[Survival of children with recurrent medulloblastoma undergoing sequential therapy: an analysis of 101 cases].

OBJECTIVE: To study the clinical features of children with recurrent medulloblastoma (MB) and treatment regimens. METHODS: A retrospective analysis was performed on 101 children with recurrent MB who were admitted to the hospital from August 1, 2011 to July 31, 2017. The children were followed up to July 31, 2020. The Kaplan-Meier method was used for survival analysis. The Cox regression model was used for multivariate regression analysis. RESULTS: Of the 101 children, 95 underwent remission induction therapy, among whom 51 had response, resulting in a response rate of 54%. The median overall survival (OS) time after recurrence was 13 months, and the 1-, 3-, and 5-year OS rates were 50.5%±5.0%, 19.8%±4.0%, and 10%±3.3% respectively. There was no significant difference in the 5-year OS rate between the children with different ages (< 3 years or 3-18 years), sexes, pathological types, or Change stages, between the children with or without radiotherapy before recurrence or re-irradiation after recurrence, and between the children with different times to recurrence (< 12 months or ≥ 12 months after surgery) (P > 0.05). There were significant differences in the 5-year OS rate between the children with or without reoperation after recurrence and between the children with different recurrence sites (P < 0.05). The children with reoperation after recurrence had a significantly longer survival time than those without reoperation (P=0.007), and the risk of death in children undergoing reoperation after recurrence was 0.389 times (95% confidence interval:0.196-0.774) that in children who did not undergo such reoperation. CONCLUSIONS: As for the recurrence of MB, although remission induction therapy again can achieve remission, such children still have a short survival time. Only reoperation can significantly prolong survival time, and therefore, early reoperation can be considered to improve the outcome of children with recurrent MB.

Methylation analysis of the SLC19A1 promoter region in Chinese children with acute lymphoblastic leukaemia.

WHAT IS KNOWN AND OBJECTIVE: Reduced folate carrier 1 (RFC1), which is encoded by the human solute carrier family 19 member 1 (SLC19A1) gene, plays an essential role in the cellular uptake of methotrexate (MTX). RFC1 expression is regulated by genetic variations and epigenetic modifications. The aim of the present study was to investigate the methylation status of the SLC19A1 promoter in peripheral blood and its association with MTX levels and toxicities in children with acute lymphoblastic leukaemia (ALL). METHODS: Serum MTX concentrations were measured using a fluorescence polarization immunoassay. Methylation quantification for SLC19A1 promoter region #17 was performed by Sequenom MassARRAY in 52 paediatric ALL patients. RESULTS AND DISCUSSION: Overall, the investigated region of the SLC19A1 promoter was in a hypermethylated state. No significant associations were detected between the methylation levels of six CpG units in the SLC19A1 promoter region #17 and clinical parameters of patients with ALL, including sex, age, immunotype and risk stratification. The methylation level of CpG_10 showed a significant positive correlation with MTX 24 hours after the initiation of infusion. No significant differences in the methylation levels of six CpG units were observed between patients with and without MTX toxicities. Due to the small sample size of this study, there was a high chance of false-positive results. A large-scale study would be required to confirm these preliminary results. WHAT IS NEW AND CONCLUSION: Our preliminary results suggested the hypermethylated status of the SLC19A1 promoter in children with ALL. The methylation levels of the SLC19A1 promoter might affect MTX exposure. These findings have implications for the mechanisms underlying the variability of MTX responses in childhood ALL.

Frequency distribution of five SNPs in human GGH gene and their effects on clinical outcomes of Chinese pediatric patients with acute lymphoblastic leukemia.

