase treatment in acute lymphoblastic leukemia (Article)

2011-01-15T00:00:00Z

Asparaginases are a cornerstone of treatment protocols for acute lymphoblastic leukemia (ALL) and are used for remission induction and intensification treatment in all pediatric regimens and in the majority of adult treatment protocols. Extensive clinical data have shown that intensive asparaginase treatment improves clinical outcomes in childhood ALL. Three asparaginase preparations are available: the native asparaginase derived from Escherichia coli (E. coli asparaginase), a pegylated form of this enzyme (PEG-asparaginase), and a product isolated from Erwinia chrysanthemi, ie, Erwinia asparaginase. Clinical hypersensitivity reactions and silent inactivation due to antibodies against E. coli asparaginase, lead to inactivation of E. coli asparaginase in up to 60% of cases. Current treatment protocols include E. coli asparaginase or PEG-asparaginase for first-line treatment of ALL. Typically, patients exhibiting sensitivity to one formulation of asparaginase are switched to another to ensure they receive the most efficacious treatment regimen possible. Erwinia asparaginase is used as a second- or third-line treatment in European and US protocols. Despite the universal inclusion of asparaginase in such treatment protocols, debate on the optimal formulation and dosage of these agents continues. This article provides an overview of available evidence for optimal use of Erwinia asparaginase in the treatment of ALL.

KIAA1524: A novel MLL translocation partner in acute myeloid leukemia (Article)

2011-01-01T00:00:00Z

The Mixed Lineage Leukemia gene on chromosome 11q23 is a frequent site of recurrent translocations in acute leukemias. Its promiscuous character is reflected by the more than 60 different translocation partners described in literature. Prompted by karyotype and atypical FISH results, we identified a new translocation partner in infant acute myeloid leukemia, KIAA1524 on 3q13.13, also known as 'Cancerous Inhibitor of Protein phosphatase 2A (CIP2A)'. This gene was recently identified as a proto-oncogene stabilizing MYC protein in gastric carcinoma. KIAA1524 has never been related to hematologic malignancies before, and the current AML case is the first case in which an MLL-KIAA1524 fusion was described.

Hypermethylation of specific microRNA genes in MLL-rearranged infant acute lymphoblastic leukemia: major matters at a micro scale (Article)

2010-11-30T00:00:00Z

MLL-rearranged acute lymphoblastic leukemia (ALL) in infants (<1 year) is the most aggressive type of childhood leukemia. To develop more suitable treatment strategies, a firm understanding of the biology underlying this disease is of utmost importance. MLL-rearranged ALL displays a unique gene expression profile, partly explained by erroneous histone modifications. We recently showed that t(4;11)-positive infant ALL is also characterized by pronounced promoter CpG hypermethylation. In this study, we investigated whether this widespread hypermethylation also affected microRNA (miRNA) expression. We identified 11 miRNAs that were downregulated in t(4;11)-positive infant ALL as a consequence of CpG hypermethylation. Seven of these miRNAs were re-activated after exposure to the de-methylating agent Zebularine. Interestingly, five of these miRNAs are associated either with MLL or MLL fusions, and for miR-152 we found both MLL and DNA methyltransferase 1 (DNMT1) as potential targeted genes. Finally, a high degree of methylation of the miR-152 CpG island was strongly correlated with a poor clinical outcome. Our data suggests that inhibitors of methylation have a potential beyond re-expression of hypermethylated protein-coding genes in t(4;11)-positive infant ALL. In this study, we provide additional evidence that they should be tested for their efficacy in MLL-rearranged infant ALL in in vivo models.Leukemia advance online publication, 30 November 2010; doi:10.1038/leu.2010.282.

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 (Article)

2010-10-14T00:00:00Z

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.

No prognostic impact of the WT1 gene single nucleotide polymorphism rs16754 in pediatric acute myeloid leukemia (Article)

2010-10-01T00:00:00Z

Future perspectives on minimizing bone density reduction in pediatric acute lymphoblastic leukemia (Article)

2010-09-01T00:00:00Z

Self-Limiting Sternal Tumors of Childhood (SELSTOC) (Article)

2010-07-15T00:00:00Z

Background. Because a sternal mass is often alarming, it is important to identify the clinical features of benign processes. Procedure. Data on clinical presentation, diagnostics, treatment and outcome of pediatric patients presenting with a sternal tumor between 2001 and 2009 were collected from medical records. Results. Among the 1,700 children who were referred to our pediatric-oncology center, 14 presented with a rapidly growing sternal mass. All patients (10 males) were Caucasian and median age was 16 (range: 7-50) months. Reported symptoms were local pain (n=7) and/or raised body temperature (n=5). No major preceding traumas were reported. Physical examination revealed solid tumors with a median diameter of 3 (range: 1-4.5)cm in a pre-sternal/parasternal location. Half of the patients showed red/blue discoloration of the skin. On radiology, dumbbell-shaped lesions extended to the area behind the sternal bone, involving the cartilage, leading to increased distance between ossification centers. Histopathology at diagnosis was available from five patients and showed aspecific chronic or acute inflammation (n=4) and a reactive osteochondromatous lesion (n=1). Laboratory infection parameters were not/only slightly raised and microbiologic cultures were negative in all patients. All tumors decreased in size within 1 month, in both patients with and without antibiotics. On physical examination the tumors disappeared within 6 months. Conclusions. This study reports 14 young children with a rapidly growing sternal mass due to aseptic inflammation, that we named self-limiting sternal tumor of childhood (SELSTOC). To prevent invasive diagnostic interventions and unnecessary treatment, we advocate a wait-and-see approach with close follow-up in the first weeks. Pediatr Blood Cancer 2010;55:81-84.

Methotrexate-induced side effects are not due to differences in pharmacokinetics in children with down syndrome and acute lymphoblastic leukemia (Article)

2010-07-01T00:00:00Z

Background Children with Down syndrome have an increased risk of developing acute lymphoblastic leukemia and a poor tolerance of methotrexate. This latter problem is assumed to be caused by a higher cellular sensitivity of tissues in children with Down syndrome. However, whether differences in pharmacokinetics play a role is unknown. Design and Methods.We compared methotrexate-induced toxicity and pharmacokinetics in a retrospective case-control study between patients with acute lymphoblastic leukemia who did or did not have Down syndrome. Population pharmacokinetic models were fitted to data from all individuals simultaneously using non-linear mixed effect modeling. Results. Overall, 468 courses of methotrexate (1-5 g/m2) were given to 44 acute lymphoblastic leukemia patients with Down syndrome and to 87 acute lymphoblastic leukemia patients without Down syndrome. Grade 3-4 gastrointestinal toxicity was significantly more frequent in the children with Down syndrome than in those without (25.5% versus 3.9%; P=0.001). The occurrence of grade 3-4 gastrointestinal toxicity was not related to plasma methotrexate area under the curve. Methotrexate clearance was 5% lower in the acute lymphoblastic leukemia patients with Down syndrome (P=0.001); however, this small difference is probably clinically not relevant, because no significant differences in methotrexate plasma levels were detected at 24 and 48 hours. Conclusions We did not find evidence of differences in the pharmacokinetics of methotrexate between patients with and without Down syndrome which could explain the higher frequency of gastrointestinal toxicity and the greater need for methotrexate dose reductions in patients with Down syndrome. Hence, these problems are most likely explained by differential pharmacodynamic effects in the tissues between children with and without Down syndrome. Although the number of patients was limited to draw conclusions, we feel that it may be safe in children with Down syndrome to start with intermediate dosages of methotrexate (1-3 g/m2) and monitor the patients carefully.

Low frequency of MLL-partial tandem duplications in paediatric acute myeloid leukaemia using MLPA as a novel DNA screenings technique (Article)

2010-07-01T00:00:00Z

Mixed-lineage leukaemia (MLL)-partial tandem duplications (PTDs) are found in 3-5% of adult acute myeloid leukaemia (AML), and are associated with poor prognosis. In adult AML, MLL-PTD is only detected in patients with trisomy 11 or internal tandem duplications of FLT3 (FLT3-ITD). To date, studies in paediatric AML are scarce, and reported large differences in the frequency of MLL-PTD, frequently utilising mRNA RT-PCR only to detect MLL-PTDs. We studied the frequency of MLL-PTD in a large cohort of paediatric AML (n = 276) and the results from two different methods, i.e. mRNA RT-PCR, and multiplex ligation-dependent probe amplification (MLPA), a method designed to detect copy number differences of specific DNA sequences. In some patients with an MLL-rearrangement, MLL-PTD transcripts were detected, but were not confirmed by DNA-MLPA, indicating that DNA-MLPA can more accurately detect MLL-PTD compared to mRNA RT-PCR. In paediatric AML, MLL-PTD was detected in 7/276 patients (2.5%). One case had a trisomy 11, while the others had normal cytogenetics. Furthermore 4 of the 7 patients revealed a FLT3-ITD, which was significantly higher compared with the other AML cases (p = 0.016). In conclusion, using DNA-MLPA as a novel screenings technique in combination with mRNA RT-PCR a low frequency of MLL-PTD in paediatric AML was found. Larger prospective studies are needed to further define the prognostic relevance of MLL-PTD in paediatric AML.

Pharmacogenetic risk factors for altered bone mineral density and body composition in pediatric acute lymphoblastic leukemia (Article)

2010-05-01T00:00:00Z

Background This study investigates pharmacogenetic risk factors for bone mineral (apparent) density (BM(A)D) and body composition in pediatric acute lymphoblastic leukemia Design and Methods We determined the influence of SNPs in 4 genes (vitamin-D receptor (VDR: BsmI/ApaI/TaqI and Cdx-2/GATA), collagen type I alpha 1 (SpI), estrogen receptor 1 (ESR1: PvuII/XbaI), glucocorticoid receptor (BclI)) on body composition, BM(A)D and fracture risk during dexamethasone-based pediatric acute lymphoblastic leukemia treatment. Body composition and BMD were measured repeatedly during and after treatment using dual energy X-ray absorptiometry. Results Non-carriers of VDR 5'-end (Cdx-2/GATA) haplotype 3 revealed a significant larger fat gain than carriers (D%fat: non-carriers: +1.76SDS, carriers: +0.77SDS, P<0.001). At diagnosis and during therapy, lumbar spine BMD was significantly higher in non-carriers of VDR 5'-end (Cdx-2/GATA) haplotype 3 than in carriers. The other SNPs did not influence BMD or fracture risk during/after treatment. The year after treatment completion, lean body mass increased in non-carriers of ESR1 (PvuII/XbaI) haplotype 3 and decreased in carriers (D lean body mass: non-car-riers:+0.28SDS, carriers: -0.55SDS, P<0.01). Conclusions Only VDR 5'-end (Cdx-2/GATA) haplotype 3 was identified as protective factor against excessive fat gain and as a risk factor for lower lumbar spine BMD during treatment. Carrying ESR1 (PvuII/XbaI) haplotype 3 negatively influenced recovery of lean body mass after pediatric acute lymphoblastic leukemia treatment.

EVI1 overexpression in distinct subtypes of pediatric acute myeloid leukemia (Article)

2010-05-01T00:00:00Z

Overexpression of the ecotropic virus integration-1 (EVI1+) gene (EVI1), localized at chromosome 3q26, is associated with adverse outcome in adult acute myeloid leukemia (AML). In pediatric AML, 3q26 abnormalities are rare, and the role of EVI1 is unknown. We studied 228 pediatric AML samples for EVI1 using gene expression profiling and RQ-PCR. EVI1 was found in 20/213 (9%) of children with de novo AML, and in 4/8 with secondary AML. It was predominantly found in MLL-rearranged AML (13/47), monosomy 7 (2/3), or FAB M6/7 (6/10), and mutually exclusive with core-binding factor AML, t(15;17), and NPM1 mutations. Fluorescent in situ hybridization (FISH) was performed to detect cryptic 3q26 abnormalities. However, none of the EVI1+ patients harbored structural 3q26 alterations. Although significant differences in 4 years pEFS for EVI1+ and EVI1-pediatric AML were observed (28% ±11 vs 44%±4, P=0.04), multivariate analysis did not identify EVI1 as an independent prognostic factor. We conclude that EVI1+ can be found in ∼10% of pediatric AML. Although EVI1+ was not an independent prognostic factor, it was predominantly found in subtypes of pediatric AML that are related with an intermediate to unfavorable prognosis. Further research should explain the role of EVI1+ in disease biology in these cases. Remarkably, no 3q26 abnormalities were identified in EVI1+ pediatric AML.

Childhood leukaemia: Towards improved tailored therapy (Article)

2010-05-01T00:00:00Z

Gene expression profiling-based dissection of MLL translocated and MLL germline acute lymphoblastic leukemia in infants (Article)

2010-04-08T00:00:00Z

Acute lymphoblastic leukemia (ALL) in infants (< 1 year) is characterized by a poor prognosis and a high incidence of MLL translocations. Several studies demonstrated the unique gene expression profile associated with MLL-rearranged ALL, but generally small cohorts were analyzed as uniform patient groups regardless of the type of MLL translocation, whereas the analysis of translocationnegative infant ALL remained unacknowledged. Here we generated and analyzed primary infant ALL expression profiles (n = 73) typified by translocations t(4;11), t(11;19), and t(9;11), or the absence of MLL translocations. Our data show that MLL germline infant ALL specifies a gene expression pattern that is different from both MLL-rearranged infantALL and pediatric precursor B-ALL. Moreover, we demonstrate that, apart from a fundamental signature shared by all MLL-rearranged infant ALL samples, each type of MLL translocation is associated with a translocation-specific gene expression signature. Finally, we show the existence of 2 distinct subgroups among t(4;11)-positive infant ALL cases characterized by the absence or presence of HOXA expression, and that patients lacking HOXA expression are at extreme high risk of disease relapse. These gene expression profiles should provide important novel insights in the complex biology of MLL-rearranged infant ALL and boost our progress in finding novel therapeutic solutions.

Bimodal distribution of genomic MLL breakpoints in infant acute lymphoblastic leukemia treatment (Article)

2010-04-01T00:00:00Z

PHF6 mutations in T-cell acute lymphoblastic leukemia (Article)

2010-04-01T00:00:00Z

Tumor suppressor genes on the X chromosome may skew the gender distribution of specific types of cancer. T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematological malignancy with an increased incidence in males. In this study, we report the identification of inactivating mutations and deletions in the X-linked plant homeodomain finger 6 (PHF6) gene in 16% of pediatric and 38% of adult primary T-ALL samples. Notably, PHF6 mutations are almost exclusively found in T-ALL samples from male subjects. Mutational loss of PHF6 is importantly associated with leukemias driven by aberrant expression of the homeobox transcription factor oncogenes TLX1 and TLX3. Overall, these results identify PHF6 as a new X-linked tumor suppressor in T-ALL and point to a strong genetic interaction between PHF6 loss and aberrant expression of TLX transcription factors in the pathogenesis of this disease.

