CNS-3 status remains an independent adverse prognosis factor in children with acute lymphoblastic leukemia (ALL) treated without cranial irradiation: Results of EORTC Children Leukemia Group study 58951.

AIM: To evaluate the prognostic significance of initial central nervous system (CNS) involvement of children with acute lymphoblastic leukemia (ALL) enrolled in the EORTC 58951 trial. PATIENTS AND METHODS: From 1998 to 2008, 1930 ALL patients were included in the randomized EORTC 58951 trial. Overall treatment intensity was adjusted according to known prognostic factors including the level of minimal residual disease after induction treatment. CNS-directed therapy comprised four to 11 courses of i.v. methotrexate (5g/m2), and 10 to 19 intrathecal chemotherapy injections, depending on risk group and CNS status. Cranial irradiation was omitted for all patients. RESULTS: The overall 8-year event-free survival (EFS) and overall survival (OS) rates were 81.3% and 88.1%, respectively. In the CNS-1, TPL+, CNS-2, and CNS-3 groups, the 8-year EFS rates were 82.1%, 77.1%, 78.3%, and 57.4%, respectively. Multivariable analysis indicated that initial CNS-3 status, but not CNS-2 or TLP+, was an independent adverse predictor of outcome. The 8-year incidence of isolated CNS relapse was 1.7% and of isolated or combined CNS relapse it was 3.7%. NCI high-risk group, male sex, CNS-2 and CNS-3 status were independent predictors for a higher incidence of any CNS relapse. CONCLUSIONS: CNS-3 status remains associated with poor prognosis and requires intensification of both systemic and CNS-directed therapy. This trial was registered at https://clinicaltrials.gov/under/NCT00003728.

Not all pediatric intestinal polyps are alike.

BACKGROUND/AIMS: In childhood, clinical presentation of intes- tinal polyps is variable. Painless rectal red blood loss is the most common presenting sign. Most polyps are sporadic, isolated and benign. However, it is important to correctly identify exceptions. Rare inherited polyposis syndromes need to be recognized because of their increased risk of intestinal and extra-intestinal malignancies. Furthermore, a correct diagnosis and treatment of rare gastro-intestinal malignancies is crucial. METHODS: Between 2016 and 2018 we encountered 4 different types of intestinal polyps. A database search was performed and patient files were checked for clinical manifestations and histo- pathology. Literature was searched to recapitulate red flags for these syndromes, probability of underlying genetic disorders and diagnostic criteria. RESULTS: Between 2016 and 2018, 28 patients presented at the Ghent University Hospital with 30 juvenile polyps. Furthermore, we diagnosed juvenile polyposis syndrome, Li Fraumeni syndrome and familial adenomatous polyposis (FAP) in 1 patient each, whilst 2 FAP patients were in follow-up. Each of these diagnoses has a different lifetime risk of (extra)-intestinal malignancy and requires a different approach and follow-up. Histopathology and genetic testing play an important role in identifying these syndromes in pediatric patients. CONCLUSION: Although most intestinal polyps in childhood are benign juvenile polyps that require no follow-up, rare inherited syndromes should be considered and correctly diagnosed since adequate follow-up is necessary to reduce morbidity and mortality from both gastrointestinal and extraintestinal complications and malignancies.

A Semi-Mechanistic Population Pharmacokinetic/Pharmacodynamic Model of Bortezomib in Pediatric Patients with Relapsed/Refractory Acute Lymphoblastic Leukemia.

