Brachytherapy for Pediatric Patients at Gustave Roussy Cancer Campus: A Model of International Cooperation for Highly Specialized Treatments.

PURPOSE: Childhood cancer is rare, and treatment is frequently associated with long-term morbidity. Disparities in survival and long-term side effects encourage the establishment of networks to increase access to complex organ-conservative strategies, such as brachytherapy. We report our experience of an international cooperation model in childhood cancers. METHODS AND MATERIALS: We examined the outcome of all children referred to our center from national or international networks to be treated according to a multimodal organ-conservative approach, including brachytherapy. RESULTS: We identified 305 patients whose median age at diagnosis was 2.2 years (range, 1.4 months to 17.2 years). Among these patients, 99 (32.4%) were treated between 2015 and 2020; 172 (56.4%) were referred from national centers; and 133 (43.6%) were international patients from 31 countries (mainly Europe). Also, 263 patients were referred for primary treatment and 42 patients were referred for salvage treatment. Genitourinary tumors were the most frequent sites, with 56.4% bladder/prostate rhabdomyosarcoma and 28.5% gynecologic tumors. In addition to brachytherapy, local treatment consisted of partial tumor resection in 207 patients (67.9%), and 39 patients (13%) had additional external radiation therapy. Median follow-up was 58 months (range, 1 month to 48 years), 93 months for national patients, and 37 months for international patients (P < .0001). Five-year local control, disease-free survival, and overall survival rates were 90.8% (95% confidence interval [CI], 87.3%-94.4%), 84.4% (95% CI, 80.1%-89.0%), and 93.3% (95% CI, 90.1%-96.5%), respectively. Patients referred for salvage treatment had poorer disease-free survival (P < .01). Implementation of image guided pulse-dose-rate brachytherapy was associated with better local control among patients with rhabdomyosarcoma referred for primary treatment (hazard ratio, 9.72; 95% CI, 1.24-71.0). At last follow-up, 16.7% patients had long-term severe treatment-related complications, and 2 patients (0.7%) had developed second malignancy. CONCLUSIONS: This retrospective series shows the feasibility of a multinational referral network for brachytherapy allowing high patient numbers in rare pediatric cancers. High local control probability and acceptable late severe complication probability could be achieved despite very challenging situations. This cooperation model could serve as a basis for generating international reference networks for high-tech radiation such as brachytherapy to increase treatment care opportunities and cure probability.

Rare KMT2A-ELL and Novel ZNF56-KMT2A Fusion Genes in Pediatric T-cell Acute Lymphoblastic Leukemia.

BACKGROUND/AIM: Previous reports have associated the KMT2A-ELL fusion gene, generated by t(11;19)(q23;p13.1), with acute myeloid leukemia (AML). We herein report a KMT2A-ELL and a novel ZNF56-KMT2A fusion genes in a pediatric T-lineage acute lymphoblastic leukemia (T-ALL). MATERIALS AND METHODS: Genetic investigations were performed on bone marrow of a 13-year-old boy diagnosed with T-ALL. RESULTS: A KMT2A-ELL and a novel ZNF56-KMT2A fusion genes were generated on der(11)t(11;19)(q23;p13.1) and der(19)t(11;19)(q23;p13.1), respectively. Exon 20 of KMT2A fused to exon 2 of ELL in KMT2A-ELL chimeric transcript whereas exon 1 of ZNF56 fused to exon 21 of KMT2A in ZNF56-KMT2A transcript. A literature search revealed four more T-ALL patients carrying a KMT2A-ELL fusion. All of them were males aged 11, 11, 17, and 20 years. CONCLUSION: KMT2A-ELL fusion is a rare recurrent genetic event in T-ALL with uncertain prognostic implications. The frequency and impact of ZNF56-KMT2A in T-ALL are unknown.

Transcriptome analysis reveals dysregulation of genes involved in oxidative phosphorylation in a murine model of retinopathy of prematurity.