The metabolic syndrome in adult survivors of childhood cancer, a review (Article)

2010-04-01T00:00:00Z

The number of adult survivors of childhood cancer in the general population has increased. As reports on the prevalence of the metabolic syndrome in adult survivors of childhood cancer are scarce, we reviewed the available literature on the components of the metabolic syndrome in adult survivors of childhood cancer. Although there is a lack of studies estimating the prevalence of metabolic syndrome directly, especially prevalence of insulin resistance, obesity, and dyslipidemia is increased in certain groups. Therefore, adult survivors of childhood cancer are at increased risk of developing cerebrovascular and cardiovascular diseases. Accordingly, it is important to identify the predisposing factors of the metabolic syndrome in cohorts of survivors, to introduce medical interventions, and to subsequently decrease the risk of cerebrovascular and cardiovascular events. Copyright

Pharmacokinetics of daunorubicin and daunorubicinol in infants with leukemia treated in the interfant 99 protocol (Article)

2010-03-01T00:00:00Z

Background. There is an extreme paucity of pharmacokinetic data for anticancer agents in infants. Therefore, we aimed at characterizing the pharmacokinetics for daunorubicin in infants and examined their relationship to age, body weight, and body surface area. Procedure. Leukemia patients treated according to the Interfant 99 protocol received 30 mg/m2 daunorubicin, with dose reduction to 3/4 for patients 6-12 months old and 2/3 for patients <6 months, respectively. Plasma samples from 21 patients (aged 0.05-1.88 years) were collected and analyzed for daunorubicin and daunorubicinol. Samples from 12 children (age 1.6-18.8 years), who received daunorubicin in an earlier investigation, were used for pharmacokinetic model building using the software NONMEM. Results. Plasma concentration time profiles could be described using a two compartment model. Daunorubicin clearance was 43.9 L hr-1m-2±65% and central volume of distribution 16.4 Lm-2±46%, whereas apparent clearance of daunorubicinol was 19.1 L hr-1m-2±32% and apparent volume of distribution 228 Lm-2±80% (mean± interindividual variability). No age-dependency in any of the BSA-normalized pharmacokinetic parameters was observed. Consequently, due to the empirical dose reduction in infants the overall exposure to daunorubicinol in infants was smaller than would be expected from older children. Patients aged <6 months experienced more infections in the induction phase than the group aged 6-12 months at diagnosis. Other toxicities were similar in both groups. Conclusion. We observed no indication of an age-dependency in the pharmacokinetics of daunorubicin.

Standardized MRD quantification in European all trials: Proceedings of the second international symposium on MRD assessment in Kiel, Germany, 18-20 September 2008 (Article)

2010-03-01T00:00:00Z

Assessment of minimal residual disease (MRD) has acquired a prominent position in European treatment protocols for patients with acute lymphoblastic leukemia (ALL), on the basis of its high prognostic value for predicting outcome and the possibilities for implementation of MRD diagnostics in treatment stratification. Therefore, there is an increasing need for standardization of methodologies and harmonization of terminology. For this purpose, a panel of representatives of all major European study groups on childhood and adult ALL and of international experts on PCR-and flow cytometry-based MRD assessment was built in the context of the Second International Symposium on MRD assessment in Kiel, Germany, 18-20 September 2008. The panel summarized the current state of MRD diagnostics in ALL and developed recommendations on the minimal technical requirements that should be fulfilled before implementation of MRD diagnostics into clinical trials. Finally, a common terminology for a standard description of MRD response and monitoring was established defining the terms complete MRD response, MRD persistence and MRD reappearance. The proposed MRD terminology may allow a refined and standardized assessment of response to treatment in adult and childhood ALL, and provides a sound basis for the comparison of MRD results between different treatment protocols.

Association of high-level MCL-1 expression with in vitro and in vivo prednisone resistance in MLL-rearranged infant acute lymphoblastic leukemia (Article)

2010-02-04T00:00:00Z

MLL-rearranged acute lymphoblastic leukemia (ALL) represents an unfavorable type of leukemia that often is highly resistant to glucocorticoids such as prednisone and dexamethasone. Because response to prednisone largely determines clinical outcome of pediatric patients with ALL, overcoming resistance to this drug may be an important step toward improving prognosis. Here, we show how gene expression profiling identifies high-level MCL-1 expression to be associated with prednisolone resistance in MLL-rearranged infant ALL, as well as in more favorable types of childhood ALL. To validate this observation, we determined MCL-1 expression with quantitative reverse transcription-polymerase chain reaction in a cohort of MLL-rearranged infant ALL and pediatric noninfant ALL samples and confirmed that high-level MCL-1 expression is associated with prednisolone resistance in vitro. In addition, MCL-1 expression appeared to be significantly higher in MLL-rearranged infant patients who showed a poor response to prednisone in vivo compared with prednisone good responders. Finally, down-regulation of MCL-1 in prednisolone-resistant MLL-rearranged leukemia cells by RNA interference, to some extent, led to prednisolone sensitization. Collectively, our findings suggest a potential role for MCL-1 in glucocorticoid resistance in MLL-rearranged infant ALL, but at the same time strongly imply that high-level MCL-1 expression is not the sole mechanism providing resistance to these drugs.

Long-term results of Dutch Childhood Oncology Group studies for children with acute lymphoblastic leukemia from 1984 to 2004 (Article)

2010-02-01T00:00:00Z

The Dutch Childhood Oncology Group (DCOG) has used two treatment strategies for children with acute lymphoblastic leukemia (ALL) based on Pinkel's St Jude Total Therapy or the Berlin-Frankfurt-Münster (BFM) backbone. In four successive protocols, 1734 children were treated. Studies ALL-6 and ALL-9 followed the Total Therapy approach; cranial irradiation was replaced by medium-dose methotrexate infusions and prolonged triple intrathecal therapy; dexamethasone was used instead of prednisone. Studies ALL-7 and ALL-8 had a BFM backbone, including more intensive remission induction, early reinduction and maintenance therapy without vincristine and prednisone pulses. The 5-year event-free survival and overall survival increased from 65.4 to 80.6% (P<0.001) and from 78.7 to 86.4% (P0.07) in ALL-7 and ALL-9, respectively. In ALL-7 and ALL-8 National Cancer Institute (NCI) high-risk criteria, male gender, T-lineage ALL and high white blood cells (WBCs) predict poor outcome. In ALL-9 NCI criteria, gender, WBC 100 × 109/l, and T-lineage ALL have prognostic impact. We conclude that the chemotherapy-only approach in children with ALL in Total Therapy-based strategies and BFM-backbone treatment does not jeopardize survival and preserves cognitive functioning. This experience is implemented in the current DCOG-ALL-10 study using a BFM backbone and minimal residual disease-based stratification.

Editorial: Educational symposium on long-term results of large prospective clinical trials for childhood acute lymphoblastic leukemia (1985-2000) (Article)

2010-02-01T00:00:00Z

Polymorphisms in genes involved in vincristine pharmacokinetics or pharmacodynamics are not related to impaired motor performance in children with leukemia (Article)

2010-02-01T00:00:00Z

Introduction: Impaired motor performance in children who completed treatment for acute lymphoblastic leukemia (ALL) may be related to polymorphisms of the metabolising gene CYP3A5 or vincristine toxicity related genes MDR-1 and MAPT. Methods: Motor performance was measured with the Movement Assessment Battery for Children (movement-ABC). DNA, from mononuclear blood cells was genotyped for CYP3A5, MDR-1 and MAPT polymorphisms. Results: Motor performance was not significantly affected by CYP3A5 *3/*3 and CYP3A5*1*3 genotypes, MDR-1 polymorphisms or MAPT haplotype. Conclusion: Our data did not show that CYP3A5, MDR-1 or MAPT polymorphisms are linked to impaired motor performance in children after treatment for ALL.

Biology and Treatment of Acute Lymphoblastic Leukemia (Article)

2010-02-01T00:00:00Z

Acute lymphoblastic leukemia (ALL), the most common type of cancer in children, is a heterogeneous disease in which many genetic lesions result in the development of multiple biologic subtypes. Today, with intensive multiagent chemotherapy, most children who have ALL are cured. The many national or institutional ALL therapy protocols in use tend to stratify patients in a multitude of different ways to tailor treatment to the rate of relapse. This article discusses the factors used in risk stratification and the treatment of pediatric ALL.

Reduced versus intensive chemotherapy for childhood acute lymphoblastic leukemia: Impact on lymphocyte compartment composition (Article)

2010-01-01T00:00:00Z

Chemotherapy for childhood acute lymphoblastic leukemia may cause severe immune damage. The lymphocyte compartment of 140 patients during and after a new strongly reduced (standard risk (SR), n = 43) and intensive chemotherapy regimen (medium risk (MR), n = 97) was studied between 2006 and 2009. Transitional and naive B cells and IgG+/A+, IgM+ and IgM only memory B cells were significantly reduced during chemotherapy; significantly more in MR group. One year after treatment CD27+IgG+/A+, IgM+ and IgM only memory B cells had still not fully recovered, but this was not confined to the MR group. The T cell compartment was less but also significantly affected during chemotherapy and recovered to normal levels. In the MR group, NK cells had not fully recovered to normal levels 1 year after treatment. Thus, intensive chemotherapy regimens cause severe, mainly B cell memory damage that persists even 1 year after treatment.

Specific promoter methylation identifies different subgroups of MLL-rearranged infant acute lymphoblastic leukemia, influences clinical outcome, and provides therapeutic options (Article)

2009-12-24T00:00:00Z

MLL-rearranged infant acute lymphoblastic leukemia (ALL) remains the most aggressive type of childhood leukemia, displaying a unique gene expression profile. Here we hypothesized that this characteristic gene expression signature may have been established by potentially reversible epigenetic modifications. To test this hypothesis, we used differential methylation hybridization to explore the DNA methylation patterns underlying MLL-rearranged ALL in infants. The obtained results were correlated with gene expression data to confirm gene silencing as a result of promoter hypermethylation. Distinct promoter CpG island methylation patterns separated different genetic subtypes of MLL-rearranged ALL in infants. MLL translocations t(4;11) and t(11;19) characterized extensively hypermethylated leukemias, whereas t(9;11)-positive infant ALL and infant ALL carrying wild-type MLL genes epigenetically resembled normal bone marrow. Furthermore, the degree of promoter hypermethylation among infant ALL patients carrying t(4; 11) or t(11;19) appeared to influence relapse-free survival, with patients displaying accentuated methylation being at high relapse risk. Finally, we show that the demethylating agent zebularine reverses aberrant DNA methylation and effectively induces apoptosis in MLL-rearranged ALL cells. Collectively these data suggest that aberrant DNA methylation occurs in the majority of MLL-rearranged infant ALL cases and guides clinical outcome. Therefore, inhibition of aberrant DNA methylation may be an important novel therapeutic strategy for MLL-rearranged ALL in infants.

Imperative of continual support by the European Community for future advances in paediatric oncology in Europe: Meeting report of the EC-funded science-communication project DIRECT "Overcoming Cancer with Research" (Article)

2009-12-01T00:00:00Z

Background/Purpose: Remarkable advances in paediatric cancer research and treatment were achieved during the last decades: In industrialised nations, overall cure rates were raised from below 20% to over 75% during the last forty years. Randomised investigator-driven clinical trials have been the backbone of this progress so far. The EU Clinical Trial Directive was introduced to improve the quality and safety of drug development; however, it has created considerable administrative "extra" workload as well as financial constraints for trials aiming at cancer treatment optimisation in children. It failed to provide harmonisation for clinical trials throughout Europe because translation into national law was highly divergent. To address the resulting challenges for future advances in European paediatric oncology including the promotion of new drug development, improved communication between all stakeholders is warranted. Methods: Supported by the EC-funded FP7 science-communication project "DIRECT", a meeting of European paediatric oncologists, representatives of patients' and parents' associations, survivors, and competent authorities was organised to investigate past successes and future challenges in paediatric oncology in Europe. Results: The DIRECT-Symposium contributed to assess what is required in the future to advance the care and closer approach to "total cure" of European children and adolescents with cancer in all medical, psychological and social aspects. Various strategies were discussed and are summarised in this article. Conclusion: EC-funded science-communication projects such as DIRECThare successful means to foster the public awareness of the necessities in caring for and curing paediat-ric cancer patients in the future. Amongst these, appreciation of the specific conditions and a need of continual funding by the EU are imperative.

WT1 mutations in T-ALL (Article)

2009-11-19T00:00:00Z

The molecular mechanisms involved in disease progression and relapse in T-cell acute lymphoblastic leukemia (T-ALL) are poorly understood. We used single nucleotide polymorphism array analysis to analyze paired diagnostic and relapsed T-ALL samples to identify recurrent genetic alterations in T-ALL. This analysis showed that diagnosis and relapsed cases have common genetic alterations, but also that relapsed samples frequently lose chromosomal markers present at diagnosis, suggesting that relapsed T-ALL emerges from an ancestral clone different from the major leukemic population at diagnosis. In addition, we identified deletions and associated mutations in the WT1 tumor suppressor gene in 2 of 9 samples. Subsequent analysis showed WT1 mutations in 28 of 211 (13.2%) of pediatric and 10 of 85 (11.7%) of adult T-ALL cases. WT1 mutations present in T-ALL are predominantly heterozygous frameshift mutations resulting in truncation of the C-terminal zinc finger domains of this transcription factor. WT1 mutations are most prominently found in T-ALL cases with aberrant rearrangements of the oncogenic TLX1, TLX3, and HOXA transcription factor oncogenes. Survival analysis demonstrated that WT1 mutations do not confer adverse prognosis in pediatric and adult T-ALL. Overall, these results identify the presence of WT1 mutations as a recurrent genetic alteration in T-ALL.

Novel prognostic subgroups in childhood 11q23/MLL-rearranged acute myeloid leukemia: Results of an international retrospective study (Article)

2009-11-19T00:00:00Z

Translocations involving chromosome 11q23 frequently occur in pediatric acute myeloid leukemia (AML) and are associated with poor prognosis. In most cases, the MLL gene is involved, and more than 50 translocation partners have been described. Clinical outcome data of the 11q23-rearranged subgroups are scarce because most 11q23 series are too small for meaningful analysis of subgroups, although some studies suggest that patients with t(9;11)(p22;q23) have a more favorable prognosis. We retrospectively collected outcome data of 756 children with 11q23- or MLL-rearranged AML from 11 collaborative groups to identify differences in outcome based on translocation partners. All karyotypes were centrally reviewed before assigning patients to subgroups. The event-free survival of 11q23/ MLL-rearranged pediatric AML at 5 years from diagnosis was 44%(± 5%), with large differences across subgroups (11% ± 5%to 92% ± 5%). Multivariate analysis identified the following subgroups as independent prognostic predictors: t(1;11)(q21;q23) (hazard ratio [HR] = 0.1, P = .004); t(6; 11)(q27;q23) (HR = 2.2, P < .001); t(10; 11)(p12;q23) (HR = 1.5, P = .005); and t(10;11)(p11.2;q23) (HR = 2.5, P = .005). We could not confirm the favorable prognosis of the t(9;11)(p22;q23) subgroup. We identified large differences in outcome within 11q23/MLL-rearranged pediatric AML and novel subgroups based on translocation partners that independently predict clinical outcome. Screening for these translocation partners is needed for accurate treatment stratification at diagnosis.

Clinical relevance of Wilms tumor 1 gene mutations in childhood acute myeloid leukemia (Article)

2009-11-17T00:00:00Z

Wilms tumor 1 (WT1) mutations have recently been identified in approximately 10% of adult acute myeloid leukemia (AML) with normal cytogenetics (CN-AML) and are associated with poor outcome. Using array-based comparative genome hybridization in pediatric CN-AML samples, we detected a WT1 deletion in one sample. The other WT1 allele was mutated. This prompted us to further investigate the role of WT1 aberrations in childhood AML. Mutations were found in 35 of 298 (12%) diagnostic pediatric AML samples. In 19 of 35 (54%) samples, more than one WT1 aberration was found: 15 samples had 2 different mutations, 2 had a homozygous mutation, and 2 had a mutation plus a WT1 deletion. WT1 mutations clustered significantly in the CN-AML subgroup (22%; P < .001) and were associated with FLT3/ITD (43 vs 17%; P < .001). WT1 mutations conferred an independent poor prognostic significance (WT1 mutated vs wild-type patients: 5-year probability of overall survival [pOS] 35% vs 66%, P = .002; probability of event-free survival 22% vs 46%, P < .001; and cumulative incidence of relapse or regression 70% vs 44%, P < .001). Patients with both a WT1 mutation and a FLT3/ITD had a dismal prognosis (5-year pOS 21%). WT1 mutations occur at a significant rate in childhood AML and are a novel independent poor prognostic marker.

Outcome of congenital acute lymphoblastic leukemia treated on the Interfant-99 protocol (Article)

2009-10-29T00:00:00Z

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 ofMLLrearrangements and CD10-negative B-lineageALLcompared 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.