INTRODUCTION: The pharmacokinetics (PK) of the 20S proteasome inhibitor bortezomib are characterized by a large volume of distribution and a rapid decline in plasma concentrations within the first hour after administration. An increase in exposure was observed in the second week of treatment, which has previously been explained by extensive binding of bortezomib to proteasome in erythrocytes and peripheral tissues. We characterized the nonlinear population PK and pharmacodynamics (PD) of bortezomib in children with acute lymphoblastic leukemia. METHODS: Overall, 323 samples from 28 patients were available from a pediatric clinical study investigating bortezomib at an intravenous dose of 1.3 mg/m2 twice weekly (Dutch Trial Registry number 1881/ITCC021). A semi-physiological PK model for bortezomib was first developed; the PK were linked to the decrease in 20S proteasome activity in the final PK/PD model. RESULTS: The plasma PK data were adequately described using a two-compartment model with linear elimination. Increased concentrations were observed in week 2 compared with week 1, which was described using a Langmuir binding model. The decrease in 20S proteasome activity was best described by a direct effect model with a sigmoidal maximal inhibitory effect, representing the relationship between plasma concentrations and effect. The maximal inhibitory effect was 0.696 pmol AMC/s/mg protein (95% confidence interval 0.664-0.728) after administration. CONCLUSION: The semi-physiological model adequately described the nonlinear PK and PD of bortezomib in plasma. This model can be used to further optimize dosing of bortezomib.

A new recurrent inversion, inv(7)(p15q34), leads to transcriptional activation of HOXA10 and HOXA11 in a subset of T-cell acute lymphoblastic leukemias.

Chromosomal translocations with breakpoints in T-cell receptor (TCR) genes are recurrent in T-cell malignancies. These translocations involve the TCRalphadelta gene (14q11), the TCRbeta gene (7q34) and to a lesser extent the TCRgamma gene at chromosomal band 7p14 and juxtapose T-cell oncogenes next to TCR regulatory sequences leading to deregulated expression of those oncogenes. Here, we describe a new recurrent chromosomal inversion of chromosome 7, inv(7)(p15q34), in a subset of patients with T-cell acute lymphoblastic leukemia characterized by CD2 negative and CD4 positive, CD8 negative blasts. This rearrangement juxtaposes the distal part of the HOXA gene cluster on 7p15 to the TCRbeta locus on 7q34. Real time quantitative PCR analysis for all HOXA genes revealed high levels of HOXA10 and HOXA11 expression in all inv(7) positive cases. This is the first report of a recurrent chromosome rearrangement targeting the HOXA gene cluster in T-cell malignancies resulting in deregulated HOXA gene expression (particularly HOXA10 and HOXA11) and is in keeping with a previous report suggesting HOXA deregulation in MLL-rearranged T- and B cell lymphoblastic leukemia as the key factor in leukaemic transformation. Finally, our observation also supports the previous suggested role of HOXA10 and HOXA11 in normal thymocyte development.

[The prognostic significance of P-glycoprotein in children with acute lymphoblastic leukemia and neuroblastoma]. / Het prognostisch belang van P-glycoproteine bij kinderen met acute lymfoblasten leukemie en neuroblastomen.

P-glycoprotein (P-gp), a pump located in the cell membrane, extrudes several clinically important drugs from the cell, and hence causes multidrug resistance (MDR). Reversing MDR is possible by using agents that inhibit the activity of P-gp. In this study, we tried to elucidate the prognostic relevance of P-gp in childhood acute lymphoblastic leukemia (ALL) and neuroblastoma. In a first prospective study in childhood ALL, an immunocytochemical APAAP assay was applied. Children scoring positive had a significantly worse outcome than those with a negative test result. In a second prospective study, the prognostic relevance of P-gp was confirmed, using the combination of immunocytochemistry and a functional flow cytometric assay. The combination of immunocytochemistry with the flow cytometric functional assay is being promoted as the most sensitive and clinically relevant amongst the different techniques. In neuroblastoma, ganglioneuroblastoma and ganglioneuroma, P-gp is detected by different assays: immunocytochemistry, flow cytometric immunological and functional tests and an in vivo imaging technique using 99mTc sestamibi. Immunocytochemistry alone did not provide a prognostic role for P-gp in neuroblastoma. On the contrary, using flow cytometric tests, many neuroblastomas scored positive and a discordance was found between the expression and activity of P-gp. P-gp was found more frequently in low-stage neuroblastoma, differentiated tumours and tumours after chemotherapy. A good correlation between flow cytometric results and imaging results was seen. Consequently, 99mTc sestamibi scintigraphy is not useful as an in vivo predictor of MDR in neuroblastoma. Unlike the findings in childhood ALL, P-gp does not contribute to MDR in neuroblastoma but seems to be a marker of differentiation.