BACKGROUND: Retinal gene expression pattern is severely altered after exposition to hyperoxia in mice with oxygen-induced retinopathy (OIR), a common model of retinopathy of prematurity. Gene ontology and signaling pathway analyses may add new insights into a better understanding of the pathogenesis of this disease. METHODS: Seven-day-old C57BL/6J mice (n = 60) were exposed to 75% oxygen for 5 days and then recovered in room air. The controls (n = 60) were kept in the normoxic conditions. Retinas were harvested immediately following hyperoxia, during the phase of maximal neovascularization, and at the time of neovascularization regression. The retinal RNA samples were evaluated for gene expression using mouse gene expression microarrays. DAVID annotation tools were used for gene ontology and pathway analyses. RESULTS: The most significantly enriched signaling pathways during the neovascularization phase of OIR were: focal adhesion; ECM-receptor interaction; PI3K-Akt; oxidative phosphorylation; and Alzheimer's, Parkinson's and Huntington's disease signaling pathways. Genes involved in apoptosis, cell proliferation, cell differentiation, and immune responses were associated with neovascularization regression. CONCLUSIONS: Performed analyses revealed the possible involvement of various signaling pathways in OIR pathomechanism, mostly specific to the OIR phase. Dysregulation of genes involved in oxidative phosphorylation may have an impact on neovascularization development.

Pulmonary vascular disease is evident in gene regulation of experimental bronchopulmonary dysplasia.

Objective: To examine the gene expression regarding pulmonary vascular disease in experimental bronchopulmonary dysplasia in young mice. Premature delivery puts babies at risk of severe complications. Bronchopulmonary dysplasia (BPD) is a common complication of premature birth leading to lifelong affection of pulmonary function. BPD is recognized as a disease of arrested alveolar development. The disease process is not fully described and no complete cure or prevention is known. The focus of interest in the search for treatment and prevention of BPD has traditionally been at airspace level; however, the pulmonary vasculature is increasingly acknowledged in the pathology of BPD. The aim of the investigation was to study the gene expression in lungs with BPD with regards to pulmonary vascular disease (PVD).Methods: We employed a murine model of hyperoxia-induced BPD and gene expression microarray technique to determine the mRNA expression in lung tissue from young mice. We combined gene expression pathway analysis and analyzed the biological function of multiple single gene transcripts from lung homogenate to study the PVD relevant gene expression.Results: There were n = 117 significantly differentially regulated genes related to PVD through down-regulation of contractile elements, up- and down-regulation of factors involved in vascular tone and tissue-specific genes. Several genes also allowed for pinpointing gene expression differences to the pulmonary vasculature. The gene Nppa coding for a natriuretic peptide, a potent vasodilator, was significantly down-regulated and there was a significant up-regulation of Pde1a (phosphodiesterase 1A), Ptger3 (prostaglandin e receptor 3), and Ptgs1 (prostaglandin-endoperoxide synthase one).Conclusion: The pulmonary vasculature is affected by the arrest of secondary alveolarization as seen by differentially regulated genes involved in vascular tone and pulmonary vasculature suggesting BPD is not purely an airspace disease. Clues to prevention and treatment may lie in the pulmonary vascular system.

Short- and long-term impact of hyperoxia on the blood and retinal cells' transcriptome in a mouse model of oxygen-induced retinopathy.

BACKGROUND: We aimed to identify global blood and retinal gene expression patterns in murine oxygen-induced retinopathy (OIR), a common model of retinopathy of prematurity, which may allow better understanding of the pathogenesis of this severe ocular prematurity complication and identification of potential blood biomarkers. METHODS: A total of 120 C57BL/6J mice were randomly divided into an OIR group, in which 7-day-old pups were maintained in 75% oxygen for 5 days, or a control group. RNA was extracted from the whole-blood mononuclear cells and retinal cells on days 12, 17, and 28. Gene expression in the RNA samples was evaluated with mouse gene expression microarrays. RESULTS: There were 38, 1370 and 111 genes, the expression of which differed between the OIR and control retinas on days 12, 17, and 28, respectively. Gene expression in the blood mononuclear cells was significantly altered only on day 17. Deptor and Nol4 genes showed reduced expression both in the blood and retinal cells on day 17. CONCLUSION: There are sustained marked changes in the global pattern of gene expression in the OIR mice retinas. An altered expression of Deptor and Nol4 genes in the blood mononuclear cells requires further investigation as they may indicate retinal neovascularization.

Immune System Regulation Affected by a Murine Experimental Model of Bronchopulmonary Dysplasia: Genomic and Epigenetic Findings.