Components of the metabolic syndrome in 500 adult long-term survivors of childhood cancer (Article)

2009-10-22T00:00:00Z

Background: Adult survivors of childhood cancer have been reported to have an increased risk of late sequels. A cluster of abnormalities that contribute to the metabolic syndrome may be expressed at a higher level and therefore result in an increased risk for diabetes mellitus and cardiovascular diseases. Patients and methods: We investigated a single-centre cohort of 500 adult survivors (228 females) of childhood cancer, median age 28 years (range 18-59 years) and median follow-up time 19 years (range 6-49 years). We measured total cholesterol, high-density lipoprotein-cholesterol, systolic and diastolic blood pressure, body mass index and the prevalence of diabetes mellitus. Data from the epidemiological Monitoring van Risicofactoren en Gezondheid in Nederland (MORGEN) study were used to calculate standard deviation scores as normative values. Results: The criteria of the metabolic syndrome were met in 13% of the total cohort. Acute lymphoblastic leukaemia (ALL) survivors treated with cranial irradiation had an increased risk of developing the metabolic syndrome compared with ALL survivors not treated with cranial irradiation (23% versus 7%, P = 0.011), probably determined by higher prevalence of overweight and hypertension. Conclusion: Adult survivors of childhood cancer, especially those treated with cranial irradiation, are at increased risk of developing the metabolic syndrome.

Cranial irradiation does not result in pituitary-gonadal axis dysfunction in very long-term male survivors of childhood acute lymphoblastic leukemia (Article)

2009-09-03T00:00:00Z

Pyruvate kinase M2 and prednisolone resistance in acute lymphoblastic leukemia (Article)

2009-09-01T00:00:00Z

A randomized trial investigating an exercise program to prevent reduction of bone mineral density and impairment of motor performance during treatment for childhood acute lymphoblastic leukemia (Article)

2009-07-15T00:00:00Z

Background. Reduced bone mineral density (BMD), altered body composition, impaired motor performance and passive ankle dorsiflexion are side effects of acute lymphoblastic leukemia (ALL) treatment. We performed a randomized study investigating whether an exercise program could prevent these side effects. Procedure. At diagnosis we randomized 51 ALL patients (median age: 5.4 years) into a group receiving a 2-year exercise program or a control group receiving standard care. BMD of total body (BMDTB), lumbar spine (BMDLS) and body composition were measured using dual energy X-ray absorptiometry, motor performance with Bayley Scales of Infant Development or Movement-ABC, and passive ankle dorsiflexion with a goniometer. The investigator was blinded to the randomization. Results. Body fat increased equally during treatment in both groups. One year after cessation of therapy more rapid decline of excessive body fat was observed in the intervention group than in the controls (P=0.01). Lean body mass, BMDTBand BMDLSof both groups decreased equally during treatment and increased equally thereafter. Both groups showed a similar decrease in passive ankle dorsiflexion and motor performance during treatment. Adherence to the intervention program varied considerably. Adherence to intervention: 11% of children exercised daily, 37% > once a week, 16% once weekly, 36% < once a week. Conclusions. The exercise program was not more beneficial than standard care in preventing reduction in BMD, motor performance and passive ankle dorsiflexion than standard care, most likely due to unsatisfactory compliance. Increased BMI and body fat in the intervention group normalized faster after cessation of chemotherapy.

Infant acute lymphoblastic leukemia: Lessons learned and future directions (Article)

2009-07-15T00:00:00Z

Compared with acute lymphoblastic leukemia (ALL) in older children, ALL in infants has a dismal outcome because rearrangements of the mixed-lineage leukemia (MLL) gene occur in about 80% of these patients, leading to an aggressive type of leukemia. With most recent therapies, about 50% long-term event-free survival is achieved, but early bone marrow relapse remains a major problem. Early intensification of chemotherapy and new innovative therapies are necessary to improve outcome. Bone marrow transplantation should be limited to a small subset of well-recognized ALL patients with a very poor prognosis. New genetic and epigenetic insights into the biology of MLL-rearranged ALL suggest new possibilities for therapies.

NRIP3: A novel translocation partner of MLL detected in a pediatric acute myeloid leukemia with complex chromosome 11 rearrangements (Article)

2009-07-01T00:00:00Z

Bone mineral density, growth, and thyroid function in long-term survivors of pediatric Hodgkin's lymphoma treated with chemotherapy only (Article)

2009-06-01T00:00:00Z

Background: The aim of this study was to investigate the long-term side effects of treatment for childhood Hodgkin's lymphoma with chemotherapy only on growth, bone mineral density (BMD), body composition, and thyroid function. Procedure: A total of 88 patients (56 male, 32 female; 17.6-42.6 yr), treated for childhood Hodgkin's lymphoma from 1974-1998 with combination chemotherapy adriamycin (doxorubicin), bleomycin, vinblastine, and dacarbazine or epirubicin, bleomycin, vinblastine, dacarbazine with or without mechlorethamine, oncovin (vincristine), procarbazine, and prednisone (MOPP) with the intention to avoid radiotherapy, participated in this study. Median follow-up was 15.5 yr (range 5.6 -30.2). BMD of lumbar spine and total body (BMD-TB), and body composition were measured using dual-energy x-ray absorptiometry. Bone mineral apparent density of the lumbar spine was calculated to correct for bone size. Free T4and TSH were measured. Results: Men treated with MOPP had a significantly reduced height with normal body proportions. Women treated with MOPP had decreased BMD-TB and bone mineral apparent density of the lumbar spine as compared with healthy controls. Percent body fat was significantly increased in female patients treated without MOPP. Body mass index was significantly increased in male patients treated without MOPP, whereas lean body mass was normal in all patients. All patients, except one, treated with chemotherapy only had normal thyroid function. However, five patients who received additional radiation to the thyroid either had abnormal levels of TSH or free T4, or used thyroid hormones. Conclusions: Lean body mass was normal in all patients; thyroid function was normal in all but one patient. The use of MOPP leads to decreased height and increased body mass index in men and decreased BMD-TB in women. Copyright

Pharmacokinetics of high-dose methotrexate in infants treated for acute lymphoblastic leukemia (Article)

2009-05-01T00:00:00Z

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/m2. 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 mM (range 9.5-313). The median systemic clearance was 6.22 L/hr/m2BSA, and tended to increase with age (P = 0.099). Boys had higher clearance than girls, 6.77 and 5.28 L/hr/ m2(P = 0.030), and tended to have lower median MTX concen-tration at 24 hr. Eight infants had MTX levels below 20 mM, 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 mM. 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.

Beneficial and harmful effects of anthracyclines in the treatment of childhood acute lymphoblastic leukaemia: A systematic review and meta-analysis (Article)

2009-05-01T00:00:00Z

Summary Anthracyclines are used to treat childhood acute lymphoblastic leukaemia (ALL) but non-randomized studies suggest that cardiotoxicity may be a problem. Individual patient data from trials in childhood ALL that randomized anthracyclines or methods of reducing cardiotoxicity were analysed by standard meta-analysis methods. Results were grouped and combined according to: addition of an anthracycline to standard therapy, type of anthracycline, mode of administration, and the use of a cardioprotectant. Data from 958 patients in 4 trials, recruiting between 1972 and 1984, showed that addition of an anthracycline reduced bone marrow relapse and, non-significantly, non-bone marrow relapse, resulting in an increased relapse-free interval. However there was a non-significant increase in induction failures, and in deaths in first remission. Event-free survival at 5 years was 56·7% with anthracycline versus 52·8% without (Odds Ratio = 0·91; 95% Confidence Interval = 0·76-1·10; P = 0·3). There were no significant differences found in other treatment comparisons. The limited data from trials did not demonstrate differences in clinically evident cardiotoxicity. Anthracyclines are effective against bone marrow relapse but have not been shown to significantly increase event free survival in childhood ALL. The evidence on type of anthracycline, method of administration or use of cardioprotectant was insufficient to be able to rule out important differences.

Polymorphisms in folate-related genes and risk of pediatric acute lymphoblastic leukemia (Article)

2009-03-05T00:00:00Z

Polymorphisms in folate pathway genes may influence the susceptibility to acute lymphoblastic leukemia (ALL). DNA was isolated from 245 pediatric ALL patients (cases) and from 500 blood bank donors (controls). Polymorphisms in methylene-tetrahydrofolate reductase (MTHFR677C>T,1298A>C), methionine synthase (MTR 2756A>G), methionine synthase reductase (MTRR 66A>G), methylenetetrahydrofolate dehydrogenase (MTHFD1 1958G>A), nicotinamide N-methyltransferase (NNMT IVS-151C>T), serine hydroxymethyl transferase (SHMT1 1420C>T), thymidylate synthase (TS 2R3R), and the reduced folate carrier (RFC1 80G>A) were detected. In ALL patients, an increased occurrence was observed of the RFC1 80AA variant (odds ratio[OR] = 2.1; 95% confidence interval[CI] = 1.3-3.2; P =.002) and the RFC1 80A allele (OR = 1.5; 95% CI, 1.1-2.1; P =.02). Likewise, the NNMT IVS-151TT genotype showed a 2.2-fold increased ALL risk (OR = 2.2; 95% CI, 1.1-4.6; P =.04). A 1.4-fold reduction in ALL risk was observed for (heterozygous or homozygous) carriers of the TS 2R allele and the MTHFR 677T allele (OR = 0.7; 95% CI, 0.5-1.0; P <.05). Furthermore, interactions between NNMT and MTHFR 677C>T and RFC were observed. NNMT IVS-151CC/MTHFR 677CT + TT patients exhibited a 2-fold reduction in ALL risk whereas RFC1 80AA/NNMT IVS-151CT + TT subjects had a 4.2-fold increase in ALL risk (P =.001). For the first time, we associate the RFC1 80G>A and NNMT IVS-151C> T variants to an increased ALL susceptibility.(Blood. 2009;113:2284-2289)

Inhibition of glycolysis modulates prednisolone resistance in acute lymphoblastic leukemia cells (Article)

2009-02-26T00:00:00Z

Treatment failure in pediatric acute lymphoblastic leukemia (ALL) is related to cellular resistance to glucocorticoids (eg, prednisolone). Recently, we demonstrated that genes associated with glucose metabolism are differentially expressed between prednisolone-sensitive and prednisolone-resistant precursor B-lineage leukemic patients. Here, we show that prednisolone resistance is associated with increased glucose consumption and that inhibition of glycolysis sensitizes prednisolone-resistant ALL cell lines to glucocorticoids. Treatment of prednisolone-resistant Jurkat and Molt4 cells with 2-deoxy-D-glucose (2-DG), lonidamine (LND), or3-bromopyruvate (3-BrPA) increased the in vitro sensitivity to glucocorticoids, while treatment of the prednisolone-sensitive cell lines Tom-1 and RS4; 11 did not influence drug cyto-toxicity. This sensitizing effect of the glycolysis inhibitors in glucocorticoid-resistant ALL cells was not found for other classes of antileukemic drugs (ie, vincris-tine and daunorubicin). Moreover, down-regulation of the expression of GAPDH by RNA interference also sensitized to prednisolone, comparable with treatment with glycolytic inhibitors. Importantly, the ability of 2-DG to reverse glucocorticoid resistance was not limited to cell lines, but was also observed in isolated primary ALL cells from patients. Together, these findings indicate the importance of the glycolytic pathway in glucocorticoid resistance in ALL and suggest that targeting glycolysis is a viable strategy for modulating prednisolone resistance in ALL.

Identification of new microRNA genes and aberrant microRNA profiles in childhood acute lymphoblastic leukemia (Article)

2009-02-19T00:00:00Z

MicroRNAs (miRNAs) control the expression of protein-coding genes in normal hematopoietic cells and, consequently, aberrant expression may contribute to leukemogenesis. To identify miRNAs relevant to pediatric acute lymphoblastic leukemia (ALL), we cloned 105 known and 8 new miRNA genes expressed in patients' leukemia cells. Instead of known miRNA genes, new miRNA genes were not evolutionarily conserved. Quantification of 19 selected miRNA genes revealed an aberrant expression in ALL as compared with normal CD34+ cells (P<0.02); both upregulated (14/19) and downregulated (5/19) expressions were observed. Eight miRNAs were differentially expressed between MLL and non-MLL precursor B-ALL cases (P<0.05). Most remarkably, miR-708 was 250- up to 6500-fold higher expressed in 57 TEL-AML1, BCR-ABL, E2A-PBX1, hyperdiploid and B-other cases than in 20 MLL-rearranged and 15 T-ALL cases (0.0001< P<0.01), whereas the expression of miR-196b was 500-fold higher in MLL-rearranged and 800-fold higher in 5 of 15 T-ALL cases as compared with the expression level in the remaining precursor B-ALL cases (P<0.001). The expression did not correlate with the maturation status of leukemia cells based on immunoglobulin and T-cell receptor rearrangements, immunophenotype or MLL-fusion partner. In conclusion, we identified new miRNA genes and showed that miRNA expression profiles are ALL subtype-specific rather than linked to the differentiation stadium associated with these subtypes.

Prognostic significance of minimal residual disease in infants with acute lymphoblastic leukemia treated within the Interfant-99 protocol (Article)

2009-02-13T00:00:00Z

Acute lymphoblastic leukemia (ALL) in infants younger than 1 year is a rare but relatively homogeneous disease (∼80% MLL gene rearranged, ∼70% CD10-negative) when compared with childhood and adult ALL. Several studies in children and adults with ALL have shown that minimal residual disease (MRD) status is a strong and independent prognostic factor. We therefore evaluated the prognostic significance of MRD in infant ALL. Ninety-nine infant patients treated according to the Interfant-99 protocol were included in this study. MRD was analyzed by real-time quantitative PCR analysis of rearranged immunoglobulin genes, T-cell receptor genes and MLL genes at various time points (TP) during therapy. Higher MRD levels at the end of induction (TP2) and consolidation (TP3) were significantly associated with lower disease-free survival. Combined MRD information at TP2 and TP3 allowed recognition of three patients groups that significantly differed in outcome. All MRD-high-risk patients (MRD levels ≥10-4 at TP3; 26% of patients) relapsed. MRD-low-risk patients (MRD level <10-4 at both TP2 and TP3) constituted 44% of patients and showed a relapse-rate of only 13%, whereas remaining patients (MRD-medium-risk patients; 30% of patients) had a relapse rate of 31%. Comparison between the current Interfant-06 stratification at diagnosis and the here presented MRD-based stratification showed that both stratifications recognized different subgroups of patients. These data indicate that MRD diagnostics has added value for recognition of risk groups in infant ALL and that MRD diagnostics can be used for treatment intervention in infant ALL as well.Leukemia advance online publication, 12 February 2009; doi:10.1038/leu.2009.17.

