Abstract：
BACKGROUND: Neuroblastoma (NB) is the most common solid tumour in childhood, and rises in the sympathetic nervous system. Here, we addressed the in silico analysis of the association between the expression of H2AFX gene involved in DNA damage response, and the survival of a cohort of 786 NB patients.
METHODS: In silico gene expression was retrieved from the publicly available dataset summarised by Cangelosi et al., including 13,696 gene expression profiles of 786 NB tumours at onset of disease. The prognostic value of H2AFX (H2A histone family member X) gene expression for event-free survival (EFS) and overall survival (OS) was evaluated by Kaplan-Meier and Cox regression analysis. The main results were validated on another openly accessible in silico database (NRC-283) containing 13,489 gene expressions in 283 NB patients. The expression of H2AFX protein was then tested by immunofluorescence on 48 primary NB samples of different tumour stages. H2AFX activity as an oncogene has been further validated in vitro by silencing the molecule in two NB cell lines, characterised by MYCN amplified or not, and performing cell growth and migration assays.
RESULTS: A strong inverse association between H2AFX expression and patients\' survival was observed and confirmed by immunofluorescence results on primary NB tissue sections. Cox regression analysis also disclosed H2AFX as an independent predictor of EFS and OS. The gene-silencing experiments strongly suggested an oncogenic role for H2AFX on NB cells, regardless of MYCN amplification.
CONCLUSIONS: H2AFX is a prognostic marker for unfavourable NB and could be considered a target for therapeutic interventions.
METHODS: In silico gene expression was retrieved from the publicly available dataset summarised by Cangelosi et al., including 13,696 gene expression profiles of 786 NB tumours at onset of disease. The prognostic value of H2AFX (H2A histone family member X) gene expression for event-free survival (EFS) and overall survival (OS) was evaluated by Kaplan-Meier and Cox regression analysis. The main results were validated on another openly accessible in silico database (NRC-283) containing 13,489 gene expressions in 283 NB patients. The expression of H2AFX protein was then tested by immunofluorescence on 48 primary NB samples of different tumour stages. H2AFX activity as an oncogene has been further validated in vitro by silencing the molecule in two NB cell lines, characterised by MYCN amplified or not, and performing cell growth and migration assays.
RESULTS: A strong inverse association between H2AFX expression and patients\' survival was observed and confirmed by immunofluorescence results on primary NB tissue sections. Cox regression analysis also disclosed H2AFX as an independent predictor of EFS and OS. The gene-silencing experiments strongly suggested an oncogenic role for H2AFX on NB cells, regardless of MYCN amplification.
CONCLUSIONS: H2AFX is a prognostic marker for unfavourable NB and could be considered a target for therapeutic interventions.
摘要：
背景：神经母细胞瘤（NB）是儿童时期最常见的实体瘤，并在交感神经系统中上升。这里,我们讨论了参与DNA损伤反应的H2AFX基因表达之间的关联的计算机分析，和786例NB患者的生存率。
方法：从Cangelosi等人总结的公开可用数据集中检索了计算机基因表达。,包括发病时786例NB肿瘤的13,696个基因表达谱。通过Kaplan-Meier和Cox回归分析评估H2AFX（H2A组蛋白家族成员X）基因表达对无事件生存期（EFS）和总生存期（OS）的预后价值。主要结果在另一个公开访问的计算机数据库（NRC-283）上进行了验证，该数据库在283例NB患者中包含13,489个基因表达。然后通过免疫荧光在不同肿瘤分期的48个原代NB样品上测试H2AFX蛋白的表达。H2AFX作为癌基因的活性已通过在两个NB细胞系中沉默该分子在体外得到进一步验证，以MYCN扩增或不扩增为特征，并进行细胞生长和迁移测定。
结果：观察到H2AFX表达与患者生存之间的强烈负相关，并通过对原发性NB组织切片的免疫荧光结果证实。Cox回归分析还显示H2AFX是EFS和OS的独立预测因子。基因沉默实验强烈提示H2AFX在NB细胞上的致癌作用，无论MYCN扩增。
结论：H2AFX是不良NB的预后标志物，可被视为治疗干预的目标。
方法：从Cangelosi等人总结的公开可用数据集中检索了计算机基因表达。,包括发病时786例NB肿瘤的13,696个基因表达谱。通过Kaplan-Meier和Cox回归分析评估H2AFX（H2A组蛋白家族成员X）基因表达对无事件生存期（EFS）和总生存期（OS）的预后价值。主要结果在另一个公开访问的计算机数据库（NRC-283）上进行了验证，该数据库在283例NB患者中包含13,489个基因表达。然后通过免疫荧光在不同肿瘤分期的48个原代NB样品上测试H2AFX蛋白的表达。H2AFX作为癌基因的活性已通过在两个NB细胞系中沉默该分子在体外得到进一步验证，以MYCN扩增或不扩增为特征，并进行细胞生长和迁移测定。
结果：观察到H2AFX表达与患者生存之间的强烈负相关，并通过对原发性NB组织切片的免疫荧光结果证实。Cox回归分析还显示H2AFX是EFS和OS的独立预测因子。基因沉默实验强烈提示H2AFX在NB细胞上的致癌作用，无论MYCN扩增。
结论：H2AFX是不良NB的预后标志物，可被视为治疗干预的目标。
