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甲基化驱动基因
IFFO 1在胰腺癌诊断和预后中的作用及对癌细胞生物学行为的影响Exploring the role of methylation-driven gene IFFO1 in pancreatic adenocarcinoma diagnosis, prognosis and cellular functions
- 中国癌症杂志 2024年34卷第11期 页码：998-1010
- 作者机构：
1. 南京医科大学附属上海一院临床医学院肿瘤科，上海 201600
2. 烟台大学生命科学学院，山东 烟台 264006
3. 上海交通大学医学院附属第一人民医院肿瘤中心，上海 201600
4. 复旦大学附属肿瘤医院肿瘤内科，复旦大学上海医学院肿瘤学系，上海 200032
- 作者简介：
[ "徐梓淇（0009-0001-2324-6543），硕士。" ]
王红霞（0000-0003-3481-6940），主任医师；
桑友洲，主治医师（0000-0003-2081-4258）。
- 基金信息：国家自然科学基金(82103666);上海市自然科学基金(22ZR1450000)
DOI：10.19401/j.cnki.1007-3639.2024.11.002
中图分类号： R735.9收稿：2024-08-07，
修回：2024-11-12，
纸质出版：2024-11-30
- 稿件说明：
移动端阅览
徐梓淇, 胡睿智, 李军建, 等. 甲基化驱动基因
IFFO 1在胰腺癌诊断和预后中的作用及对癌细胞生物学行为的影响[J]. 中国癌症杂志, 2024,34(11):998-1010.Ziqi XU, Ruizhi HU, Junjian LI, et al. Exploring the role of methylation-driven gene IFFO1 in pancreatic adenocarcinoma diagnosis, prognosis and cellular functions[J]. China Oncology, 2024, 34(11): 998-1010.
徐梓淇, 胡睿智, 李军建, 等. 甲基化驱动基因
IFFO 1在胰腺癌诊断和预后中的作用及对癌细胞生物学行为的影响[J]. 中国癌症杂志, 2024,34(11):998-1010. DOI： 10.19401/j.cnki.1007-3639.2024.11.002.Ziqi XU, Ruizhi HU, Junjian LI, et al. Exploring the role of methylation-driven gene IFFO1 in pancreatic adenocarcinoma diagnosis, prognosis and cellular functions[J]. China Oncology, 2024, 34(11): 998-1010. DOI： 10.19401/j.cnki.1007-3639.2024.11.002.
摘要
背景与目的：
DNA异常甲基化与肿瘤的发生、发展密切相关。本研究旨在探索胰腺癌（pancreatic adenocarcinoma，PAAD）甲基化驱动基因（methylation-driven gene，MDG）中间丝家族孤儿蛋白1（intermediate filament family orphan 1，
IFFO
1）在PAAD中的表达、对PAAD细胞侵袭转移的影响，以及作为诊断和预后标志物的潜力。
方法：
从癌症基因组图谱（The Cancer Genome Atlas，TCGA）数据库、国际癌症基因组联盟（International Cancer Genome Consortium，ICGC）数据库、高通量基因表达（Gene Expression Omnibus，GEO）数据库获取PAAD及其癌旁组织的mRNA表达数据（TCGA-PAAD-mRNA）、DNA甲基化芯片数据（TCGA-PAAD-meth、GSE53051、PACA-AU）以及健康人血液的DNA甲基化芯片数据（GSE69270）。通过差异表达分析联合差异甲基化分析筛选PAAD的MDG。在TCGA数据库中，应用Pearson相关性检验验证IFFO1启动子甲基化水平与其表达水平之间的相关性。同时，通过Kaplan-Meier生存分析评估IFFO1启动子甲基化水平、表达水平与PAAD预后的关系。27例PAAD患者的癌组织及其对应癌旁组织的病理切片来自上海交通大学医学院附属第一人民医院。本研究涉及的所有样本均通过上海交通大学医学院附属第一人民医院人类伦理委员会的审核批准（伦理编号：院伦审[2017
]
53号）。利用免疫组织化学染色（immunohistochemistry stai
ning，IHC）检测27例PAAD患者的癌组织及其对应癌旁组织中IFFO1的表达。根据IFFO1表达的中位值，将TCGA数据库中的患者分为高表达组和低表达组，并进行差异分析、基因本体论（gene ontology，GO）富集分析及基因集富集分析（gene set enrichment analysis，GSEA）。使用蛋白质印迹法（Western blot）及实时荧光定量聚合酶链反应（real-time fluorescence quantitative polymerase chain reaction，RTFQ-PCR）检验正常胰腺导管上皮细胞系H6C7与PAAD细胞系MIA PaCa2、BxPC-3、AsPC-1和Capan-2中IFFO1的表达差异。通过细胞划痕实验和侵袭实验检测过表达IFFO1对PAAD细胞AsPC-1和Capan-2的迁移和侵袭能力的影响；通过受试者工作特征（receiver operating characteristic，ROC）曲线和Kaplan-Meier生存分析评估IFFO1甲基化水平在TCGA泛癌队列中的诊断和预后预测价值。
结果：
通过对5组数据的交叉筛选，筛选出41个PAAD的MDG。其中，
IFFO
1与PAAD的预后关系最为密切[风险比（hazard ratio，HR）=0.28，
P
<
0.001
]
。IFFO1在PAAD中高甲基化而低表达，其启动子的甲基化水平与表达水平呈显著负相关关系（
r
=-0.55，
P
<
0.001）。IHC结果显示，IFFO1在PAAD组织中的表达显著低于癌旁组织（
P
<
0.05）。TCGA生存分析结果表明，高甲基化或低表达IFFO1的患者总生存期更差（
P
<
0.05）。GO和GSEA富集分析结果均表明，迁移和运动的负向调控通路在IFFO1高表达患者中富集。Western blot和RTFQ-PCR结果显示，IFFO1在正常胰腺导管上皮细胞系H6C7中的表达显著高于PAAD细胞系MIA PaCa2、BxPC-3、AsPC-1和Capan-2。过表达IFFO1可显著抑制PAAD细胞株AsPC-1和Capan-2的迁移和侵袭。进一步泛癌分析结果提示，IFFO1普遍存在启动子高甲基化而低表达的现象，其甲基化水平在多种癌症中均表现出良好的诊断和预后预测价值。
结论：
启动子高甲基化导致IFFO1在PAAD中低表达，IFFO1能够抑制PAAD细胞的侵袭和迁移能力。IFFO1甲基化有望成为PAAD诊断和预后的新型标志物。
Abstract
Background and purpose:
Abnormal DNA methylation is closely associated with the onset and progression of tumors. This study aimed to investigate the expression of intermediate filament family orphan 1 (
IFFO
1)
a methylation-driven gene (MDG) in pancreatic adenocarcinoma (PAAD)
along with its effects on the invasion and metastasis of PAAD cells
as well as its potential as a diagnostic and prognostic biomarker.