A subtype of childhood acute lymphoblastic leukaemia with poor treatment outcome: a genome-wide classification study (Article)

2009-02-01T00:00:00Z

Background: Genetic subtypes of acute lymphoblastic leukaemia (ALL) are used to determine risk and treatment in children. 25% of precursor B-ALL cases are genetically unclassified and have intermediate prognosis. We aimed to use a genome-wide study to improve prognostic classification of ALL in children. Methods: We constructed a classifier based on gene expression in 190 children with newly diagnosed ALL (German Cooperative ALL [COALL] discovery cohort) by use of double-loop cross-validation and validated this in an independent cohort of 107 newly diagnosed patients (Dutch Childhood Oncology Group [DCOG] independent validation cohort). Hierarchical cluster analysis with classifying gene-probe sets revealed a new ALL subtype, the underlying genetic abnormalities of which were characterised by comparative genomic hybridisation-arrays and molecular cytogenetics. Findings: Our classifier predicted ALL subtype with a median accuracy of 90·0% (IQR 88·3-91·7) in the discovery cohort and correctly identified 94 of 107 patients (accuracy 87·9%) in the independent validation cohort. Without our classifier, 44 children in the COALL cohort and 33 children in the DCOG cohort would have been classified as B-other. However, hierarchical clustering showed that many of these genetically unclassified cases clustered with BCR-ABL1-positive cases: 30 (19%) of 154 children with precursor B-ALL in the COALL cohort and 14 (15%) of 92 children with precursor B-ALL in the DCOG cohort had this BCR-ABL1-like disease. In the COALL cohort, these patients had unfavourable outcome (5-year disease-free survival 59·5%, 95% CI 37·1-81·9) compared with patients with other precursor B-ALL (84·4%, 76·8-92·1%; p=0·012), a prognosis similar to that of patients with BCR-ABL1-positive ALL (51·9%, 23·1-80·6%). In the DCOG cohort, the prognosis of BCR-ABL1-like disease (57·1%, 31·2-83·1%) was worse than that of other precursor B-ALL (79·2%, 70·2-88·3%; p=0.026), and similar to that of BCR-ABL1-positive ALL (32·5%, 2·3-62·7%). 36 (82%) of the patients with BCR-ABL1-like disease had deletions in genes involved in B-cell development, including IKZF1, TCF3, EBF1, PAX5, and VPREB1; only nine (36%) of 25 patients with B-other ALL had deletions in these genes (p=0·0002). Compared with other precursor B-ALL cells, BCR-ABL1-like cells were 73 times more resistant to L-asparaginase (p=0·001) and 1·6 times more resistant to daunorubicin (p=0·017), but toxicity of prednisolone and vincristine did not differ. Interpretation: New treatment strategies are needed to improve outcome for this newly identified high-risk subtype of ALL. Funding: Dutch Cancer Society, Sophia Foundation for Medical Research, Paediatric Oncology Foundation Rotterdam, Centre of Medical Systems Biology of the Netherlands Genomics Initiative/Netherlands Organisation for Scientific Research, American National Institute of Health, American National Cancer Institute, and American Lebanese Syrian Associated Charities.

Favorable prognostic impact of NPM1 gene mutations in childhood acute myeloid leukemia, with emphasis on cytogenetically normal AML (Article)

2009-01-01T00:00:00Z

Nucleophosmin (NPM1) mutations occur frequently in adult cytogenetically normal acute myeloid leukemia (CN-AML) and confer favorable outcome. We investigated the frequency and prognostic significance of NPM1 mutations in childhood AML (n=298), specifically focusing on the CN-AML subgroup (n=100). Mutations were found in 8.4%, and clustered significantly in the CN-AML subgroup (22%). No mutations were found in patients below the age of 3 years; in CN-AML, there was an increasing incidence above this age. In the overall group, NPM1 mutations conferred an independent favorable prognostic impact on event-free survival (5-year pEFS 66 vs 39%; P=0.02), which did not translate into a significantly better overall survival (5-year pOS 68 vs 56%; P=0.30). However, when the favorable cytogenetic subgroups [inv(16) and t(8;21)] were excluded from the NPM1 wild-type group, the difference in pOS was borderline statistically significant (68 vs 45%; P=0.07). In the CN-AML cohort, NPM1 mutations were an independent prognostic factor on pEFS (80 vs 39%; P=0.01), and pOS (85 vs 60%; P=0.06), which was not influenced by FLT3/ITD. However, in NPM1 wild-type CN-AML, FLT3/ITD-positive patients had a significantly worse outcome (pEFS 48 vs 18%; P<0.001). We conclude that NPM1 mutations confer a favorable prognosis in childhood AML and in CN-AML in particular.

New Genetic Abnormalities and Treatment Response in Acute Lymphoblastic Leukemia (Article)

2009-01-01T00:00:00Z

Numerous genetic abnormalities have been identified in acute lymphoblastic leukemia (ALL). Here we review the recurrent abnormalities with emphasis on those recently discovered, and discuss their association with chemotherapy resistance or sensitivity and with clinical response to therapy. Also, the role of genetic abnormalities in leukemogenesis and their potential as therapeutic targets will be discussed.

The SWI/SNF chromatin-remodeling complex and glucocorticoid resistance in acute lymphoblastic leukemia (Article)

2008-12-17T00:00:00Z

Background: Glucocorticoids are used in the curative treatment of acute lymphoblastic leukemia (ALL). Resistance to glucocorticoids is an important adverse prognostic factor in newly diagnosed ALL patients but its mechanism is unknown. Because SWI/SNF complex-mediated chromatin remodeling is required for glucocorticoid transcriptional activity in vitro, we investigated whether expression of subunits of the SWI/SNF complex was related to glucocorticoid resistance in ALL. Methods: Gene expression and in vitro sensitivity to prednisolone and dexamethasone were assessed in a training set of primary ALL cells from 177 children with newly diagnosed ALL and a validation set of cells from an independent cohort of 95 ALL patients. The global test method was used to select pathways whose genes were associated with drug sensitivity. Genes involved in chromatin remodeling were identified by use of the Gene Ontology database. Short hairpin RNA (shRNA) was used to knock down mRNA expression of SMARCA4 in glucocorticoid-sensitive Jurkat human ALL cells. Spearman rank correlation, multiple linear regression, and logistic regression were used to investigate associations between gene expression and glucocorticoid sensitivity. All statistical tests were two-sided. Results: Statistically significant associations between decreased expression in ALL cells of genes for core subunits of the SWI/SNF complex - SMARCA4, ARID1A, and SMARCB1 - and resistance to prednisolone and dexamethasone were identified in the training cohort. In the validation cohort, expression of SMARCA4 (P <. 001 and r = -0.43), ARID1A (P =. 016 and r = -0.29), and SMARCB1 (P =. 019 and r = -0.29) in ALL cells was statistically significantly associated with dexamethasone sensitivity, and SMARCA4 expression (P =. 018 and r = -0.28) was statistically significantly associated with prednisolone sensitivity. Prednisolone resistance was higher in SMARCA4 shRNA-transfected Jurkat cells (drug concentration lethal to 50% of the leukemia cells [LC50] = 277 μM) than in control shRNA-transfected cells (LC50= 174 μM, difference = 103 μM, 95% confidence interval of the difference = 100 to 106 μM; P <. 001, t test). Conclusion: Decreased expression of as many as three subunits of the SWI/SNF complex appears to be associated with glucocorticoid resistance in primary ALL cells.

Pharmacokinetics, pharmacodynamics, efficacy, and safety of a new recombinant asparaginase preparation in children with previously untreated acute lymphoblastic leukemia: A randomized phase 2 clinical trial (Article)

2008-12-15T00:00:00Z

The pharmacokinetics, pharmacodynam-ics, efficacy, and safety of a new recom-binant Escherichia coli - asparaginase preparation was compared withAsparagi-nase medac. Thirty-two children with acute lymphoblastic leukemia were randomized to receive one of both agents at a dose of 5000 U/m2every 3 days, for a total of 8 doses during induction treatment. The serum activity-time profile after the first dose of recombinant asparagi-nase was similar to that of Asparaginase medac. The trough serum activities were greater than the desired threshold of 100 U/L in both treatment groups. Asparagine was completely depleted in serum and in cerebrospinal fluid, whereas glutamine levels were only moderately influenced. No significant difference between the 2 treatments regarding the degree of aspar-agine depletion, duration of depletion, complete remission rate, and minimal residual disease status at the end of induction, overall frequency or intensity of adverse events was seen. Observed adverse reactions are known as possible and labeled side effects of asparaginase treatment and chemotherapy. We conclude that the new recombinant asparaginase and other native Asparaginase medac are bioequivalent and have the same pharma-codynamic effects and the same direct toxicity profile in children with acute lym-phoblastic leukemia. This trial was registered at http://www. controlled-trials.com as no. ISRCTN 75734403.

CD34 expression is associated with poor survival in pediatric T-cell acute lymphoblastic leukemia (Article)

2008-12-01T00:00:00Z

Background. Children with T-lineage acute lymphoblastic leukemia (T-ALL) have an inferior outcome with combination chemotherapy compared to B-lineage ALL, and still about 30% of the patients relapse within the first 2 years following diagnosis. As CD34 has been related with poor outcome in ALL in general, we investigated the prognostic significance of the stem cell marker CD34, as well as the association of CD34 positivity with the expression of several multidrug resistance (MDR) genes. Procedure. In this retrospective study, we investigated the prognostic significance of the expression of the early T-cell differentiation marker CD34 and the expression of MDR genes in relation to outcome in a cohort of 72 newly diagnosed pediatric T-ALL patients. Results. CD34 expression was related to a poor 5-year disease-free-survival and a poor 5-year overall survival. Using the Cox proportional hazard model, CD34 expression predicted for increased risk for relapse and death. Expression of CD34 was associated with elevated MDR1 and MRP1 mRNA expression levels. For the entire T-ALL cohort, these expression levels of MDR1 or MRP1 did not independently predict for poor outcome. Conclusions. We conclude that CD34-positive T-ALL has a relatively poor survival that is not explained by the mRNA expression levels of MDR1, LRP, or MRP1.

Biological background of pediatric medulloblastoma and ependymoma: A review from a translational research perspective (Article)

2008-12-01T00:00:00Z

Survival rates of pediatric brain tumor patients have significantly improved over the years due to developments in diagnostic techniques, neurosurgery, chemotherapy, radiotherapy, and supportive care. However, brain tumors are still an important cause of cancer-related deaths in children. Prognosis is still highly dependent on clinical characteristics, such as the age of the patient, tumor type, stage, and localization, but increased knowledge about the genetic and biological features of these tumors is being obtained and might be useful to further improve outcome for these patients. It has become clear that the deregulation of signaling pathways essential in brain development, for example, sonic hedgehog (SHH), Wnt, and Notch pathways, plays an important role in pathogenesis and biological behavior, especially for medulloblastomas. More recently, data have become available about the cells of origin of brain tumors and the possible existence of brain tumor stem cells. Newly developed array-based techniques for studying gene expression, protein expression, copy number aberrations, and epigenetic events have led to the identification of other potentially important biological abnormalities in pediatric medulloblastomas and ependymomas. Copyright 2008 by the Society for Neuro-Oncology.

Differential expression and prognostic significance of SOX genes in pediatric medulloblastoma and ependymoma identified by microarray analysis. (Article)

2008-10-01T00:00:00Z

The objective of this study was to identify differentially expressed and prognostically important genes in pediatric medulloblastoma and pediatric ependymoma by Affymetrix microarray analysis. Among the most discriminative genes, three members of the SOX transcription factor family were differentially expressed. Both SOX4 and SOX11 were significantly overexpressed in medulloblastoma (median, 11-fold and 5-fold, respectively) compared with ependymoma and normal cerebellum. SOX9 had greater expression in ependymoma (median, 16-fold) compared with normal cerebellum and medulloblastoma (p<0.001 for all comparisons). The differential expression of the SOX genes was confirmed at the protein level by immunohistochemical analysis. Survival analysis of the most discriminative probe sets for each subgroup showed that 35 and 13 probe sets were predictive of survival in patients with medulloblastoma and ependymoma, respectively. There was a trend toward better survival with increasing SOX4 expression in medulloblastoma. SOX9 expression was predictive for favorable outcome in ependymoma. The mRNA levels of BCAT1, a mediator of amino acid breakdown, were higher (median, 15-fold) in medulloblastoma patients with metastases compared with those without metastasized disease (p<0.01). However, the correlation between BCAT1 expression and metastatic medulloblastoma could not be confirmed at the protein level. The potential prognostic effect of the genes associated with outcome should be evaluated in ongoing studies using larger groups of patients. Furthermore, our findings support further analysis of the functional properties of the selected genes, especially SOX4 and BCAT1 for medulloblastoma and SOX9 for ependymoma, to evaluate the use of these genes as potential tumor markers, prognostic markers, and drug targets in pediatric brain tumors.

Molecular-genetic insights in paediatric T-cell acute lymphoblastic leukaemia (Article)

2008-10-01T00:00:00Z

Paediatric T-cell acute lymphoblastic leukaemia (T-ALL) is an aggressive malignancy of thymocytes that accounts for about 15% of ALL cases and for which treatment outcome remains inferior compared to B-lineage acute leukaemias. In T-ALL, leukemic transformation of maturating thymocytes is caused by a multistep pathogenesis involving numerous genetic abnormalities that drive normal T-cells into uncontrolled cell growth and clonal expansion. This review provides an overview of the current knowledge on onco- and tumor suppressor genes in T-ALL and suggests a classification of these genetic defects into type A and type B abnormalities. Type A abnormalities may delineate distinct molecular-cytogenetic T-ALL subgroups, whereas type B abnormalities are found in all major T-ALL subgroups and synergize with these type A mutations during T-cell pathogenesis.

Impaired dexamethasone-related increase of anticoagulants is associated with the development of osteonecrosis in childhood acute lymphoblastic leukemia (Article)

2008-10-01T00:00:00Z

Coagulation alterations may be involved in osteonecrosis in childhood acute lymphoblastic leukemia. Retrospectively, we evaluated the available coagulation parameters at diagnosis and during induction treatment of 161 acute lymphoblastic leukemia patients: 24 with symptomatic osteonecrosis (median age: 13.8 years, range 4.0-17.2) and 137 without osteonecrosis (median age: 4.9 years, range 1.0-16.7). Coagulation parameters of both groups were similar at diagnosis. After four weeks of treatment including dexamethasone, levels of antithrombin and protein S were significantly less in osteonecrosis-positive than in osteonecrosis-negative patients. Subsequently, after four doses of asparaginase and tapering dexamethasone, these coagulation parameters equally decreased in both groups. Consequently, nadirs of antithrombin and protein S were significantly lower in osteonecrosis-positive than in osteonecrosis- negative patients, even reaching levels below lower normal limits in the osteonecrosis-positive group. A reduced dexamethasone related increase of antithrombin and protein S, and subsequent decline below normal levels after introduction of asparaginase, may result in a hypercoagulable state, contributing to development of symptomatic osteonecrosis.

Polymorphisms in folate-related genes: Association with side effects of high-dose methotrexate in childhood acute lymphoblastic leukemia (Article)

2008-09-18T00:00:00Z

Circumvention of glucocorticoid resistance in childhood leukemia (Article)

2008-09-01T00:00:00Z

In this study, we determined if in vitro resistance to prednisolone and dexamethasone could be circumvented by cortivazol or methylprednisolone, or reversed by meta-iodobenzylguanidine in pediatric lymphoblastic and myeloid leukemia. As there were strong correlations between the LC50 values (drug concentration inducing 50% leukemic cell kill, LCK) of the different glucocorticoids and median prednisolone/methylprednisolone, prednisolone/dexamethasone and prednisolone/cortivazol LC50 ratios did not differ between the leukemia subtypes, we conclude that none of the glucocorticoids had preferential anti-leukemic activity. Meta-iodobenzylguanidine however, partially reversed glucocorticoid resistance in 19% of the lymphoblastic leukemia samples.

High expression of CD40 on B-cell precursor acute lymphoblastic leukemia blasts is an independent risk factor associated with improved survival and enhanced capacity to up-regulate the death receptor CD95 (Article)

2008-08-15T00:00:00Z

CD40 and CD27, members of the tumor necrosis factor receptor (TNFR) family, are critical regulators of lymphocyte growth and differentiation. In B-cell precursor acute lymphoblastic leukemia (BCP-ALL), we prospectively assessed the impact of CD40 and CD27 on outcome in 121 children treated according to the CoALL06-97 protocol. Expression of both CD40 and CD27 was found to be significantly higher in low- than in high-risk patients as defined by standard clinical risk parameters such as age and white blood cell count. In addition, in multivariable analysis, a very high percentage of CD40+blasts at diagnosis was identified as an independent favorable prognostic factor for relapse-free survival. Of note, high CD40 expression particularly protected against late relapse. In B cells, CD40 is known to enhance both antigen-presenting capacity and sensitivity to pro-apoptotic signals. Yet, although CD40 ligation does result in significant up-regulation of CD80/CD86 in our cohort, it is up-regulation of the death receptor CD95 that significantly correlates with the percentage of CD40+blasts. Thus very high expression of CD40 on BCP-ALL blasts is an independent prognostic marker indicative of superior relapse-free survival that may in part be due to CD40- dependent death receptor up-regulation.