[Detection of minimal residual disease in children with neuroblastoma]. / Detectie van minimale residuele ziekte bij kinderen met neuroblastoom.

The presence of residual cells in bone marrow specimens of stage 4 patients, is an important prognostic parameter in neuroblastoma. Sensitive and specific standardized techniques are a prerequisite to detect and quantify these residual cells. In this respect, we evaluated an immunocytochemical test, a multiparameter flow cytometric assay and quantitative real-time RT-PCR. The latter technique was the most sensitive one in our hands. Although it is a simple and highly reproducible test, RT-PCR does not allow visualisation of neuroblastoma cells. On the contrary, the immunocytochemical APAAP assay is slightly less sensitive but can discriminate neuroblastoma cells morphologically. The combination of both assays seems to be ideal for monitoring residual cells in bone marrow of neuroblastoma patients. The flow cytometric multiparameter assay particularly offers additional information in GD2-negative neuroblastoma. Preliminary results of our study in children with stage 4 neuroblastoma reveals important information for the responsible physician and favours the prognostic role of minimal residual disease in neuroblastoma. Current multicenter trials in neuroblastoma trying to elucidate the relevance of residual cells at different time point during treatment are ongoing and will ultimately result in a consensus about this intruiging prognostic paramater.

MicroRNA-193b-3p acts as a tumor suppressor by targeting the MYB oncogene in T-cell acute lymphoblastic leukemia.

The MYB oncogene is a leucine zipper transcription factor essential for normal and malignant hematopoiesis. In T-cell acute lymphoblastic leukemia (T-ALL), elevated MYB levels can arise directly through T-cell receptor-mediated MYB translocations, genomic MYB duplications or enhanced TAL1 complex binding at the MYB locus or indirectly through the TAL1/miR-223/FBXW7 regulatory axis. In this study, we used an unbiased MYB 3'untranslated region-microRNA (miRNA) library screen and identified 33 putative MYB-targeting miRNAs. Subsequently, transcriptome data from two independent T-ALL cohorts and different subsets of normal T-cells were used to select miRNAs with relevance in the context of normal and malignant T-cell transformation. Hereby, miR-193b-3p was identified as a novel bona fide tumor-suppressor miRNA that targets MYB during malignant T-cell transformation thereby offering an entry point for efficient MYB targeting-oriented therapies for human T-ALL.

IKZF1 deletion is an independent prognostic marker in childhood B-cell precursor acute lymphoblastic leukemia, and distinguishes patients benefiting from pulses during maintenance therapy: results of the EORTC Children's Leukemia Group study 58951.

The added value of IKZF1 gene deletion (IKZF1(del)) as a stratifying criterion in B-cell precursor acute lymphoblastic leukemia (BCP-ALL) is still debated. We performed a comprehensive analysis of the impact of IKZF1(del) in a large cohort of children (n=1223) with BCR-ABL1-negative BCP-ALL treated in the EORTC-CLG trial 58951. Patients with IKZF1(del) had a lower 8-year event-free survival (EFS, 67.7% versus 86.5%; hazard ratio (HR)=2.41; 95% confidence interval (CI)=1.75-3.32; P<0.001). Importantly, despite association with high-risk features such as high minimal residual disease, IKZF1(del) remained significantly predictive in multivariate analyses. Analysis by genetic subtype showed that IKZF1(del) increased risk only in the high hyperdiploid ALLs (HR=2.57; 95% CI=1.19-5.55; P=0.013) and in 'B-other' ALLs, that is, lacking classifying genetic lesions (HR=2.22; 95% CI=1.45-3.39; P<0.001), the latter having then a dramatically low 8-year EFS (56.4; 95% CI=44.6-66.7). Among IKZF1(del)-positive patients randomized for vincristine-steroid pulses during maintenance, those receiving pulses had a significantly higher 8-year EFS (93.3; 95% CI=61.3-99.0 versus 42.1; 95% CI=20.4-62.5). Thus, IKZF1(del) retains independent prognostic significance in the context of current risk-adapted protocols, and is associated with a dismal outcome in 'B-other' ALL. Addition of vincristine-steroid pulses during maintenance may specifically benefit to IKZF1(del) patients in preventing relapses.