Methods:
mRNA expression data (TCGA-PAAD-mRNA)
DNA methylation data (TCGA-PAAD-meth
GSE53051
PACA-AU) of PAAD and adjacent normal tissues
as well as DNA methylation data of healthy individuals’ blood (GSE69270)
were obtained from the The Cancer Genome Atlas (TCGA)
International Cancer Genome Consortium (IC
GC) and Gene Expression Omnibus (GEO) databases. By performing differential expression analysis combined with differential methylation analysis
we screened for MDG in PAAD. In the TCGA database
Pearson correlation tests were employed to verify the relationship between IFFO1 promoter methylation level and its expression level. Additionally
Kaplan-Meier survival analysis was conducted to evaluate the relationship among IFFO1 promoter methylation level
expression level
and the prognosis of PAAD. Pathological sections of cancer tissues and corresponding adjacent tissues from 27 PAAD patients were obtained from Shanghai General Hospital
Shanghai Jiao Tong University School of Medicine. All samples involved in this study were approved by the human ethics committee of Shanghai General Hospital
Shanghai Jiao Tong University School of Medicine (ethics number: hospital ethics review[2017
]
No.53). Immunohistochemistry staining (IHC) was utilized to detect the expression of IFFO1 in cancer tissues and corresponding adjacent tissues from 27 PAAD patients. Based on the median expression level of IFFO1
patients in the TCGA database were classified into high-expression and low-expression groups. Subsequently
differential analysis
gene ontology (GO) enrichment analysis and gene set enrichment analysis (GSEA) were performed. Western blot and real-time fluorescence quantitative polymerase chain reaction (RTFQ-PCR) were employed to assess the expression variations of IFFO1 between the normal pancreatic ductal epithelial cell line H6C7 and the PAAD cell lines MIA PaCa2
BxPC-3
AsPC-1 and Capan-2. The impact of IFFO1 overexpression on the migration and invasion capacities of PAAD cell lines AsPC-1 and Capan-2 was evaluated using scratch and invasion assays. Additionally
receiver operating characteristic (ROC) curves and Kaplan-Meier survival analysis were utilized to assess the diagnostic and prognostic significance of IFFO1 methylation levels in the TCGA pan-cancer cohort.
Results:
Through the cross-screening
of five datasets
41 MDG in PAAD were identified. Among these
IFFO
1 was found to be the gene most closely associated with the prognosis of PAAD [hazard ratio (HR)=0.28
P
<
0.001
]
. IFFO1 exhibited high methylation and low expression levels in PAAD. Moreover
a significant negative correlation was observed between the methylation level of its promoter and its expression level (
r
=-0.55
P
<
0.001). IHC results indicated that IFFO1 expression was significantly lower in PAAD tissues than in adjacent non-tumor tissues (
P
<
0.05). TCGA survival analysis demonstrated that patients with high methylation or low expression of IFFO1 had poorer overall survival (
P
<
0.05). Both GO and GSEA analyses indicated that the pathway “Negative regulation of cell migration” was enriched in patients with high IFFO1 expression. Western blot and RTFQ-PCR results demonstrated that IFFO1 expression in normal pancreatic ductal epithelial cells H6C7 was significantly higher compared with PAAD cell lines MIA PaCa2
BxPC-3
AsPC-1
and Capan-2. Overexpression of IFFO1 significantly inhibited the migration and invasion of the PAAD cell lines AsPC-1 and Capan-2. Additionally
pan-cancer analysis revealed that IFFO1 exhibited abnormal promoter methylation and low expression across various cancer types
with its methylation levels demonstrating significant diagnostic and prognostic prediction value among different tumors.
Conclusion:
Promoter hypermethylation results in decreased expression of IFFO1 in PAAD. IFFO1 may suppress the invasion and migration abilities of PAAD cells. Furthermore
IFFO1 methylation holds great promise as a novel biomarker for the diagnosis and prognosis of PAAD.
关键词
Keywords
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