He Shiqing,Wang Meide,Sha Yongliang.Construction and validation of a prognostic model for neuroblastoma based upon glycosyltransferase genes[J].Journal of Clinical Pediatric Surgery,2026,(04):333-343.[doi:10.3760/cma.j.cn101785-20260124-00038]
基于糖基转移酶基因构建并验证神经母细胞瘤预后模型
- Title:
- Construction and validation of a prognostic model for neuroblastoma based upon glycosyltransferase genes
- Keywords:
- Neuroblastoma; Glycosyltransferases; Prognosis; Models; Statistical; Risk Assessment
- 摘要:
- 目的 基于糖基转移酶基因(glycosyltransferases,GTs)构建并验证神经母细胞瘤(neuroblastoma,NB)预后模型。方法 以基因表达综合数据库(gene expression omnibus,GEO)中的GEO85047为训练集,TARGET数据库为验证集。依据文献报道的213个GTGs,采用单因素及多因素Cox回归分析、蛋白质-蛋白质相互作用(protein-protein interaction,PPI)及最小绝对值收缩和选择算子(least absolute shrinkage and selection operator,LASSO)回归,筛选预后相关GTs并构建预后模型。计算患者风险评分,以评分中位值为阈值将患者分为低风险组和高风险组,通过Kaplan-Meier曲线、ROC曲线及Cox回归验证模型的稳定性和预测效能;采用单因素和多因素Cox回归分析评估模型及其他临床因素与肿瘤预后之间的相关性;对高、低风险组差异基因进行功能富集分析,通过单样本基因集富集分析(single sample gene set enrichment analysis,ssGSEA)探究模型与免疫细胞浸润的关系。选取81例NB肿瘤组织和25例神经节神经瘤肿瘤组织标本,采用免疫组化检测肿瘤组织中B3GALT4表达水平。选取NB细胞系9464D和975A2,分为Scramble组(转染B3GALT4敲低慢病毒空载体)、Sh-B3GALT4组(转染B3GALT4敲低慢病毒)、Vector组(转染B3GALT4过表达慢病毒空载体)及OE-B3GALT4组(转染B3GALT4过表达慢病毒),通过Western blot、实时荧光定量聚合酶链式反应(quantitative real-time polymerase chain reaction,qRT-PCR)、细胞增殖实验(cell counting kit-8,CCK-8)、集落形成实验和Transwell实验,检测细胞增殖、侵袭及迁移能力。结果 本研究成功构建由 PIGZ、GALNT7、GALNT2、GALNT14、EXTL2、DPY19L3、CHPF2、B3GNT5、B3GALT4及ALG3基因组成的预后模型。生存分析显示,高风险组患儿的预后显著劣于低风险组(P<0.01);训练集模型预测1年、3年及5年生存率曲线下面积(area under curve,AUC)分别为0.807、0.825及0.837;验证集分别为0.654、0.675及0.662。单因素、多因素Cox回归分析显示,年龄、分期和预后模型均是影响患儿预后的影响因素。高风险组差异基因主要富集于蛋白靶向内质网、细胞膜及核转录的mRNA分解过程等通路;高风险组患儿CD4+T细胞、CD8+T细胞浸润数量显著多于低风险组(P<0.05)。B3GALT4基因在NB组织中呈低表达;下调 B3GALT4可促进NB细胞的增殖、侵袭及迁移能力。结论 基于糖基转移酶基因构建的NB预后模型可有效预测患儿预后及肿瘤免疫微环境;上调 B3GALT4基因表达可抑制NB细胞恶性表型,该基因有望成为NB新的分子标志物。
- Abstract:
- Objective To construct and validate a prognosis model for neuroblastoma (NB) based on glycosyltransferases (GTs).Methods GEO85047 from the Gene Expression Omnibus (GEO) database was used as training set while the TARGET database as validation set.Based upon 213 GTGs reported in the literature,single/multi-factor Cox regression analysis,protein-protein interaction (PPI) and least absolute shrinkage and selection operator (LASSO) regression were employed for screening prognostic-related GTs and constructing a prognosis model.The risk score was calculated.They were assigned into low-risk and high-risk groups based upon median value.Kaplan-Meier curve,ROC curve and Cox regression were utilized for observing the stability and prognostic efficacy of the model.Univariate and multivariate Cox regression analyses were used for evaluating the correlation between the model,other clinical factors and tumor prognosis.Functional enrichment analysis of differentially expressed genes between high-risk and low-risk groups was performed.Single sample gene set enrichment analysis (ssGSEA) was used for examining the relationship between the model and immune cell infiltration.A total of 81 NB and 25 ganglioneuroblastoma tumor tissue specimens were selected for immunohistochemical detection of B3GALT4 expression.And 9464D and 975A2 in NB cell lines were divided into Scramble group (transfection with B3GALT4 knockdown lentiviral vector),Sh-B3GALT4 group (transfection with B3GALT4 knockdown lentiviral vector),Vector group (transfection with B3GALT4 overexpression lentiviral vector) and OE-B3GALT4 group (B3GALT4 overexpression lentiviral vector).Western blot,quantitative real-time polymerase chain reaction (qRT-PCR),CCK-8,clone formation and Transwell assay were employed for detecting cell proliferation,invasion and migratory capabilities.Results A prognosis model consisting of PIGZ,GALNT7,GALNT2,GALNT14,EXTL2,DPY19L3,CHPF2,B3GNT5,B3GALT4 and ALG3 genes was constructed.Survival analysis revealed that high-risk group had significantly worse prognosis than low-risk group (P<0.01).The AUC of the prognostic model for 1/3/5-year survival rate was 0.807,0.825 and 0.837 (training set); 0.654,0.675 and 0.662 (validation set).Univariate and multivariate Cox regression analysis indicated that age,stage and the prognosis model were independent risk factors for prognosis.The high-risk group became enriched predominantly in pathways such as protein targeting endoplasmic reticulum,cell membrane and mRNA degradation processes of nuclear transcription.The number of CD4+ T cells and CD8+ T cells infiltrating in high-risk group was higher than that in low-risk group (P<0.05).B3GALT4 was lowly expressed in NB tissues.Down-regulation of B3GALT4 could promote the proliferation,invasion and migratory capabilities of NB cells.Conclusions The NB prognosis model constructed based upon glycosyltransferase genes may predict the prognosis of children and immune microenvironment.Up-regulation of B3GALT4 arrests the malignant phenotype of NB.It is expected to become a new molecular marker for NB.
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备注/Memo
收稿日期:2026-1-24。
基金项目:江苏省卫生健康委科研项目(Z2024033);徐州医科大学附属医院发展基金资助项目(XYFM202407);徐州市卫生健康委科技项目(XWKYHT20250021);徐州医科大学附属医院发展基金资助项目(XYFM202335)
通讯作者:沙永亮,Email:sylv4_2012@sina.com