Clinical significance of P-glycoprotein (P-gp) expression in childhood acute lymphoblastic leukemia. Results of a 6-year prospective study.

UNLABELLED: P-glycoprotein (P-gp), a cellular drug-efflux pump is thought to be one of the major causes of multidrug resistance in malignancies. Since therapeutic strategies are being developed to circumvent drug resistance by inhibiting P-gp, prospective studies concerning the clinical relevance of P-gp in childhood leukemia are warranted. METHODS: P-gp was studied in 102 consecutive cases of de novo childhood ALL and in 34 relapsed patients. An immunocytochemical technique with two monoclonal antibodies (C219,4E3) was used on bone marrow and blood smears. RESULTS: 12/34 (35%) children were scored positive at relapse compared to 12/102 (12%) children with newly diagnosed ALL (p = 0.006). No correlation between P-gp expression and clinical and hematological parameters was seen. All patients were treated according to the EORTC-CLCG protocols (survival at 5 years = 85%). 20/102 patients relapsed. The mortality rate in the P-gp positive group was significantly worse (Logrank P = 0.009) than in the P-gp negative patients. In the relapsed patient population 10/12 P-gp positive cases experienced an unfavourable outcome compared with 10/22 P-gp negative patients [Risk Ratio 2.21 (0.90-5.45)]. CONCLUSIONS: P-glycoprotein expression in newly diagnosed childhood ALL is an independent prognostic parameter for dismal outcome. P-gp positivity at relapse tends towards an adverse clinical outcome compared to the P-gp negative relapsed population.

Discordance of P-glycoprotein expression and function in acute leukemia.

Since March 1996, 93 consecutive samples of 45 adults and 41 children, suffering from acute leukemia, were evaluated prospectively for P-glycoprotein (P-gp) expression and function by flow cytometry. P-gp antigen expression was determined with the monoclonal antibodies 4E3 and MRK16. Transport function was assessed by measuring the modulating effect of verapamil on the intracellular retention of Rhodamine 123. P-gp positivity at relapse was not significantly more frequently than at initial diagnosis in our study group, neither with the immunologic assay, nor with the functional test. There was no correlation between the results of the immunologic and the functional test. Discordant test results were observed in 11/41 children (12/44 samples) and 14/45 adults (15/49 samples), independent of the type of leukemia. 2/12 and 2/15 samples scored positive with one of the monoclonal antibodies and were functionally inactive. In 10/12 and 13/15 samples however, an efflux pump was active and dependent of verapamil, but without detectable antigen. The functional flow cytometric assay allows evaluation of non-P-gp efflux pumps and, therefore, is more clinically relevant since it may better identify patients who would benefit from the use of P-gp inhibitors.

T-cell receptor Vß skewing frequently occurs in refractory cytopenia of childhood and is associated with an expansion of effector cytotoxic T cells: a prospective study by EWOG-MDS.

Immunosuppressive therapy (IST), consisting of antithymocyte globulin and cyclosporine A, is effective in refractory cytopenia of childhood (RCC), suggesting that, similar to low-grade myelodysplastic syndromes in adult patients, T lymphocytes are involved in suppressing hematopoiesis in a subset of RCC patients. However, the potential role of a T-cell-mediated pathophysiology in RCC remains poorly explored. In a cohort of 92 RCC patients, we prospectively assessed the frequency of T-cell receptor (TCR) ß-chain variable (Vß) domain skewing in bone marrow and peripheral blood by heteroduplex PCR, and analyzed T-cell subsets in peripheral blood by flow cytometry. TCRVß skewing was present in 40% of RCC patients. TCRVß skewing did not correlate with bone marrow cellularity, karyotype, transfusion history, HLA-DR15 or the presence of a PNH clone. In 28 patients treated with IST, TCRVß skewing was not clearly related with treatment response. However, TCRVß skewing did correlate with a disturbed CD4(+)/CD8(+) T-cell ratio, a reduction in naive CD8(+) T cells, an expansion of effector CD8(+) T cells and an increase in activated CD8(+) T cells (defined as HLA-DR(+), CD57(+) or CD56(+)). These data suggest that T lymphocytes contribute to RCC pathogenesis in a proportion of patients, and provide a rationale for treatment with IST in selected patients with RCC.