L-Asparaginase and the effect of age on coagulation and fibrinolysis in childhood acute lymphoblastic leukemia (Article)

2008-08-01T00:00:00Z

Alterations in haemostasis are frequently observed in children with acute lymphoblastic leukemia (ALL). It was the objective of this study to analyse age-related disturbances in coagulation and fibrinolysis parameters during the induction phase of the antileukemic treatment. Sixty-four children were classified by age into three groups (1-5, 6-10, 11-16 years), and studied during induction treatment of ALL including four weeks of dexamethasone, followed by two weeks tapering of dexamethasone during which 6,000 IU/m2native L-Asparaginase (total 4 doses) was administered intravenously twice weekly. Blood samples were collected immediately before each L-Asparaginase infusion to analyze procoagulant (fibrinogen, factor [F] II, FV, FVII, F IX, F X) and anticoagulant factors (antithrombin [AT], protein C, protein S), parameters of thrombin generation (F1+2,TAT) and fibrinolysis (α2-antiplasmin, plasminogen, PAP, D-dimer). Children were in a hypercoagulable state after four weeks of dexamethasone due to upregulation of coagulation parameters. Upregulation was highest in the two youngest age groups. During L-Asparaginase treatment the 11- to 16-year-olds showed lower values in procoagulant and, even more, in anticoagulant factor levels compared to the younger children.Activation markers of thrombin generation and fibrinolysis did not change over time during the study period. Decreased synthesis of α2-antiplasmin and plasminogen during L-Asparaginase treatment resulted in less potential of clot lysis by plasmin in children older than 11 years of age. In conclusion, a more severe decline of anticoagulant and fibrinolytic parameters in children between 11 and 16 years of age underline that these children are at higher risk of thrombosis during ALL induction treatment.

Various components of the insulin-like growth factor system in tumor tissue, cerebrospinal fluid and peripheral blood of pediatric medulloblastoma and ependymoma patients (Article)

2008-08-01T00:00:00Z

The insulin-like growth factor (IGF) system plays an important role in neuronal development and may contribute to the development of brain tumors. In this study, we studied mRNA expression levels of IGFs, insulin-like growth factor binding proteins (IGFBPs) and insulin-like growth factor receptors (IGFRs) in 27 pediatric medulloblastomas, 13 pediatric ependymomas and 5 control cerebella. Compared to normal cerebellum, mRNA levels of IGFBP-2 and IGFBP-3 were significantly increased in medulloblastomas and ependymomas. IGFBP-2 expression was indicative of poor prognosis in medulloblastomas, whereas IGFBP-3 mRNA levels were especially high in anaplastic ependymomas. IGFBP-5 and IGF-II mRNA levels were significantly increased in ependymomas compared to control cerebellum. Protein expression levels of IGFs and IGFBPs were analyzed in the cerebrospinal fluid (CSF) of 16 medulloblastoma, 4 ependymoma and 23 control patients by radioimmunoassay to determine whether they could be used as markers for residual disease after surgery. No aberrant CSF protein expression levels were found for ependymoma patients. In medulloblastoma patients, the IGFBP-3 protein levels were significantly higher than in ependymoma patients and controls. Moreover, enhanced levels of proteolytic fragments of IGFBP-3 were found in the CSF of medulloblastoma patients, being in concordance with a significantly increased IGFBP-3 proteolytic activity in the CSF of these patients. In conclusion, our data suggest that the IGF system is of importance in pediatric medulloblastomas and ependymomas. Larger studies should be conducted to validate the predictive values of the levels of intact IGFBP-3 and proteolytic fragments in CSF in the follow-up of medulloblastomas.

Perceived and actual motor competence of children treated for cancer with vincristine containing chemotherapy (Article)

2008-08-01T00:00:00Z

In Childhood Acute Lymphoblastic Leukemia, Blasts at Different Stages of Immunophenotypic Maturation Have Stem Cell Properties (Article)

2008-07-08T00:00:00Z

We examined the leukemic stem cell potential of blasts at different stages of maturation in childhood acute lymphoblastic leukemia (ALL). Human leukemic bone marrow was transplanted intrafemorally into NOD/scid mice. Cells sorted using the B precursor differentiation markers CD19, CD20, and CD34 were isolated from patient samples and engrafted mice before serial transplantation into primary or subsequent (up to quaternary) recipients. Surprisingly, blasts representative of all of the different maturational stages were able to reconstitute and reestablish the complete leukemic phenotype in vivo. Sorted blast populations mirrored normal B precursor cells with transcription of a number of stage-appropriate genes. These observations inform a model for leukemia-propagating stem cells in childhood ALL.

Monoallelic or biallelic LMO2 expression in relation to the LMO2 rearrangement status in pediatric T-cell acute lymphoblastic leukemia (Article)

2008-07-01T00:00:00Z

Increased total-Tau levels in cerebrospinal fluid of pediatric hydrocephalus and brain tumor patients (Article)

2008-07-01T00:00:00Z

Total Tau (t-Tau), hyperphosphorylated Tau (p-Tau(181P)) and β-amyloid(1-42)in cerebrospinal fluid (CSF) have shown to be markers of neuronal and axonal degeneration in various neurological and neurodegenerative diseases. The aim of this study was to evaluate the influence of the presence of a brain tumor and hydrocephalus on t-Tau, p-Tau(181P)and β-amyloid(1-42)levels in CSF of pediatric patients. t-Tau, p-Tau(181P)and β-amyloid(1-42)levels were simultaneously quantified by xMAP®technology in 22 lumbar and 15 ventricular CSF samples from newly diagnosed pediatric brain tumor patients and 39 lumbar and 12 ventricular CSF samples from pediatric patients without a brain tumor. t-Tau, p-Tau(181P)and β-amyloid(1-42)levels in both lumbar and ventricular CSF were not significantly correlated with age. t-Tau levels in lumbar CSF were elevated in brain tumor patients, being especially high in medulloblastoma patients. Lumbar CSF p-Tau(181P)levels were lower in brain tumor patients compared to normal controls. Ventricular levels of t-Tau, p-Tau(181P)and β-amyloid(1-42)were not significantly different between the brain tumor patients and non-tumor patients, but t-Tau levels were significantly increased in patients with radiological signs of hydrocephalus. Two patients with an infected ventriculo-peritoneal drain also had high CSF t-Tau levels. In conclusion, high t-Tau levels in CSF are found in pediatric patients with a brain tumor, patients with hydrocephalus and patients with a serious CNS infection, reflecting neuronal and axonal damage. Ongoing studies should determine whether these neurodegenerative markers in CSF can be used to monitor neuronal and axonal degeneration in these patients during therapy and long-term follow up.

Pharmacokinetic, pharmacodynamic and intracellular effects of PEG-asparaginase in newly diagnosed childhood acute lymphoblastic leukemia: Results from a single agent window study (Article)

2008-06-30T00:00:00Z

L-asparaginase is an effective drug for treatment of children with acute lymphoblastic leukemia (ALL). The effectiveness is thought to result from depletion of asparagine in serum and cells. We investigated the clinical response in vivo of 1000 IU/m2 pegylated (PEG)-asparaginase and its pharmacokinetic, pharmacodynamic and intracellular effects in children with newly diagnosed ALL before start of combination chemotherapy. The in vivo window response was significantly related to immunophenotype and genotype: 26/38 common/pre B-ALL cases, especially those with hyperdiploidy and TELAML1 rearrangement, demonstrated a good clinical response compared to 8/17 T-ALL (P=0.01) and BCRABL-positive ALL (P=0.04). A poor in vivo clinical window response was related to in vitro resistance to L-asparaginase (P=0.02) and both were prognostic factors for long-term event-free survival (hazard ratio 6.4, P=0.004; hazard ratio 3.7, P=0.01). After administration of one in vivo dose of PEG-asparaginase no changes in apoptotic parameters or in intracellular levels of twenty amino acids in leukemic cells could be measured, in contradiction to the changes found after in vitro exposure. This may be explained by the rapid removal of apoptotic cells from the circulation in vivo. One additional dose of PEG-asparaginase upfront ALL treatment did not lead to other severe toxicities.

Eye-movement analysis of search effectiveness (Article)

2008-06-01T00:00:00Z

Advances in eye-tracking technology have promoted its widespread use to understand and improve target searches in psychology, industrial engineering, human factors, medical diagnostics, and marketing. Eye movements are the realization of a complex, unobserved spatiotemporal attention process with many sources of variation. Eye-tracking data often have been aggregated and/or summarized descriptively, because few adequate statistical models are available for their analysis. This article proposes a model that may serve to uncover the latent attention processes of people searching for targets in complex scenes. It recognizes the spatial nature of eye movements and represents two latent attention states, a localization state and an identification state, between which people may switch over time according to a Markov process. A saliency map, based on low-level perceptual features and the scene's organization, guide target searches in the localization state. In the identification state, people verify whether a selected candidate object is the target. The model is applied to analyze commercial eye-tracking data from more than 100 people engaged in a target search task on a computer-simulated retail shelf display. Rapid switching between attention states over time is revealed. Estimates of the feature and saliency maps are provided and found to be related to search performance. The results facilitate the evaluation of the effectiveness of alternative visual search strategies.

T-cell receptor Vβ CDR3 oligoclonality frequently occurs in childhood refractory cytopenia (MDS-RC) and severe aplastic anemia (Article)

2008-06-01T00:00:00Z

(Very) severe acquired aplastic anemia ((v)SAA) and myelodysplastic syndrome (MDS) are rare diseases in childhood. (V)SAA is a bone marrow (BM) failure syndrome characterized by immune-mediated destruction of hematopoietic progenitors. MDS is a malignant clonal stem cell disorder, of which the hypoplastic variant is, in case of absence of a cytogenetic clone, difficult to separate from (v)SAA. Recently, studies provided a molecular signature of autoimmunity in adult (v)SAA, by showing oligoclonality based on the length of the TCR Vβ CDR3 region. We investigated retrospectively the frequency and the discriminative value of TCR Vβ CDR3 oligoclonality in pediatric (v)SAA and MDS patients. Peripheral blood (PB) and/or BM mononuclear cell samples of pediatric patients with (v)SAA (n=38), refractory cytopenia (MDS-RC) (n=28) and 18 controls were analysed via TCR Vβ heteroduplex PCR analysis of extracted RNA. A skewed TCR Vβ CDR3 repertoire was found in 21/38 (v)SAA and in 17/28 RC patients in contrast to 2/18 in the control group. These data suggest an overlapping group of RC and SAA patients that may share a common immune-mediated pathogenesis. Prospective studies are required to establish the clinical value of TCR Vβ CDR3 repertoire analysis to predict the clinical response in these patients.

Lung dendritic cells are stimulated by ultrafine particles and play a key role in particle adjuvant activity (Article)

2008-05-01T00:00:00Z

Background: The adjuvant activity of air pollution particles on allergic airway sensitization is well known, but the cellular mechanisms underlying this adjuvant potential are not clear. Objective: We sough to study the role of dendritic cells and the costimulatory molecules CD80 and CD86 in the adjuvant activity of ultrafine carbon black particles (CBP). Methods: The proliferation of CFSE-labeled DO11.10 CD4 cells was studied after intranasal exposure to particles and ovalbumin (OVA). Next the frequency of myeloid dendritic cells (mDCs) and plasmacytoid dendritic cells and their expression of CD80 and CD86 were studied in the peribronchial lymph nodes (PBLNs). The expression of costimulatory molecules was also studied on bone marrow-derived mDCs after exposure to CBPs in vitro, and the importance of costimulation in CBP adjuvant activity was assessed by using CD80/CD86-deficient mice or cytotoxic T lymphocyte-associated antigen 4 (CTLA4)-Ig in vivo. Results: Our data show that CBPs plus OVA caused proliferation of DO11.10 CD4 cells and high levels of cytokine production in the PBLNs. Furthermore, the combined CBP plus OVA exposure increased the number of mDCs and expression of costimulatory molecules in the PBLNs. In addition, CBPs upregulated the expression of CD80/CD86 molecules on dendritic cells in vitro, which are necessary for the particle adjuvant effects in vivo. Conclusion: Together this study shows the importance of dendritic cells and costimulation in particle adjuvant activity. Furthermore, we show for the first time that CBPs can also directly induce maturation of dendritic cells.

The recurrent SET-NUP214 fusion as a new HOXA activation mechanism in pediatric T-cell acute lymphoblastic leukemia (Article)

2008-05-01T00:00:00Z

T-cell acute lymphoblastic leukemia (TALL) is mostly characterized by specific chromosomal abnormalities, some occurring in a mutually exclusive manner that possibly delineate specific T-ALL subgroups. One subgroup, including MLLrearranged, CALM-AFWor inv (7)(p15q34) patients, is characterized by elevated expression of HOXA genes. Using a gene expression-based clustering analysis of 67 T-ALL cases with recurrent molecular genetic abnormalities and 25 samples lacking apparent aberrations, we identified 5 new patients with elevated HOXA levels. Using microarray-based comparative genomic hybridization (array-CGH), a cryptic and recurrent deletion, del (9)(q34.11q34.13), was exclusively identified in 3 of these 5 patients. This deletion results in a conserved SET-NUP214 fusion product, which was also identified in the T-ALL cell line LOUCY. SET-NUP214 binds in the promoter regions of specific HOXA genes, where it interacts with CRM1 and DOT1L, which may transcriptionally activate specific members of the HOXA cluster. Targeted inhibition of SET-NUP214 by siRNA abolished expression of HOXA genes, inhibited proliferation, and induced differentiation in LOUCY but not in other T-ALL lines. We conclude that SETNUP214 may contribute to the pathogenesis of T-ALL by enforcing T-cell differentia.

Leukemia-associated NF1 inactivation in patients with pediatric T-ALL and AML lacking evidence for neurofibromatosis (Article)

2008-04-15T00:00:00Z

Neurofibromatosis type 1 (NF1) is an autosomal dominant genetic disorder caused by mutations in the NF1 gene. Patients with NF1 have a higher risk to develop juvenile myelomonocytic leukemia (JMML) with a possible progression toward acute myeloid leukemia (AML). In an oligo array comparative genomic hybridization-based screening of 103 patients with pediatric T-cell acute lymphoblastic leukemia (T-ALL) and 71 patients with MLL-rearranged AML, a recurrent cryptic deletion, del(17)(q11.2), was identified in 3 patients with T-ALL and 2 patients with MLL-rearranged AML. This deletion has previously been described as a microdeletion of the NF1 region in patients with NF1. However, our patients lacked clinical NF1 symptoms. Mutation analysis in 4 of these del(17)(q11.2)-positive patients revealed that mutations in the remaining NF1 allele were present in 3 patients, confirming its role as a tumor-suppressor gene in cancer. In addition, NF1 inactivation was confirmed at the RNA expression level in 3 patients tested. Since the NF1 protein is a negative regulator of the R AS pathway (RAS-GTPase activating protein), homozygous NF1 inactivation represent a novel type I mutation in pediatric MLL-rearranged AML and T-ALL with a predicted frequency that is less than 10%. NF1 inactivation may provide an additional proliferative signal toward the development of leukemia.

Cooperative genetic defects in TLX3 rearranged pediatric T-ALL (Article)

2008-04-01T00:00:00Z

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive neoplastic disorder, in which multiple genetic abnormalities cooperate in the malignant transformation of thymocytes. About 20% of pediatric T-ALL cases are characterized by TLX3 expression due to a cryptic translocation t(5;14)(q35;q32). Although a number of collaborating genetic events have been identified in TLX3 rearranged T-ALL patients (NOTCH1 mutations, p15/p16 deletions, NUP214-ABL1 amplifications), further elucidation of additional genetic lesions could provide a better understanding of the pathogenesis of this specific T-ALL subtype. In this study, we used array-CGH to screen TLX3 rearranged T-ALL patients for new chromosomal imbalances. Array-CGH analysis revealed five recurrent genomic deletions in TLX3 rearranged T-ALL, including del(1)(p36.31), del(5)(q35), del(13)(q14.3), del(16)(q22.1) and del(19)(p13.2). From these, the cryptic deletion, del(5)(q35), was exclusively identified in about 25% of TLX3 rearranged T-ALL cases. In addition, 19 other genetic lesions were detected once in TLX3 rearranged T-ALL cases, including a cryptic WT1 deletion and a deletion covering the FBXW7 gene, an U3-ubiquitin ligase that mediates the degradation of NOTCH1, MYC, JUN and CyclinE. This study provides a genome-wide overview of copy number changes in TLX3 rearranged T-ALL and offers great new challenges for the identification of new target genes that may play a role in the pathogenesis of T-ALL.