Methotrexate (MTX) is widely used in the treatment of childhood acute lymphoblastic leukemia (ALL). Gamma-glutamyl hydrolase (GGH) plays an important role in the disposition of MTX. The aim of this study was to investigate the frequency distribution of five SNPs in the human GGH gene and their effects on serum MTX concentrations and clinical outcomes in Chinese children with ALL. Genotyping of 149 pediatric patients for GGH rs11545078 C>T, rs11545077 G>A, rs1800909 T>C, rs11545076 T>G, and rs3758149 C>T was performed using the Sequenom MassARRAY system. Serum MTX concentrations were determined using a fluorescence polarization immunoassay. The five SNPs studied were in strong linkage. The minor allele frequencies for rs11545078, rs11545077, rs1800909, rs11545076, and rs3758149 were 5.3, 15.0, 14.3, 15.0, and 15.0%, respectively. Four haplotypes (CGTTC, CACGT, TACGT, and TATGT) were observed at frequencies of 84.9, 9.8, 4.5, and 0.8%, respectively. The median C/D ratios of serum MTX at 24 h and 42 h in children with variant haplotypes (12.30 and 0.08 µmol/L per g/m², respectively) were higher than those in wild haplotype carriers (11.85 and 0.07 µmol/L per g/m², respectively). The event-free survival of patients with variant haplotypes (89.2%) was significantly better than that of patients with wild haplotypes (71.9%, P < 0.05). The relapse rate in children with variant haplotypes (8.1%) was lower than that in children with wild haplotypes (15.6%). These findings have implications for the efficacious use of MTX in childhood ALL patients.

[Clinical effect of surgery combined with chemotherapy and radiotherapy in children with central primitive neuroectodermal tumor and prognostic analysis].

OBJCTIVE: To study the clinical effect of surgery combined with chemotherapy and radiotherapy in children with central primitive neuroectodermal tumor (cPNET), as well as the risks factors for poor prognosis. METHODS: A retrospective analysis was performed for the clinical data of 42 children who were diagnosed with cPNET from June 2012 to September 2018. RESULTS: The 42 children had a median overall survival (OS) time of 2.0 years and a median event-free survival (EFS) time of 1.3 years; the 1-, 3-, and 5-year OS rates were 76.2%±6.6%, 41.4%±8.7%, 37.3%±8.8% respectively, and the 1-, 3-, and 5-year EFS rates were 64.3%±7.4%, 32.7%±8.0%, 28.0%±8.1% respectively. The univariate analysis showed that there were significant differences in the OS and EFS rates among the children with different patterns of surgical resection, chemotherapy cycles, and risk grades (P<0.05), and there was also a significant difference in the OS rate between the children receiving radiotherapy and those not receiving radiotherapy (P<0.05). The multivariate Cox regression analysis showed that chemotherapy cycles and risk grade were independent influencing factors for EFS and OS rates (P<0.05). The EFS and OS rates increased with the increase in chemotherapy cycles and the reduction in risk grade. CONCLUSIONS: Multimodality therapy with surgery, chemotherapy, and radiotherapy is an effective method for the treatment of cPNET in children. Early diagnosis and treatment and adherence to chemotherapy for as long as possible may improve EFS and OS rates.

The role of transcription factor Sp1 in the regulation of gamma-glutamyl hydrolase gene expression by the rs3758149 polymorphism in CEM/C1 cells.

Gamma-Glutamyl hydrolase (GGH) plays an important role in the disposition of anti-folate analogs. Several studies noted the pharmacological relevance of rs3758149 C/T polymorphism located in the human GGH promoter. The present study aimed to investigate the role of rs3758149 C/T polymorphism and transcription factors in the regulation of GGH expression in human acute lymphoblastic leukemia (ALL) CEM/C1 cells. Compared with the rs3758149 T allele, the C allele showed significantly higher transcriptional activity in luciferase reporter assays, as well as a stronger binding affinity for the nuclear protein extracts in an electrophoretic mobility shift assay. Sp1 was identified as the target transcription factor that exhibited allele-specific binding to the location of rs3758149 C/T polymorphism in the chromatin immunoprecipitation assay. Overexpression of Sp1 led to enhanced GGH promoter activity and GGH mRNA expression in allele-specific manners. These findings suggested that Sp1 acted as a positive regulator of human GGH transcription through the rs3758149 polymorphism in CEM/C1 cells. This study contributed to the present understanding of the mechanisms underlying variable responses of ALL to anti-folates.

[Risk factors for recurrence within 2 years in children with medulloblastoma].