BACKGROUND: Bronchopulmonary dysplasia (BPD) is a common cause of abrupted lung development after preterm birth. BPD may lead to increased rehospitalization, more severe and frequent respiratory infections, and life-long reduced lung function. The gene regulation in lungs with BPD is complex, with various genetic and epigenetic factors involved. OBJECTIVES: The aim of this study was to examine the regulatory relation between gene expression and the epigenome (DNA methylation) relevant for the immune system after hyperoxia followed by a recovery period in air using a mouse model of BPD. METHODS: Newborn mice pups were subjected to an immediate hyperoxic condition from birth and kept at 85% O2 levels for 14 days followed by a 14-day period in room air. Next, mice lung tissue was used for RNA and DNA extraction with subsequent microarray-based assessment of lung transcriptome and supplementary methylome analysis. RESULTS: The immune system-related transcriptomeregulation was affected in mouse lungs after hyperoxia. A high proportion of genes relevant in the immune system exhibited significant expression alterations, e.g., B cell-specific genes central to the cytokine-cytokine receptor interaction, the PI3K-AKT, and the B cell receptor signaling pathways. The findings were accompanied by significant DNA hypermethylation observed in the PI3K-AKT pathway and immune system-relevant genes. CONCLUSIONS: Oxygen damage could be partly responsible for the increased susceptibility and abnormal response to respiratory viruses and infections seen in premature babies with BPD through dysregulated genes.

Neonatal Ogg1/Mutyh knockout mice have altered inflammatory gene response compared to wildtype mice in the brain and lung after hypoxia-reoxygenation.

Background 8-Oxoguanine DNA-glycosylase 1 (OGG1) and mutY DNA glycosylase (MUTYH) are crucial in the repair of the oxidative DNA lesion 7,8-dihydro-8-oxoguanine caused by hypoxia-reoxygenation injury. Our objective was to compare the gene expression changes after hypoxia-reoxygenation in neonatal Ogg1-Mutyh double knockout mice (OM) and wildtype mice (WT), and study the gene response in OM after hyperoxic reoxygenation compared to normoxic. Methods Postnatal day 7 mice were subjected to 2 h of hypoxia (8% O2) followed by reoxygenation in either 60% O2 or air, and sacrificed right after completed reoxygenation (T0h) or after 72 h (T72h). The gene expression of 44 a priori selected genes was examined in the hippocampus/striatum and lung. Results We found that OM had an altered gene response compared to WT in 21 genes in the brain and 24 genes in the lung. OM had a lower expression than WT of inflammatory genes in the brain at T0h, and higher expression at T72h in both the brain and lung. In the lung of OM, five genes were differentially expressed after hyperoxic reoxygenation compared to normoxic. Conclusion For the first time, we report that Ogg1 and Mutyh in combination protect against late inflammatory gene activation in the hippocampus/striatum and lung after neonatal hypoxia-reoxygenation.

Temporal Patterns of Gene Expression Profiles in the Neonatal Mouse Lung after Hypoxia-Reoxygenation.

BACKGROUND: One out of four children with neonatal asphyxia has lung involvement. Still, there has been little research on injury mechanisms of hypoxia-reoxygenation in the neonatal lung. OBJECTIVES: To make a temporal profile of the gene expression changes of 44 a priori selected genes after hypoxia-reoxygenation in the newborn mouse lung, and to compare the changes after hyperoxic and normoxic reoxygenation. METHODS: Postnatal day 7 mice were randomized to 2-hour hypoxia (8% O2) and 30-min reoxygenation in either 60% O2 or air. After 0-72 h of observation, gene expression changes and protein concentrations in whole lung homogenates were examined. RESULTS: Immediately after completed reoxygenation, 7 genes of mediators of inflammation were downregulated, and there was an antiapoptotic gene expression pattern. Three DNA glycosylases were downregulated, while genes involved in cell cycle renewal indicated both increased and decreased cell cycle arrest. Sod1 (T2.5h median H60: 1.01, H21: 0.88, p = 0.005; T5h median H60: 1.04, H21: 0.85, p = 0.038) and Il1b (T0h median H60: 0.86, H21: 1.08, p = 0.021) were significantly differentially expressed when comparing hyperoxic and normoxic reoxygenation. CONCLUSION: In this newborn mouse lung hypoxia-reoxygenation model, we found downregulation of genes of mediators of inflammation, an antiapoptotic gene expression pattern, and downregulation of DNA glycosylases. Sod1 and Il1b were significantly differentially expressed when comparing reoxygenation using 60% O2 with air.

Increased expression of inflammatory genes in the neonatal mouse brain after hyperoxic reoxygenation.