Methemoglobinemia and hemolytic anemia after rasburicase administration in a child with leukemia.

CASE: We report a case of simultaneous methemoglobinemia and hemolytic anemia, probably related to the use of rasburicase, in a child starting treatment for acute lymphoblastic leukemia (ALL). In addition, the patient developed symptoms of pancreatitis. All complications resolved after 3 days of supportive therapy. CONCLUSION: Although usually well tolerated in pediatrics, physicians prescribing rasburicase should be aware of these possible life-threatening adverse reactions.

Methemoglobinemia and hemolytic anemia after rasburicase administration in a child with leukemia.

CASE: We report a case of simultaneous methemoglobinemia and hemolytic anemia, probably related to the use of rasburicase, in a child starting treatment for acute lymphoblastic leukemia (ALL). In addition, the patient developed symptoms of pancreatitis. All complications resolved after 3 days of supportive therapy. CONCLUSION: Although usually well tolerated in pediatrics, physicians prescribing rasburicase should be aware of these possible life-threatening adverse reactions.

The use of propranolol in the treatment of periocular infantile haemangiomas: a review.

Infantile haemangiomas (IH) are benign vascular tumours characterised by their very rapid growth. Although usually innocuous, periocular IH can cause serious visual loss through induction of strabismic, deprivational or anisometropic astigmatism. Common treatment modalities for these IH include intralesional and systemic oral steroids; however, both treatments are associated with potentially severe side effects. A report was published recently demonstrating the impressive effect of propranolol in the treatment of IH. This exciting finding has provoked a paradigm shift in the management of this condition. So far little has been reported in the specific ophthalmologic literature, although case reports are emerging. This review gives an overview of the recent findings and includes the authors' experience with 10 patients treated with propranol.

Surveillance of cytomegalovirus (CMV) DNAemia in pediatric allogeneic stem cell transplantation: incidence and outcome of CMV infection and disease.

Cytomegalovirus (CMV) remains a serious problem after hematopoietic stem cell transplantation (HSCT). To investigate the incidence of CMV infection and outcome we retrospectively analyzed 70 consecutive pediatric allogeneic HSCTs monitored by CMV polymerase chain reaction (PCR), with at least 1-year follow-up or until death. All patients at risk for CMV infection (CMV-seropositive patients and CMV-seronegative recipients transplanted from CMV-seropositive donors) received hyperimmune anti-CMV globulins whereas in the group of HSCT patients with both donor and recipient CMV negativity, polyvalent immunoglobulins were given, both at a dose of 400 mg/kg. All patients received acyclovir at prophylactic doses for at least 6 months. Patients were monitored twice a week by CMV PCR. Patients with 2 positive results for CMV DNAemia received ganciclovir for 14 days and continued until 2 consecutive negative results were obtained. The incidence of CMV DNAemia was 12.8% (9/70) in the whole group, with significant higher risk for patients with CMV-seropositive recipient status, 8 out of 22 (36%), vs. patients with seronegative status, 1 out of 48 (2%) (P=0.0002). Three out of 9 patients with DNAemia developed CMV disease despite adequate preemptive treatment. The transplant-related mortality was higher in the CMV-seropositive recipient group (P=0.05). Age, use of hyperimmune anti-CMV globulins at a high dose, and the low incidence of graft-versus-host disease might be contributing factors to this low incidence.

Expression of the MDR1 gene product P-glycoprotein in childhood neuroblastoma.