Decrease in peripheral muscle strength and ankle dorsiflexion as long-term side effects of treatment for childhood cancer (Article)

2008-04-01T00:00:00Z

Background. This study investigated muscle strength, passive ankle dorsiflexion, and their association with motor performance in children after treatment for acute lymphoblastic leukemia, Wilms tumor, B-non-Hodgkin lymphoma, and malignant mesenchymal tumors. Procedure. Muscle strength was assessed with a hand-held dynamometer and ankle dorsiflexion with a goniometer in 92 and 64 survivors, respectively. Motor performance was measured with the Movement Assessment Battery for Children (movement-ABC). Age at testing: 6.1-12.9 years. Mean time since completing treatment: 3.3 years. Results were compared to 155 healthy controls. Results. Muscle strength of the survivors was reduced in ankle dorsiflexors on both sides (P < 0.001), wrist dorsiflexors on the non-dominant side (P < 0.001), and pinch grip on the non-dominant (P = 0.001) and dominant side (P = 0.01). Passive ankle dorsiflexion of the survivors was significantly less on both sides (P < 0.01). Movement-ABC percentile score was affected by pinch grip strength on the non-dominant (P < 0.004), and dominant side (P = 0.024) but not by strength of other muscle groups or by passive ankle dorsiflexion. Conclusion. Peripheral muscle strength and ankle dorsiflexion are reduced in the long-term in children treated for cancer with chemotherapy. However, neither decreased muscle strength nor reduced ankle dorsiflexion could completely explain reduced scores on the movement-ABC.

Biology and Treatment of Acute Lymphoblastic Leukemia (Article)

2008-02-01T00:00:00Z

Influence of tumor site and histology on long-term survival in 193 children with extracranial germ cell tumors (Article)

2008-02-01T00:00:00Z

Aims: Although germ cell tumors (GCT) supposedly share the same cell type of origin, their clinical course differs considerably depending on tumor site and histology. The aim of this work was to study long-term survival stratified for tumor site and tumor histology. Materials and Methods: The medical records of 193 consecutive infants and children with extracranial GCT were studied. The GCT arose in the following anatomical sites: sacrococcygeal (n = 70), ovary (n = 66), testis (n = 20), retroperitoneum (n = 12), neck (n = 8), mediastinum (n = 7), and miscellaneous (n = 10). Histological analysis revealed 152 teratomas (mature: 115, immature: 37), 27 yolk sac tumors, 8 mixed tumors, 2 dysgerminomas, 2 gonadoblastomas, 1 choriocarcinoma and 1 embryonal carcinoma.

Prognostic significance of molecular-cytogenetic abnormalities in pediatric T-ALL is not explained by immunophenotypic differences (Article)

2008-01-01T00:00:00Z

Pediatric T-cell acute lymphoblastic leukemia (T-ALL) is characterized by chromosomal rearrangements possibly enforcing arrest at specific development stages. We studied the relationship between molecular-cytogenetic abnormalities and T-cell development stage to investigate whether arrest at specific stages can explain the prognostic significance of specific abnormalities. We extensively studied 72 pediatric T-ALL cases for genetic abnormalities and expression of transcription factors, NOTCH1 mutations and expression of specific CD markers. HOX11 cases were CD1 positive consistent with a cortical stage, but as 4/5 cases lacked cytoplasmatic-β expression, developmental arrest may precede β-selection. HOX11L2 was especially confined to immature and pre-AB developmental stages, but 3/17 HOX11L2 mature cases were restricted to the γδ-lineage. TAL1 rearrangements were restricted to the αβ-lineage with most cases being TCR-αβ positive. NOTCH1 mutations were present in all molecular-cytogenetic subgroups without restriction to a specific developmental stage. CALM-AF10 was associated with early relapse. TAL1 or HOX11L2 rearrangements were associated with trends to good and poor outcomes, respectively. Also cases with high vs low TAL1 expression levels demonstrated a trend toward good outcome. Most cases with lower TAL1 levels were HOX11L2 or CALM-AF10 positive. NOTCH1 mutations did not predict for outcome. Classification into T-cell developmental subgroups was not predictive for outcome.

Inhibin B is superior to FSH as a serum marker for spermatogenesis in men treated for Hodgkin's lymphoma with chemotherapy during childhood (Article)

2007-12-01T00:00:00Z

BACKGROUND: The aim of this study was to evaluate the long-term gonadal sequelae after treatment for childhood Hodgkin's lymphoma with combination chemotherapy, using up to date fertility parameters and andrological evaluation, including for the first time inhibin B. METHODS: There were 56 male patients treated from 1974-1998 for childhood Hodgkin's lymphoma with combination chemotherapy ABVD or EBVD (adriamycin/epirubicin, bleomycin, vinblastine, dacarbazine) with or without MOPP (mechlorethamine, vincristin, prednisone, procarbazine) with the intention to avoid radiotherapy. These men were studied 15.5 years (range 5.6-30.2 years) after cessation of therapy. Serum follicle stimulating hormone (FSH), luteinizing hormone (LH), inhibin B, testosterone, sex hormone-binding globulin (SHBG), sperm concentration and sperm DNA integrity were determined. RESULTS: In men treated with MOPP, median FSH and LH were significantly increased (P < 0.001) and inhibin B (17.5 versus 143 ng/l; P < 0.001) and sperm concentration (1.05 versus 49.5 × 106/ml; P < 0.05) were significantly decreased compared with patients treated without MOPP. The number of MOPP courses was significantly correlated with FSH and inhibin B levels. Only inhibin B showed an independent correlation with sperm concentration (r = 0.86; P < 0.001). CONCLUSIONS: The use of MOPP chemotherapy causes permanent gonadal damage in the far majority of male survivors of childhood Hodgkin's lymphoma and inhibin B is the most valuable serum marker for gonadal function.

High nerve growth factor receptor (p75NTR) expression is a favourable prognostic factor in paediatric B cell precursor-acute lymphoblastic leukaemia (Article)

2007-11-01T00:00:00Z

Nerve growth factor (NGF) plays a pivotal role in cellular survival/death decisions with the low affinity receptor p75NTR predominately transmitting anti-proliferative signals. In spite of its established role in B-cell function and identification as a prognostically favourable marker in a number of malignancies, little is known about the expression pattern and prognostic significance of p75NTR in B cell precursor-acute lymphoblastic leukaemia (BCP-ALL). p75NTR expression was prospectively studied on primary ALL-blasts in a cohort of paediatric patients with common ALL (n = 86) and preB-ALL (n = 34) treated within the Co-operative study group for childhood acute lymphoblastic leukaemia (CoALL) protocol, CoALL06-97. Flow cytometric analysis showed that almost half of the patients expressed no or negligible amounts of p75NTR (<10%). The median expression in patients expressing p75NTR beyond that threshold was 49% (range 11-100%). In patients classified as low-risk at diagnosis, p75NTR expression was significantly higher than in high-risk patients (P = 0.001). Of note, p75NTR expression was lower in the 21 patients who subsequently developed relapse compared with those remaining in remission (P = 0.038). Accordingly, relapse-free survival was significantly better in patients expressing high surface p75NTR (P = 0.041). Thus, in this prospective analysis, high p75NTR expression was a strong prognostic marker that identified a group of paediatric ALL patients with favourable outcome.

Role of mutation independent constitutive activation of FLT3 in juvenile myelomonocytic leukemia (Article)

2007-11-01T00:00:00Z

FLT3 gene mutations have been identified as prognostic factors in myeloid malignancies. Furthermore, FLT3 can be activated by wild type overexpression or ligand-dependent in leukemic cells co-expressing FLT3 ligand (FLT3L). So far no data are available on FLT3/FLT3L expression and activation in JMML. In 51 clinical JMML samples, activating mutations were screened, FLT3 and FLT3L mRNA levels were assessed and the sensitivity of JMML cells to the FLT3 inhibitor PKC412 was tested by MTT assays. No evidence for constitutively activation of FLT3/FLT3L was found in JMML, indicating that FLT3 inhibitors are unlikely to be effective in JMML.

D-HPLC analysis of the entire FLT3 gene in MLL rearranged and hyperdiploid acute lymphoblastic leukemia (Article)

2007-11-01T00:00:00Z

MLL rearranged and hyperdiploid acute lymphoblastic leukemia (ALL) are characterized by high-level FLT3 expression and constitutive FLT3 activation. As known activating FLT3 mutations are often absent in these patients, we screened the entire FLT3 coding sequence in MLL rearranged and hyperdiploid ALL cases for yet unidentified additional genetic alterations using denaturing D-HPLC. Both in MLL rearranged and hyperdiploid ALL we found that a small minority of samples, 7% and 10% respectively, carried genetic alterations. Although some of these alterations may induce FLT3 activation, the majority of these patients carry wild-type FLT3 genes.

Influence of two different regimens of concomitant treatment with asparaginase and dexamethasone on hemostasis in childhood acute lymphoblastic leukemia [11] (Article)

2007-11-01T00:00:00Z

No adverse effect of vincristine on handwriting in children after completion of therapy (Article)

2007-11-01T00:00:00Z

Background. Long-term writing difficulties in children after treatment with vincristine for acute lymphoblastic leukemia, Wilms tumor, B non-Hodgkin lymphoma, and malignant mesenchymal tumors, were investigated. Procedure. Handwriting of 33 survivors and 33 controls matched for age, sex, and grade, was assessed with the BHK-scale. The examiner was blinded for whether a child was a case or a control. Results. No significant difference in writing speed was found. Mean difference in number of letters produced during 5 min was 6.4 (±67.1, range -103 to +169). No significant difference was found in quality of writing scores; mean difference in points was 1.5 (±7.7, range -19 to +22). Cumulative vincristine dose, age at diagnosis or time since completion of treatment did not affect writing speed or quality. Conclusion. Chemotherapy, including vincristine, does not lead to long-term problems in speed or quality of writing in children treated for cancer.

Anti-Müllerian hormone is a sensitive serum marker for gonadal function in women treated for Hodgkin's lymphoma during childhood (Article)

2007-10-01T00:00:00Z

Purpose: The aim of this study was to evaluate the long-term effects of combination chemotherapy treatment for girls with Hodgkin's lymphoma (HL) on gonadal function using anti-Müllerian hormone (AMH) and inhibin B as ovarian reserve parameters. Patients and Methods: LH, FSH, inhibin B, and AMH were measured in 32 women treated from 1974 to 1998 for pediatric HL with chemotherapy, with the intention to avoid radiotherapy. All patients [median age 25.0 yr (range 19.2-40.4 yr)] were in complete remission with a median follow-up time of 14.0 yr (range 5.7-24.5 yr) after therapy. All patients were treated with combination chemotherapy doxorubicin, bleomycin, vinblastine and dacarbazine (ABVD) or EBVD with or without mechlorethamine, vincristine, procarbazine, and prednisone (MOPP). Because of incomplete remission or relapse, involved field radiotherapy was needed in seven of 32 women. Results were compared with a healthy control group. Results: Patients treated with six or more cycles of MOPP combination chemotherapy had significantly higher levels of FSH and lower serum levels of inhibin B and AMH, compared with healthy women [FSH, 17.0 vs. 6.0 U/liter (P < 0.05); inhibin B, 23.0 vs. 112.5 ng/liter (P < 0.01); AMH, 0.39 vs. 2.10 μg/liter (P < 0.01)]. AMH was also significantly lower, compared with women treated without MOPP (median 0.39 vs. 1.40 μg/liter; P = 0.01). Conclusions: Women treated during childhood for HL with MOPP seem to have a distinctly lower ovarian reserve as measured by lower AMH values at early adulthood, compared with healthy women. Moreover, AMH seems to be the only predictor that is sufficiently sensitive to detect this decrease in ovarian reserve. Copyright

Prognostic significance of high-level FLT3 expression in MLL-rearranged infant acute lymphoblastic leukemia [8] (Article)

2007-10-01T00:00:00Z

FBW7 mutations in leukemic cells mediate NOTCH pathway activation and resistance to γ-secretase inhibitors (Article)

2007-08-06T00:00:00Z

γ-secretase inhibitors (GSIs) can block NOTCH receptor signaling in vitro and therefore offer an attractive targeted therapy for tumors dependent on deregulated NOTCH activity. To clarify the basis for GSI resistance in T cell acute lymphoblastic leukemia (T-ALL), we studied T-ALL cell lines with constitutive expression of the NOTCH intracellular domain (NICD), but that lacked C-terminal truncating mutations in NOTCH1. Each of the seven cell lines examined and 7 of 81 (8.6%) primary T-ALL samples harbored either a mutation or homozygous deletion of the gene FBW7, a ubiquitin ligase implicated in NICD turnover. Indeed, we show that FBW7 mutants cannot bind to the NICD and define the phosphodegron region of the NICD required for FBW7 binding. Although the mutant forms of FBW7 were still able to bind to MYC, they do not target it for degradation, suggesting that stabilization of both NICD and its principle downstream target, MYC, may contribute to transformation in leukemias with FBW7 mutations. In addition, we show that all seven leukemic cell lines with FBW7 mutations were resistant to the MRK-003 GSI. Most of these resistant lines also failed to down-regulate the mRNA levels of the NOTCH targets MYC and DELTEX1 after treatment with MRK-003, implying that residual NOTCH signaling in T-ALLs with FBW7 mutations contributes to GSI resistance. JEM

Chemotherapy does not influence intestinal amino acid uptake in children (Article)

2007-08-01T00:00:00Z

Chemotherapy will frequently induce intestinal damage (mucositis). Enteral nutrition is then often withheld for fear of impaired intestinal absorption as shown in animal models. There is no clinical evidence, however, that absorption is indeed compromised during chemotherapy-induced mucositis. The aim of this study was to evaluate systemic availability of dietary amino acids (leucine) during chemotherapy-induced mucositis. We studied eight childhood cancer patients (age 1.5-16 y) on 2 d, i.e. the day before chemotherapy and 3-5 d after. Chemotherapy-induced oral mucositis and diarrhea were scored on a World Health Organization toxicity scale. Stable isotope tracers were used to measure first-pass splanchnic leucine uptake and whole-body leucine kinetics. Patients showed increased mucositis and/or diarrhea toxicity scores (p < 0.0001) after chemotherapy. Systemic availability of enterally administered leucine was not significantly affected by chemotherapy (before 60%, after 90%, p = 0.46). Interestingly, five patients already showed a negative leucine balance before chemotherapy. In conclusion, most children receiving chemotherapy are already catabolic before start of a new cycle of chemotherapy. Amino acid transport as measured by leucine uptake in the intestine is not affected by chemotherapy-induced mucositis.

Alterations in epithelial and mesenchymal intestinal gene expression during doxorubicin-induced mucositis in mice (Article)

2007-08-01T00:00:00Z

In the current study we aimed to gain insight into epithelial-mesenchymal cross-talk and progenitor compartment modulation during doxorubicin (DOX)-induced mucositis in mice. Intestinal segments were collected on various days after DOX treatment. DOX-induced damage at day 1-2 was characterized by increased epithelial proliferation and apoptosis and a decrease in the expression of epithelial differentiation markers. Concurrently, T-cell factor-4 (TCF4) levels increased and the epithelial differentiation enhancing factor, bone morphogenic protein-4 (BMP4), decreased. During severe damage (day 3), BMP4 levels were significantly increased, which inversely correlated with epithelial proliferation. At the same time, the expression of the epithelial differentiation markers was increasing again. At day 7, BMP4 levels were down-regulated, while the levels of the epithelial differentiation markers and TCF4 were normalized again. These data suggest that in response to DOX-induced damage, BMP4 and TCF4 are modulated in such a way that homeostasis of the progenitor compartment is partly preserved.