BACKGROUND: Hyperoxic reoxygenation following hypoxia increases the expression of inflammatory genes in the neonatal mouse brain. We have therefore compared the temporal profile of 44 a priori selected genes after hypoxia and hyperoxic or normoxic reoxygenation. METHODS: Postnatal day 7 mice were subjected to 2 h of hypoxia (8% O2) and 30 min reoxygenation with 60% or 21% O2. After 0 to 72 h observation, mRNA and protein were examined in the hippocampus and striatum. RESULTS: There were significantly higher gene expression changes in six genes after hyperoxic compared to normoxic reoxygenation. Three genes had a generally higher expression throughout the observation period: the inflammatory genes Hmox1 (mean difference: 0.52, 95% confidence interval (CI): 0.15-1.01) and Tgfb1 (mean difference: 0.099, CI: 0.003-0.194), and the transcription factor Nfkb1 (mean difference: 0.049, CI: 0.011-0.087). The inflammatory genes Cxcl10 and Il1b, and the DNA repair gene Neil3, had a higher gene expression change after hyperoxic reoxygenation at one time point only. Nineteen genes involved in inflammation, transcription regulation, apoptosis, angiogenesis, and glucose transport had significantly different gene expression changes with time in all intervention animals. CONCLUSION: We confirm that hyperoxic reoxygenation induces a stronger inflammatory gene response than reoxygenation with air.

Coronavirus causes lower respiratory tract infections less frequently than RSV in hospitalized Norwegian children.

BACKGROUND: We have described occurrence and clinical manifestations of human coronaviruses (HCoV) in hospitalized Norwegian children with respiratory tract infection (RTI) and compared them with a group of respiratory syncytial virus (RSV)-infected children. METHODS AND POPULATION: We used in-house TaqMan multiplex real-time polymerase chain reaction to test nasopharyngeal samples from 536 RTI episodes in 452 children who were admitted during the 2006-2007 winter. Twenty-one viruses, including HCoV-OC43, HCoV-NL63, HCoV-229E, HCoV-HKU1, and RSV were tested. The amount of viral nucleic acid was recorded semiquantitatively based on the cycle threshold value. RESULTS: A total of 665 positive polymerase chain reaction tests were recorded in 536 nasopharyngeal specimens. Coronavirus was found in 68 (12.7%): HCoV-OC43, n = 44 (8.2%), and HCoV-NL63, n = 24 (4.5%). Only RSV and rhinovirus were detected more frequently. Neither HCoV-229E nor HCoV-HKU1 was detected. Among children with HCoV-OC43, 73.0% tested positive for at least one other virus, compared with 41.2% with HCoV-NL63 and 40.3% with RSV (P = 0.03 and P < 0.01, respectively). Children with HCoV-OC43 and HCoV-NL63 were older than children with RSV (median age, 19 vs. 10 months, P = 0.01). Lower respiratory tract infection (LRTI) was half as common in children with HCoV-OC43 (48.6%) and HCoV-NL63 (47.1%) as in children with RSV (82.3%) (both P < 0.01). After adjusting for age, chronic disease, LRTI, and co-detection of other viruses in a multiple logistic regression analysis, HCoV was associated with a shorter fever period and shorter hospitalization time than RSV. CONCLUSIONS: HCoV-OC43 and HCoV-NL63 are common among hospitalized Norwegian children with RTI. Children with HCoV-OC43 and HCoV-NL63 have LRTI less frequently and may need a shorter hospital stay than children with RSV.

Human bocavirus in children: mono-detection, high viral load and viraemia are associated with respiratory tract infection.

BACKGROUND AND OBJECTIVES: Human bocavirus 1 (HBoV1) has recently been detected in children with respiratory tract infections (RTI). In order to study whether HBoV1 can cause RTI, we investigated its presence in children with upper RTI (URTI), lower RTI (LRTI) and a control group of children without RTI. STUDY DESIGN: Nasopharyngeal aspirates (NPA) and blood samples were collected from children admitted to hospital with RTI from 6 June 2007 to 28 February 2009 (n=1154), and from children admitted for elective surgery who had no RTI (n=162). Using polymerase chain reaction (PCR), the NPAs were examined for 17 infectious agents including HBoV1. Blood samples were tested with HBoV1-PCR only. RESULTS: HBoV1 was detected in NPAs from 10% of patients and 17% of controls. Adjusted for age, gender and the presence of other viruses, HBoV1 was not associated with RTI. In the HBoV1-positive NPAs, at least one other virus was detected in 75% and the virus appeared alone in 25%. Adjusted for age and gender, the detection of HBoV1 as the sole virus was associated with RTI, but not with LRTI. Viraemia was found only in children with RTI. The study showed that it was associated with RTI and LRTI. A high HBoV1-load was associated with LRTI, but not with RTI. No interactions between HBoV1 and other infectious agents were found. CONCLUSIONS: Our data support the hypothesis that HBoV1 causes RTI in children, because detection of HBoV1 alone, viraemia and high viral load are associated with RTI and/or LRTI in this age group. However, HBoV1 is common in healthy children.