BACKGROUND: In cancer treated with chemotherapy, multidrug resistance is characterized by increased genetic expression of P-glycoprotein (P-gp), which acts as an ATP-dependent drug-efflux pump. However, the clinical significance of the expression of the multidrug resistance gene (MDR1) product P-gp in neuroblastoma (NB) is still a matter of debate. In this study, the role of the expression of P-gp in NB was evaluated. METHODS: NB tumor imprints and NB positive bone marrow smears from 23 children before and after multidrug chemotherapy were examined for P-gp expression by antialkaline phosphatase immunocytochemical analysis. RESULTS: Before chemotherapy, only 10% of the NB samples showed positivity for P-gp. At diagnosis, no difference in P-gp expression was found between primary tumor cells and NB cells from metastases to bone marrow. P-gp positivity was only observed in patients with nonlocalized disease. P-gp positivity was never found in tumor cells that were histologically well differentiated. No clear correlation of P-gp positivity with poor prognostic parameters, such as chromosome 1p deletion or MYCN amplification, were found. Multidrug chemotherapy did not induce enhanced expression of P-gp in the neuroblasts. However, at clinical recurrence, P-gp expression was found in the metastatic NB cells of five of seven bone marrow samples examined. CONCLUSIONS: The prognostic relevance of P-gp expression in NB was not clear from the results of this study. To resolve the uncertainties, a standardization of the methodology and more prospective studies are needed to determine whether routine analysis of P-gp is worth adding to the other prognostic parameters that are evaluated in NB patients. The finding that metastatic cells are capable of expressing MDR1, in contrast to the NB cells of the primary tumor, would certainly be an interesting topic for further study as work directed at understanding the progression to metastasis continues.

P-glycoprotein is an independent prognostic factor predicting relapse in childhood acute lymphoblastic leukaemia: results of a 6-year prospective study.

P-glycoprotein (P-gp), a cellular drug-efflux pump, is thought to be one of the major causes of multidrug resistance (MDR) in malignancies. Since therapeutic strategies are being developed to circumvent drug resistance by inhibiting P-gp function, large prospective studies evaluating the clinical relevance of P-gp in childhood acute lymphoblastic leukaemia (ALL) are warranted. P-gp expression was evaluated over a period of 6 years in 102 consecutive patients with de novo childhood ALL and in 35 children with relapse of ALL. Bone marrow and blood smears were studied immunocytochemically with two monoclonal antibodies at initial diagnosis and at relapse. P-gp expression was found in 14 (14%) patients at initial diagnosis. After induction treatment, complete remission was achieved in 100/102 patients (98%), of whom 19 relapsed. Cumulative event-free survival was significantly higher in the P-gp-negative group compared with the P-gp-positive population (Logrank P = 0.02). Multivariate analysis showed the results to be independent of age, WBC count and karyotype, and concomitantly underlined the importance of MDR1 phenotype detection in childhood ALL. P-gp expression was more frequently found at relapse (34%) than at primary diagnosis (P = 0.01). In the relapsed patient group, P-gp-positive patients had a 2-fold greater risk for adverse clinical outcome than the P-gp-negative relapsed patients. P-gp expression was not induced by exposure to previous chemotherapy since the majority of P-gp-negative patients remained negative at relapse. P-glycoprotein expression in newly diagnosed childhood ALL is an independent adverse prognostic parameter with a predictive value for relapse.

Repetitive analyses of P-glycoprotein in chronic myeloid leukaemia.

P-glycoprotein, a pump located in the plasma cell membrane, extrudes several clinically important drugs from the cell, and hence causes multidrug resistance. Reversing clinical drug resistance is possible by using agents that inhibit the activity of P-glycoprotein. We describe the results of sequential flow cytometric determinations of P-glycoprotein expression and activity in two patients suffering from acute lymphoblastic transformation of chronic myeloid leukaemia. Neither P-glycoprotein expression, nor its activity could be detected in the initial sample of the first patient. In the second patient, no P-glycoprotein expression was found at diagnosis. However, after chemotherapy containing P-glycoprotein substrates, a significant expression was found in both patients and the functional flow cytometric test was positive. In order to achieve an accurate selection of patients that might benefit from the clinical use of P-gp inhibitors, repeated analyses are indicated in each patient suffering from acute leukaemia, during the course of the illness.