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

2007-07-21T00:00:00Z

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.

Identification of novel biomarkers in pediatric primitive neuroectodermal tumors and ependymomas by proteome-wide analysis (Article)

2007-06-01T00:00:00Z

The aim of this study was to identify aberrantly expressed proteins in pediatric primitive neuroectodermal tumors (PNETs) and ependymomas. Tumor tissue of 29 PNET and 12 ependymoma patients was subjected to 2-dimensional difference gel electrophoresis. Gel analysis resulted in 79 protein spots being differentially expressed between PNETs and ependymomas (p < 0.01, fold change difference in expression >2). Three proteins, stathmin, annexin A1, and calcyphosine, were chosen for validation by immunohistochemistry. Stathmin was expressed 2.6-fold higher in PNETs than in ependymomas, and annexin A1 and calcyphosine were expressed 2.5- and 37.6-fold higher, respectively, in ependymomas. All PNETs showed strong staining for stathmin, and all ependymomas were strongly positive for annexin A1, whereas control tissues were negative. Calcyphosine immunoreactivity was observed in 59% of the ependymomas and was most profound in ependymoma tissue showing epithelial differentiation. mRNA expression levels of stathmin, annexin A1, and calcyphosine significantly correlated (Rs = 0.65 [p < 0.0001], Rs = 0.50 [p = 0.001], and Rs = 0.72 [p < 0.0001], respectively) with protein expression levels. In conclusion, using a proteome-wide approach, stathmin, annexin A1, and calcyphosine were successfully identified as tumor-specific proteins in pediatric PNETs and ependymomas. Ongoing studies are focused on characterizing the role of these proteins as tumor markers and potential drug targets in pediatric brain tumors.

Duplication of the MYB oncogene in T cell acute lymphoblastic leukemia (Article)

2007-05-01T00:00:00Z

We identified a duplication of the MYB oncogene in 8.4% of individuals with T cell acute lymphoblastic leukemia (T-ALL) and in five T-ALL cell lines. The duplication is associated with a threefold increase in MYB expression, and knockdown of MYB expression initiates T cell differentiation. Our results identify duplication of MYB as an oncogenic event and suggest that MYB could be a therapeutic target in human T-ALL.

Renin-induced hypertension in Wilms tumor patients (Article)

2007-05-01T00:00:00Z

Background. Since the report that hypertension associated with Wilms tumor (WT) may be renin-induced, no larger series than 13 patients have been published. Nevertheless, angiotensin converting enzyme (ACE) inhibitors have become treatment of choice for hypertension in WT patients. The purpose of this study was to investigate the correlation between plasma renin levels and blood pressure in a larger cohort of WT patients. Procedure. In this retrospective, single-center study, data on blood pressure and plasma renin were analyzed in 86 WT patients treated according to the consecutive SIOP protocols 6, 9, 93-01, and 2001. Results. At diagnosis, 47 WT patients suffered from hypertension (55%). In 31 of these patients plasma renin levels were analyzed; increased plasma renin levels were found in 25/31 patients (81%). In contrast, normal plasma renin levels were measured in 8/13 of the patients with a normal blood pressure (P = 0.012). Twenty-eight children received antihypertensive treatment before surgery, in 25 of them blood pressure normalized before surgery. Blood pressure was normal directly after surgery in all patients but 4, in whom blood pressure recovered to normal within a few weeks. Conclusions. This retrospective study shows that hypertension in WT patients is associated with elevated plasma renin levels, indicating that ACE inhibitors may be a good therapeutic option in at least a subset of WT patients with hypertension before nephrectomy.

Protection against chemotherapy induced mucositis by TGF-β2 in childhood cancer patients: Results from a randomized cross-over study (Article)

2007-05-01T00:00:00Z

Background. Mucositis is one of the most frequent and severe side-effect of chemotherapy in childhood-cancer patients for which there is no prophylaxis available. The efficacy and feasibility of a TGF-β2-enriched feeding for preventing oral and gastro-intestinal-mucositis in childhood-cancer patients were studied. Procedure. The study was designed as a two-period cross-over, randomized, double-blinded, placebo, controlled trial. Patients who had a high risk for developing mucositis and who would receive two comparable cycles of chemotherapy were eligible for the study. During one cycle of chemotherapy, TGF-β2-enriched feeding was administered; during the other, a 'placebo' (not enriched) feeding was used. WHO toxicity scales of diarrhea, oral mucositis, fever, anal lesions and nausea/vomiting were scored daily. In addition, the incidence of occurrence of blood cultures, antibiotic therapy, and interventions or diagnostics related to mucositis were measured. Results. The feasibility of the study was good: 83% of the patients completed two cycles and 86% of the study-feeding was effectively consumed. Administration of TGF-β2was safe as serum TGF-β2did not increase, and renal and liver function were not affected during TGF-β2consumption compared to normal feeding. Differences in toxicity, scored during the whole observation period and the number of days with WHO 3/4 toxicity, were not significantly different between cycles with TGF-β2enriched and normal feeding. Conclusions. TGF-β2administration via feeding is well tolerated and safe. Although this study might have had limitations to show potential benefit of TGF-β2, it does not provide evidence that TGF-β2decreases the incidence or degree of mucositis induced by combination chemotherapy in childhood-cancer patients.

Genomewide identification of prednisolone-responsive genes in acute lymphoblastic leukemia cells (Article)

2007-05-01T00:00:00Z

Glucocorticoids are keystone drugs in the treatment of childhood acute lymphoblastic leukemia (ALL). To get more insight in signal transduction pathways involved in glucocorticoid-induced apoptosis, Affymetrix U133A GeneChips were used to identify transcriptionally regulated genes on 3 and 8 hours of prednisolone exposure in leukemic cells of 13 children as compared with nonexposed cells. Following 3 hours of exposure no significant changes in gene expression could be identified. Following 8 hours of exposure, 51 genes were differentially expressed (P < .001 and false discovery rate < 10%) with 39 genes being up-regulated (median, 2.4-fold) and 12 genes were downregulated (median, 1.7-fold). Twenty-one of those genes have not been identified before to be transcriptionally regulated by prednisolone. Two of the 3 most highly up-regulated genes were tumor suppressor genes, that is, thioredoxin-interacting protein (TXNIP; 3.7-fold) and zinc finger and BTB domain containing 16 (ZBTB16; 8.8-fold). About 50% of the differentially expressed genes were functionally categorized in 3 major routes, namely MAPK pathways (9 genes), NF-κB signaling (11 genes), and carbohydrate metabolism (5 genes). Biologic characterization of these genes and pathways might elucidate the action of glucocorticoids in ALL cells, possibly suggesting causes of glucocorticoid resistance and new potential targets for therapy.

Immunobiological diversity in infant acute lymphoblastic leukemia is related to the occurrence and type of MLL gene rearrangement (Article)

2007-04-01T00:00:00Z

The aim of this study was to identify immunobiological subgroups in 133 infant acute lymphoblastic leukemia (ALL) cases as assessed by their immunophenotype, immunoglobulin (Ig) and T-cell receptor (TCR) gene rearrangement pattern, and the presence of mixed lineage leukemia (MLL) rearrangements. About 70% of cases showed the pro-B-ALL immunophenotype, whereas the remaining cases were common ALL and pre-B-ALL. MLL translocations were found in 79% of infants, involving MLL-AF4 (41%), MLL-ENL (18%), MLL-AF9 (11%) or another MLL partner gene (10%). Detailed analysis of Ig/TCR rearrangement patterns revealed IGH, IGK and IGL rearrangements in 91, 21 and 13% of infants, respectively. Cross-lineage TCRD, TCRG and TCRB rearrangements were found in 46, 17 and 10% of cases, respectively. As compared to childhood precursor-B-ALL, Ig/TCR rearrangements in infant ALL were less frequent and more oligoclonal. MLL-AF4 and MLL-ENL-positive infants demonstrated immature rearrangements, whereas in MLL-AF9-positive leukemias more mature rearrangements predominated. The immature Ig/TCR pattern in infant ALL correlated with young age at diagnosis, CD10 negativity and predominantly with the presence and the type of MLL translocation. The high frequency of immature and oligoclonal Ig/TCR rearrangements is probably caused by early (prenatal) oncogenic transformation in immature B-lineage progenitor cells with germline Ig/TCR genes combined with a short latency period.

The clinical value of follow-up examinations in childhood T-cell acute lymphoblastic leukemia and T-cell non-Hodgkin lymphoma (Article)

2007-04-01T00:00:00Z

Background. The aim of this study was to evaluate the value of follow-up investigations of T-cell acute lymphoblastic leukemia (T-ALL) and T-cell non-Hodgkin's lymphoma (T-NHL), including cerebrospinal fluid (CSF) examination, bone marrow (BM) aspiration, peripheral blood (PB) count, serum lactate dehydrogenase (LDH) and chest X-rays in patients with an initial mediastinal enlargement. Procedure. We reviewed clinical records of all T-ALL patients from 1987 to 2002 and all T-NHL patients from 1977 to 2002, seen at a single institution. Results. Of 48 T-ALL patients, 15 suffered from a relapse, 6 (40%) were asymptomatic at the time of relapse. T-ALL (13/30) with mediastinal enlargement at first diagnosis relapsed versus 2/16 of those without mediastinal enlargement. However, at relapse, only one patient had a mediastinal mass, which in addition was symptomatic. Of 39 T-NHL patients, 6 patients relapsed. Forty percent of relapsed T-ALL and 17% of relapsed T-NHL were asymptomatic. The seven asymptomatic relapses were detected by CSF (n = 4), BM (n = 2) or blood count (n = 1) examinations. All T-ALL and T-NHL patients with a mediastinal relapse were symptomatic. Conclusions. This study suggests that routine CSF examinations during treatment can detect relapses of T-ALL and T-NHL before onset of symptoms, which might be of clinical value. Relapses are rarely detected by BM or blood examinations and whether this translates in a clinical benefit is unlikely. Routine chest X-rays are not useful.

Methotrexate-induced mucositis in mucin 2-deficient mice (Article)

2007-01-01T00:00:00Z

The mucin Muc2 or Mycin2 (Muc2), which is the main structural component of the protective mucus layer, has shown to be upregulated during chemotherapy-induced mucositis. As Muc2 has shown to have protective capacities, upregulation of Muc2 may be a counter reaction of the intestine protecting against mucositis. Therefore, increasing Muc2 protein levels could be a therapeutic target in mucositis prevention or reduction. Our aim was to determine the role of Muc2 in chemotherapy-induced mucositis. Mucositis was induced in Muc2 knockout (Muc2-/-) and wild type (Muc2+/+) mice by injecting methotrexate (MTX). Animals were weighed and sacrificed on Days 2-6 after MTX treatment and jejunal segments were analyzed. Before MTX treatment, the small intestine of Muc2+/+and Muc2-/-mice were similar with respect to epithelial morphology and proliferation. Moreover, sucrase-isomaltase and trefoil factor-3 protein expression levels were comparable between Muc2+/+and Muc2-/-mice. Up to Day 3 after MTX treatment, percentages of weight-loss did not differ. Thereafter, Muc2+/+mice showed a trend towards regaining weight, whereas Muc2-/-mice continued to lose weight. Surprisingly, MTX-induced intestinal damage of Muc2-/-and Muc2+/+mice was comparable. Prior to MTX-injection, tumor necrosis factor-α and interleukin-10 mRNAs were upregulated in Muc2-/-mice, probably due to continuous exposure of the intestine to luminal antigens. Muc2 deficiency does not lead to an increase in chemotherapy-induced mucositis. A possible explanation is the mechanism by which Muc2 deficiency may trigger the immune system to release interleukin-10, an anti-inflammatory cytokine before MTX-treatment.

Medical end-of-life decisions for children in the Netherlands. (Article)

2005-09-01T00:00:00Z

BACKGROUND: Most end-of-life decision-making studies have, until now, involved either the general population or newborn infants. OBJECTIVE: To assess the frequency of end-of-life decisions preceding child death and the characteristics of the decision-making process in the Netherlands. METHODS: Two studies were performed. The first was a death certificate study in which all 129 physicians reporting the death of a child aged between 1 and 17 years in the period August to December 2001 received a written questionnaire; the second was an interview study in which face-to-face interviews were held with 63 physicians working in pediatric hospital departments. RESULTS: Some 36% of all deaths of children between the ages of 1 and 17 years during the relevant period were preceded by an end-of-life decision: 12% by a decision to refrain from potentially life-prolonging treatment; 21% by the alleviation of pain or symptoms with a possible life-shortening effect; and 2.7% by the use of drugs with the explicit intention of hastening death. The latter decision was made at the child's request in 0.7% and at the request of the family in 2% of cases. The interview study examined 76 cases of end-of-life decision making. End-of-life decisions were discussed with all 9 competent and 3 partly competent children, with the parents in all cases, with other physicians in 75 cases, and with nurses in 66 cases. CONCLUSIONS: While not inconsiderable, the percentage of end-of-life decisions was lower for children than for adults and newborn infants. Most children are not considered to be able to participate in the decision-making process. Decisions are generally discussed with parents and other caregivers and, if possible, with the child.

Genetic variations in the glucocorticoid receptor gene are not related to glucocorticoid resistance in childhood acute lymphoblastic leukemia. (Article)

2005-08-15T00:00:00Z

Glucocorticoid sensitivity is an important prognostic factor in pediatric acute lymphoblastic leukemia (ALL). For its antileukemic effect, glucocorticoid binds the intracellular glucocorticoid receptor (GR) subsequently regulating transcription of downstream genes. We analyzed whether genetic variations within the GR gene are related to differences in the cellular response to glucocorticoids. METHODS: In leukemic samples of 57 children, the GR gene was screened for nucleotide variations using a PCR/single-strand conformational polymorphism sequencing strategy. Data were linked to in vivo and in vitro glucocorticoid resistance. RESULTS: No somatic mutations were detected in the GR gene coding region, but six polymorphisms (i.e., ER22/23EK, N363S, BclI, intron mutation 16 bp upstream of exon 5, H588H, and N766N) were identified. In 67% of ALL cases, at least one minor allele of these polymorphisms was detected. Although only borderline significant, the incidence for the N363S polymorphism minor allele was higher (12% versus 6%, P = 0.06) and for the ER22/23EK minor allele lower (4% versus 7.6%, P = 0.1) than in a healthy, comparable population. The different genotypes of the polymorphisms were not related to prednisone resistance. In conclusion, polymorphisms but not somatic mutations in the GR gene coding region occur in leukemic blasts of children with ALL. Our data suggest that these genetic variations are not a major contributor for differences in cellular response to glucocorticoids in childhood ALL. The higher incidence of the N363S minor allele and the lower incidence of the ER22/23EK minor allele in our ALL population as compared with a normal population warrants further research.