Rotavirus gastroenteritis in Norway: analysis of prospective surveillance and hospital registry data.

We aimed to evaluate rotavirus morbidity and describe rotavirus epidemiology in hospitalized children in Norway to provide information before the introduction of new rotavirus vaccines. We retrospectively reviewed 14,973 gastroenteritis hospitalizations in children aged <5 y for the period 1995 to 2004, and prospectively surveyed for rotavirus in 311 children aged <5 y admitted with diarrhoea to 3 hospitals in 2006-2008. The proportion of rotavirus among all gastroenteritis hospitalizations was estimated at 14.5% from the retrospective data and at 62.9% in the prospective data. The annual incidence of rotavirus hospitalizations is estimated to be 3 per 1000 children <5 y of age, corresponding to approximately 900 (range 735-1092) hospitalizations each year. Children aged 6-23 months accounted for 61% of all confirmed rotavirus cases, and average duration of hospital stay for rotavirus cases was 1.3 days. We observed a predominance of rotavirus infections from March through May, similar to the seasonality of diarrhoea-associated hospitalizations with viral and unspecified aetiology. No rotavirus-associated deaths were reported. It is estimated that two thirds of all gastroenteritis hospitalizations in children <5 y of age may be attributable to rotavirus in Norway. Continued surveillance and further studies are needed to assess the full burden of rotavirus disease and its economic impact in Norway.

Detection and characterization of group A rotaviruses in children hospitalized with acute gastroenteritis in Norway, 2006-2008.

To assess the genetic diversity of rotavirus strains in Norway, the distribution of rotavirus genotypes was studied in children admitted to hospital with acute gastroenteritis. The detection of rotavirus in stool samples was compared using an enzyme-linked immunosorbent assay (ELISA), an immunochromatographic test and RT-PCR. Children <5 years of age admitted to hospital with diarrhea in three large hospitals were enrolled prospectively from March 2006 to February 2008. Rotavirus was detected in 58% of the children by the immunochromatographic test, in 63% by ELISA and 72% by RT-PCR. A total of 219 (70%) rotavirus isolates were characterized in order to determine the genotype. The predominant G types included G1 (53%), G9 (16%), and G3 (13%), and the frequency of G3 varied more than G9 between seasons (8-20%). The P[8] genotype was identified in 188 (86%) of samples, and the globally common genotype combinations G1P[8], G2P[4], G3P[8], G4P[8], and G9P[8] accounted together for >80% of infection. No unusual rotavirus strains were detected, and only four samples contained mixed infections. This study demonstrates that ELISA has similar specificity but lower sensitivity compared to RT-PCR. The immunochromatographic test had the lowest sensitivity and specificity compared to the other assays. Rotaviruses causing severe gastroenteritis leading to hospitalization of children <5 years of age in Norway include the common genotypes, however, a considerable geographical and seasonal variation was observed in the distribution of these genotypes. These data may be important for assessing the need for introducing rotavirus vaccines into immunization programs in Norway.

Human bocavirus commonly involved in multiple viral airway infections.

BACKGROUND: Human bocavirus (HBoV) was recently discovered in children with acute respiratory tract infections. We have included a PCR for HBoV in a study on airway infections in children. OBJECTIVES: To study the occurrence of HBoV in Norwegian children, and to evaluate the results of a semiquantitative PCR. STUDY DESIGN: During a 4-month period in the winter season 2006/2007 we collected nasopharyngeal aspirations from children who were admitted to the Department of Pediatrics. All samples were examined for 17 agents with real-time PCR. RESULTS: HBoV was detected in 45 of 376 samples (12%). The occurrence of HBoV was stable during the study period. Multiple viral infections were present in 78% of the samples (42% double, 20% triple and 16% quadruple infections). RS-virus, enterovirus and human metapneumovirus were the most frequently codetected agents. In samples with a high load for HBoV, significantly fewer multiple infections were found than in the other samples. Eighty-eight percent of the 25 patients with HBoV recorded as either the only or the dominating virus, and 50% of the other patients, had lower respiratory tract infection. The difference was statistically significant. CONCLUSIONS: HBoV was frequently detected in nasopharyngeal aspirates from children with airway infections in Norway. Multiple viral infections were common among the HBoV-infected patients. Semiquantitative PCR results may be useful for interpretation of clinical relevance.