OBJECTIVE: To investigate the risk factors for recurrence of medulloblastoma (MB) within 2 years and their influence on progression-free survival (PFS). METHODS: A retrospective analysis was performed for the clinical data of 123 children with MB who were admitted from January to December, 2017. According to the presence or absence of recurrence, they were divided into recurrence group with 30 children and non-recurrence group with 93 children. The risk factors for recurrence within 2 years were analyzed, and PFS was compared between the children with different risk factors. RESULTS: Large-cell/anaplastic type and M stage were risk factors for MB recurrence within 2 years. The risk of recurrence in the children with M+ MB was 3.525 times that in those with M0 MB, and the risk of recurrence in the children with large-cell/anaplastic MB was 3.358 times that in those with classic MB (P<0.05). The survival analysis showed that the median PFS time was 20 months in the children with M+ MB, and the 20-month PFS rate was 50% ±â€…11% in the children with M+ MB and 81% ±â€…5% in those with M0 MB (P<0.05). The 20-month PFS rate was 80% ±â€…5% in the children with classic MB, 65% ±â€…10% in those with desmoplastic/nodular MB, 86% ±â€…13% in those with MB with extensible nodularity, and 36% ±â€…20% in those with large-cell/anaplastic MB (P<0.05). CONCLUSIONS: Recurrence is an important influencing factor for the prognosis of MB, and M+ stage and large-cell/anaplastic MB are risk factors for recurrence. Children with such risk factors tend to have a low PFS rate.

[Effect of bevacizumab in treatment of children with optic pathway glioma].

OBJECTIVE: To investigate the effect of bevacizumab in the treatment of children with optic pathway glioma (OPG). METHODS: A retrospective analysis was performed for the clinical data of 30 children with OPG who underwent chemotherapy. According to whether bevacizumab was used, they were divided into conventional chemotherapy (carboplatin, vincristine and etoposide) group with 12 children and combined chemotherapy (bevacizumab, carboplatin, vincristine and etoposide) group with 18 children. The children were followed up to 6 months after chemotherapy, and the two groups were compared in terms of visual acuity and tumor size before and after chemotherapy and adverse reactions during chemotherapy. RESULTS: The combined chemotherapy group had a significantly higher proportion of children achieving tumor regression than the conventional chemotherapy group (P<0.05), while there were no significant differences between the two groups in the proportion of children with improved visual acuity or adverse reactions (P>0.05). No chemotherapy-related death was observed in either group. CONCLUSIONS: Bevacizumab combined with conventional chemotherapy can effectively reduce tumor size. Compared with conventional chemotherapy, such combination does not increase adverse reactions and can thus become a new direction for the treatment of OPG in children.

Association between MTHFR microRNA binding site polymorphisms and methotrexate concentrations in Chinese pediatric patients with acute lymphoblastic leukemia.

BACKGROUND: The pharmacokinetics and therapeutic response to methotrexate (MTX) display large variability in the treatment of acute lymphoblastic leukemia (ALL). The aim of the present study was to investigate the association of two microRNA (miRNA) binding site polymorphisms (rs3737966 G > A and rs35134728 DEL/TTC) in the 3'-untranslated region of MTHFR with serum MTX concentrations, in a Chinese pediatric population with ALL. METHODS: Genotyping for MTHFR rs3737966 and rs35134728 in 144 children with ALL was performed using the Sequenom MassArray system (Sequenom, San Diego, CA, USA). Serum MTX concentrations were measured by a fluorescence polarization immunoassay 24 h (C24h ) and 42 h (C42h ) after administration. The effects of the polymorphisms on concentration-to-dose (C/D) ratios of MTX were assessed. RESULTS: Complete linkage disequilibrium between rs3737966 and rs35134728 polymorphisms (r2  = 1) was found in the study population. The minor allele frequency observed in the present study (17.4%) was significantly lower than those in European and African samples reported in the 1000 Genomes Project (42.9% and 63.9%, respectively; p < 0.01). The C/D ratios of MTX at 24 and 42 h for the TTC/TTC-A/A haplotype carriers (11.74 and 0.07 µmol/l per g/m2 , respectively) were significantly lower than those in DEL/DEL-G/G or DEL/TTC-G/G haplotype carriers (12.49 and 0.09 µmol/l per g/m2 , respectively; p < 0.05). Computational predictions suggested that the two polymorphisms overlapped with putative binding sites of several miRNAs. CONCLUSIONS: The rs3737966 and rs35134728 polymorphisms in MTHFR were associated with serum MTX concentrations. The findings of the present study indicate that miRNAs might be involved in the post-transcriptional regulation of MTHFR.