Expression levels of TEL, AML1, and the fusion products TEL-AML1 and AML1- TEL versus drug sensitivity and clinical outcome in t(12;21)-positive pediatric acute lymphoblastic leukemia. (Article)

2005-04-15T00:00:00Z

PURPOSE: t(12;21)(p13; q22), present in approximately 25% of pediatric precursor B-ALL, is highly sensitivity to L-asparaginase and the prognosis depends on the intensity of the treatment protocol. This study analyzes the relationship between the mRNA expression of the genes and fusion products involved in t(12;21), in vitro sensitivity to prednisolone, vincristine, and L-asparaginase, and long-term clinical outcome in t(12;21)+ acute lymphoblastic leukemia (ALL) patients. EXPERIMENTAL DESIGN: Long-term clinical outcome in 45 t(12;21)+ ALL patients was related to mRNA expression of TEL, AML1, TEL-AML1, and AML1-TEL, determined by real-time quantitative PCR, and the in vitro sensitivity to prednisolone, vincristine, and L-asparaginase, using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays. RESULTS: A significant approximately 3.5-fold lower TEL expression in t(12;21)+ compared with t(12;21)- ALL samples (P = 0.006) and normal controls (P = 0.004) was found. Expression of AML1 did not differ between t(12;21)+ and t(12;21)- ALL. However, AML1 expression in the leukemic cells was 2-fold higher compared with normal controls (P = 0.02). The TEL-AML1 fusion product was expressed in all t(12;21)+ cases, whereas the reciprocal fusion product AML1-TEL was expressed in only 76%. High expression levels of TEL-AML1 [hazard ratio (HR), 1.3; 95% confidence interval (95% CI), 1.10-1.57; P = 0.003], AML1-TEL (HR, 4.9; 95% CI, 1.99-12.40; P = 0.001) and AML1 (HR, 1.1; 95% CI, 1.03-1.22; P = 0.006) were associated with a poor long-term clinical outcome within t(12;21)+ ALL. Cellular drug resistance towards prednisolone, vincristine, and L-asparaginase could not explain this predictive value. Multivariate analysis including age and WBC showed that only high AML1-TEL expression is an independent poor prognostic factor in t(12;21)+ childhood ALL. CONCLUSION: High AML1-TEL expression is an independent poor prognostic factor in t(12;21)+ childhood ALL.

Effect of polymorphisms in folate-related genes on in vitro methotrexate sensitivity in pediatric acute lymphoblastic leukemia (Article)

2005-01-01T00:00:00Z

We studied whether common polymorphisms in genes involved in folate metabolism affect methotrexate (MTX) sensitivity. Ex vivo MTX sensitivity of lymphoblasts obtained from pediatric patients with acute lymphoblastic leukemia (ALL; n = 157) was determined by the in situ thymidylate synthase inhibition assay after either continuous (21 hours; TSI(50, cont)) or short-term (3 hours; TSI(50, short)) MTX exposure. DNA was isolated from lymphoblasts obtained from cytospin slides. Polymorphisms in methylenetetrahydrofolate reductase (MTHFR 677C>T, MTHFR 1298A>C), methionine synthase (MTR 2756A>G), methionine synthase reductase (MTRR 66A>G), methylenetetrahydrofolate dehydrogenase (MTHFD1 1958G>A), serine hydroxymethyl transferase (SHMT1 1420C>T), thymidylate synthase (TS 2R3R), and the reduced folate carrier (RFC 80G>A) were detected by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) or real-time PCR. Patients with the MTHFR 1298AC variant or the MTRR 66 G-allele showed decreased in vitro MTX sensitivity measured under both test conditions. SHMT1 1420TT homozygotes only showed decreased MTX sensitivity in the TSI(50, cont). In conclusion, polymorphisms in the folate-related genes MTHFR, MTRR, and SHMT1 are related to MTX resistance in pediatric patients with ALL.

Asparagine synthetase expression is linked with L-asparaginase resistance in TEL-AML1-negative but not TEL-AML1-positive pediatric acute lymphoblastic leukemia (Article)

2005-01-01T00:00:00Z

Resistance to L-asparaginase in leukemic cells may be caused by an elevated cellular expression of asparagine synthetase (AS). Previously, we reported that high AS expression did not correlate to L-asparaginase resistance in TEL-AML1-positive B-lineage acute lymphoblastic leukemia (ALL). In the present study we confirmed this finding in TEL-AML1-positive patients (n = 28) using microarrays. In contrast, 35 L-asparaginase-resistant TEL-AML1-negative B-lineage ALL patients had a significant 3.5-fold higher AS expression than 43 sensitive patients (P < .001). Using real-time quantitative polymerase chain reaction (RTQ-PCR), this finding was confirmed in an independent group of 39 TEL-AML1-negative B-lineage ALL patients (P = .03). High expression of AS was associated with poor prognosis (4-year probability of disease-free survival [pDFS] 58% +/- 11%) compared with low expression (4-year pDFS 83% +/- 7%; P = .009). We conclude that resistance to l-asparaginase and relapse risk are associated with high expression of AS in TEL-AML1-negative but not TEL-AML1-positive B-lineage ALL.

Targeting FLT3 in primary MLL-gene-rearranged infant acute lymphoblastic leukemia (Article)

2005-01-01T00:00:00Z

Acute lymphoblastic leukemia (ALL) in infants is characterized by rearrangements of the mixed lineage leukemia (MLL) gene, drug resistance, and a poor treatment outcome. Therefore, novel therapeutic strategies are needed to improve prognosis. Recently, we showed that FLT3 is highly expressed in MLL rearranged ALL (MLL). Here we demonstrate FLT3 expression in infants with MLL (n = 41) to be significantly higher compared to both infant (n = 8; P < .001) and noninfant patients with ALL (n = 23; P = .001) carrying germline MLL genes. Furthermore, leukemic cells from infants with MLL were significantly more sensitive to the Fms-like tyrosine kinase 3 (FLT3) inhibitor PKC412 (N-benzoyl staurosporine) than noninfant ALL cells, and at least as sensitive as internal tandem duplication-positive (ITD+) AML cells. Surprisingly, activation loop mutations only occurred in about 3% (1 of 36) of the cases and no FLT3/ITDs were observed. However, measuring FLT3 phosphorylation in infants with MLL expressing varying levels of wild-type FLT3 revealed that high-level FLT3 expression is associated with ligand-independent FLT3 activation. This suggests that infant MLL cells displaying activated FLT3 as a result of overexpression can be targeted by FLT3 inhibitors such as PKC412. However, at concentrations of PKC412 minimally required to fully inhibit FLT3 phosphorylation, the cytotoxic effects were only fractional. Thus, PKC412-induced apoptosis in infant MLL cells is unlikely to be a consequence of FLT3 inhibition alone but may involve inhibition of multiple other kinases by this drug.

Decreased PARP and procaspase-2 protein levels are associated with cellular drug resistance in childhood acute lymphoblastic leukemia (Article)

2005-01-01T00:00:00Z

Drug resistance in childhood acute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML) is associated with impaired ability to induce apoptosis. To elucidate causes of apoptotic defects, we studied the protein expression of Apaf-1, procaspases-2, -3, -6, -7, -8, -10, and poly(adenosine diphosphate [ADP]-ribose) polymerase (PARP) in cells from children with acute lymphoblastic leukemia (ALL; n = 43) and acute myeloid leukemia (AML; n = 10). PARP expression was present in all B-lineage samples, but absent in 4 of 15 T-lineage ALL samples and 3 of 10 AML cases, which was not caused by genomic deletions. PARP expression was a median 7-fold lower in T-lineage ALL (P < .001) and 10-fold lower in AML (P < .001) compared with B-lineage ALL. PARP expression was 4-fold lower in prednisolone, vincristine and L-asparaginase (PVA)-resistant compared with PVA-sensitive ALL patients (P < .001). Procaspase-2 expression was 3-fold lower in T-lineage ALL (P = .022) and AML (P = .014) compared with B-lineage ALL. In addition, procaspase-2 expression was 2-fold lower in PVA-resistant compared to PVA-sensitive ALL patients (P = .042). No relation between apoptotic protease-activating factor 1 (Apaf-1), procaspases-3, -6, -7, -8, -10, and drug resistance was found. In conclusion, low baseline expression of PARP and procaspase-2 is related to cellular drug resistance in childhood acute lymphoblastic leukemia.

Gene-expression patterns in drug-resistant acute lymphoblastic leukemia cells and response to treatment (Article)

2004-01-01T00:00:00Z

BACKGROUND: Childhood acute lymphoblastic leukemia (ALL) is curable with chemotherapy in approximately 80 percent of patients. However, the cause of treatment failure in the remaining 20 percent of patients is largely unknown. METHODS: We tested leukemia cells from 173 children for sensitivity in vitro to prednisolone, vincristine, asparaginase, and daunorubicin. The cells were then subjected to an assessment of gene expression with the use of 14,500 probe sets to identify differentially expressed genes in drug-sensitive and drug-resistant ALL. Gene-expression patterns that differed according to sensitivity or resistance to the four drugs were compared with treatment outcome in the original 173 patients and an independent cohort of 98 children treated with the same drugs at another institution. RESULTS: We identified sets of differentially expressed genes in B-lineage ALL that were sensitive or resistant to prednisolone (33 genes), vincristine (40 genes), asparaginase (35 genes), or daunorubicin (20 genes). A combined gene-expression score of resistance to the four drugs, as compared with sensitivity to the four, was significantly and independently related to treatment outcome in a multivariate analysis (hazard ratio for relapse, 3.0; P=0.027). Results were confirmed in an independent population of patients treated with the same medications (hazard ratio for relapse, 11.85; P=0.019). Of the 124 genes identified, 121 have not previously been associated with resistance to the four drugs we tested. CONCLUSIONS: Differential expression of a relatively small number of genes is associated with drug resistance and treatment outcome in childhood ALL.

Resistance to different classes of drugs is associated with impaired apoptosis in childhood acute lymphoblastic leukemia (Article)

2003-01-01T00:00:00Z

Resistance of leukemic cells to chemotherapeutic agents is associated with an unfavorable outcome in pediatric acute lymphoblastic leukemia (ALL). To investigate the underlying mechanisms of cellular drug resistance, the activation of various apoptotic parameters in leukemic cells from 50 children with ALL was studied after in vitro exposure with 4 important drugs in ALL therapy (prednisolone, vincristine, l-asparaginase, and daunorubicin). Exposure to each drug resulted in early induction of phosphatidylserine (PS) externalization and mitochondrial transmembrane (Deltapsim) depolarization followed by caspase-3 activation and poly(ADP-ribose) polymerase (PARP) inactivation in the majority of patients. For all 4 drugs, a significant inverse correlation was found between cellular drug resistance and (1) the percentage of cells with PS externalization (<.001 < P <.008) and (2) the percentage of cells with Deltapsim depolarization (.002 < P <.02). However, the percentage of cells with caspase-3 activation and the percentage of cells with PARP inactivation showed a significant inverse correlation with cellular resistance for prednisolone (P =.001; P =.001) and l-asparaginase (P =.01; P =.001) only. This suggests that caspase-3 activation and PARP inactivation are not essential for vincristine- and daunorubicin-induced apoptosis. In conclusion, resistance to 4 unrelated drugs is associated with defect(s) upstream or at the level of PS externalization and Deltapsim depolarization. This leads to decreased activation of apoptotic parameters in resistant cases of pediatric ALL

Differential mRNA expression of Ara-C-metabolizing enzymes explains Ara-C sensitivity in MLL gene-rearranged infant acute lymphoblastic leukemia (Article)

2003-01-01T00:00:00Z

Infant acute lymphoblastic leukemia (ALL) is characterized by a high incidence of mixed lineage leukemia (MLL) gene rearrangements, a poor outcome, and resistance to chemotherapeutic drugs. One exception is cytosine arabinoside (Ara-C), to which infant ALL cells are highly sensitive. To investigate the mechanism underlying Ara-C sensitivity in infants with ALL, mRNA levels of Ara-C-metabolizing enzymes were measured in infants (n = 18) and older children (noninfants) with ALL (n = 24). In the present study, infant ALL cells were 3.3-fold more sensitive to Ara-C (P =.007) and accumulated 2.3-fold more Ara-CTP (P =.011) upon exposure to Ara-C, compared with older children with ALL. Real-time quantitative reverse trancriptase-polymerase chain reaction (RT-PCR) (TaqMan) revealed that infants express 2-fold less of the Ara-C phosphorylating enzyme deoxycytidine kinase (dCK) mRNA (P =.026) but 2.5-fold more mRNA of the equilibrative nucleoside transporter 1 (hENT1), responsible for Ara-C membrane transport (P =.001). The mRNA expression of pyrimidine nucleotidase I (PN-I), cytidine deaminase (CDA), and deoxycytidylate deaminase (dCMPD) did not differ significantly between both groups. hENT1 mRNA expression inversely correlated with in vitro resistance to Ara-C (r(s) = -0.58, P =.006). The same differences concerning dCK and hENT1 mRNA expression were observed between MLL gene-rearranged (n = 14) and germ line MLL cases (n = 25). An oligonucleotide microarray screen (Affymetrix) comparing patients with MLL gene-rearranged ALL with those with nonrearranged ALL also showed a 1.9-fold lower dCK (P =.001) and a 2.7-fold higher hENT1 (P =.046) mRNA expression in patients with MLL gene-rearranged ALL. We conclude that an elevated expression of hENT1, which transports Ara-C across the cell membrane, contributes to Ara-C sensitivity in MLL gene-rearranged infant ALL.

Sensitivity to L-asparaginase is not associated with expression levels of asparagine synthetase in t(12;21)+ pediatric ALL (Article)

2003-01-01T00:00:00Z

The (12;21) translocation resulting in TEL/AML1 gene fusion is present in about 25% of childhood precursor B-lineage acute lymphoblastic leukemia (ALL) and is associated with a good prognosis and a high cellular sensitivity to L-asparaginase (L-Asp). ALL cells are thought to be sensitive to L-Asp due to lower asparagine synthetase (AS) levels. Resistance to L-Asp may be caused by an elevated cellular level of AS or by the ability of resistant cells to rapidly induce the expression of the AS gene on L-Asp exposure. AS may be a target regulated by t(12;21). We studied the relationship between t(12;21) and the mRNA level of AS to investigate a possible mechanism underlying L-Asp sensitivity. Real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR) analysis surprisingly revealed that 30 patients positive for t(12;21) expressed 5-fold more AS mRNA compared with 17 patients negative for t(12;21) (P =.008) and 11 samples from healthy controls (P =.016). The mRNA levels of AS between t(12;21)(-) ALL and healthy controls did not differ. No difference was found between ALL patients positive or negative for t(12;21) in the capacity to up-regulate AS after in vitro L-Asp exposure, excluding a defective capacity for t(12;21) cells in up-regulating AS on L-Asp exposure. Moreover, no correlation was observed between AS mRNA expression and sensitivity to L-Asp. We conclude that the sensitivity of t(12;21)(+) childhood ALL to L-Asp is not associated with the expression level of the AS gene. Furthermore, we contradict the general thought that leukemic cells specifically lack AS compared with normal bone marrow and blood cells.

Kwaad bloed: van jonge mensen, de dingen die voorbijgaan (Inaugural Lecture)

2001-04-20T00:00:00Z

MDR1 gene-related clonal selection and P-glycoprotein function and expression in relapsed or refractory acute myeloid leukemia (Article)

2001-01-01T00:00:00Z

The expression of P-glycoprotein (P-gp), encoded by the MDR1 gene, is an independent adverse prognostic factor for response and survival in de novo acute myeloid leukemia (AML). Little is known about MDR1 expression during the development of disease. The present study investigated whether MDR1 gene- related clonal selection occurs in the development from diagnosis to relapsed AML, using a genetic polymorphism of the MDR1 gene at position 2677. Expression and function of P-gp were studied using monoclonal antibodies MRK16 and UIC2 and the Rhodamine 123 retention assay with or without PSC 833. No difference was found in the levels of P-gp function and expression between diagnosis and relapse in purified paired blast samples from 30 patients with AML. Thirteen patients were homozygous for the genetic polymorphism of MDR1 (n = 7 for guanine, n = 6 for thymidine), whereas 17 patients were heterozygous (GT). In the heterozygous patients, no selective loss of one allele was observed at relapse. Homozygosity for the MDR1 gene (GG or TT) was associated with shorter relapse-free intervals (P =.002) and poor survival rates (P =.02), compared with heterozygous patients. No difference was found in P-gp expression or function in patients with AML with either of the allelic variants of the MDR1 gene. It was concluded that P-gp function or expression is not upregulated at relapse/refractory disease and expression of one of the allelic variants is not associated with altered P-gp expression or function in AML, consistent with the fact that MDR1 gene-related clonal selection does not occur when AML evolves to recurrent disease. (Blood. 2001;97:3605-3611)