Genetic Landscape and Its Prognostic Impact in Children With Langerhans Cell Histiocytosis.

CONTEXT.­: Langerhans cell histiocytosis (LCH) is a rare myeloid neoplasm that predominantly affects young children. OBJECTIVE.­: To investigate genetic alterations and their correlation with clinical characteristics and prognosis in pediatric LCH. DESIGN.­: We performed targeted sequencing to detect mutations in LCH lesions from pediatric patients. RESULTS.­: A total of 30 genomic alterations in 5 genes of the MAPK pathway were identified in 187 of 223 patients (83.9%). BRAF V600E (B-Raf proto-oncogene, serine/threonine kinase) was the most common mutation (51.6%), followed by MAP2K1 (mitogen-activated protein kinase kinase 1) alterations (17.0%) and other BRAF mutations (13.0%). ARAF (A-Raf proto-oncogene, serine/threonine kinase) and KRAS (KRAS proto-oncogene, GTPase) mutations were relatively rare (2.2% and 0.9%, respectively). Additionally, FNBP1 (formin-binding protein 1)::BRAF fusion and MAP3K10 (mitogen-activated protein kinase kinase 10) mutations A17T and R823C were identified in 1 case each, with possible constitutive activation of ERK1/2 phosphorylation. BRAF V600E was more frequent in patients with risk organ involvement, while MAP2K1 mutation was more prevalent in patients with single-system LCH (P = .001). BRAF V600E was associated with craniofacial bone, skin, liver, spleen, and ear involvement (all P < .05). Patients with other BRAF mutations had a higher proportion of spinal column involvement (P = .006). Univariate analysis showed a significant difference in progression-free survival among the 4 molecular subgroups for patients treated with first-line therapy (P = .02). According to multivariate analysis, risk organ involvement was the strongest independent adverse prognostic factor (hazard ratio, 8.854; P < .001); BRAF or MAP2K1 mutation was not an independent prognostic factor. CONCLUSIONS.­: Most pediatric patients with LCH carry somatic mutations involving the MAPK pathway, correlating with clinical characteristics and outcomes for first-line chemotherapy.

MiR-595 Suppresses the Cellular Uptake and Cytotoxic Effects of Methotrexate by Targeting SLC19A1 in CEM/C1 Cells.

The human solute carrier family 19 member 1 (SLC19A1) is the gene coding for reduced folate carrier 1 (RFC1). In our previous work, we showed that the miR-595-related polymorphism, rs1051296 G>T, which was located in the 3'-untranslated region (3'-UTR) of SLC19A1, was associated with high methotrexate (MTX) plasma concentrations in patients with paediatric acute lymphoblastic leukaemia (ALL). This study aimed to investigate the role of miR-595 in the regulation of SLC19A1 expression and its effects on the cellular uptake and cytotoxicity of MTX in ALL CEM/C1 cells. Luciferase reporter assay was performed to validate SLC19A1 as a miR-595 target. RFC1 protein expression was determined via Western blotting. Intracellular MTX concentrations were measured by enzyme-linked immunosorbent assay (ELISA). Cell viability and apoptosis were assessed using Cell Counting Kit-8 (CCK-8) assay and flow cytometer, respectively. Compared to the negative control, miR-595 mimics induced a significant decrease in the relative luciferase activity by binding to the 3'-UTR of SLC19A1 harbouring the rs1051296 T allele (p < 0.01). Treatment of CEM/C1 cells with miR-595 mimics substantially reduced RFC1 protein expression, intracellular MTX levels, MTX-induced cytotoxicity and apoptosis rates compared to those of negative control. However, opposite results were observed in cells transfected with a miR-595 inhibitor. These findings suggested that miR-595 acts as a phenotypic regulator of MTX sensitivity in CEM/C1 cells by targeting SLC19A1. This study helped us to understand the mechanisms underlying the variable MTX responses observed in patients with ALL.