| 19 | 0 | 6 |
| 下载次数 | 被引频次 | 阅读次数 |
本研究通过整合生物信息学分析与功能实验验证，系统解析环氧化物水解酶4(epoxide hydrolase 4, EPHX4)在胰腺导管腺癌(pancreatic ductal adenocarcinoma, PDAC)中的生物学特性及临床价值。基于TCGA、 GTEx和HPA数据库的多组学分析EPHX4在泛癌及胰腺导管腺癌中的表达水平，通过Kaplan-Meier法评估EPHX4 mRNA水平与PDAC患者生存率之间的相关性，基于单因素和多因素Cox回归分析及Nomogram列线图探究EPHX4基因在PDAC中的表达水平及其与患者临床信息和预后的相关性，利用UALCAN数据库分析EPHX4在PDAC中与不同临床特征、癌症分期分级的相关性，并构建EPHX4相关蛋白网络和进行功能富集分析，通过不同算法和GEPIA数据库对EPHX4在免疫浸润细胞中的表达进行预测。通过CCK-8、集落形成、细胞划痕和Transwell实验检测敲低EPHX4基因表达对细胞增殖、存活、迁移和侵袭能力的影响。结果显示，EPHX4在泛癌组织中普遍高表达(P<0.001),其中胰腺导管腺癌组织中的EPHX4 mRNA和蛋白质表达水平显著高于正常组织。临床预后分析表明，EPHX4高表达组患者中位总生存期显著缩短(HR=1.32, 95%CI:1.14～1.53,P=0.000 2),且与PDAC患者的EPHX4表达量、年龄及病理N分期密切相关。基因富集通路分析的数据显示，EPHX4可能参与调控MAPK、 PI3K-Akt、 Wnt等多个与PDAC进展相关的生物通路，免疫浸润和免疫检查点分析结果显示，EPHX4的表达水平与肿瘤微环境中初始CD4~+T细胞、 M2型巨噬细胞及活化的CD8 T细胞的含量呈显著负相关(P<0.05)。体外实验证实，在BXPC-3和PANC-1细胞中敲低EPHX4基因后均能显著抑制胰腺导管腺癌PDAC细胞的体外增殖、迁移和侵袭能力，其增殖、迁移和侵袭能力均降低至对照组的50%以上，差异有统计学意义(P<0.05)。本研究首次揭示EPHX4作为胰腺导管腺癌独立预后标志物的临床价值，其通过多维度调控肿瘤恶性表型和免疫微环境的分子机制为胰腺导管腺癌精准治疗提供了新靶点。
Abstract：This study systematically elucidated the biological characteristics and clinical significance of epoxide hydrolase 4(epoxide hydrolase 4, EPHX4) in pancreatic ductal adenocarcinoma(PDAC) by integrating bioinformatics analysis and functional experimental validation. Multi-omics analysis based on the TCGA, GTEx, and HPA databases revealed the expression levels of EPHX4 in PDAC. The Kaplan-Meier method was employed to assess the association between EPHX4 mRNA levels and the survival rate of PDAC patients. Univariate and multivariate Cox regression analyses, along with a nomogram, were used to explore the correlation between EPHX4 expression levels and clinical characteristics as well as patient prognosis in PDAC. The UALCAN database was utilized to analyze the relationship between EPHX4 expression and various clinical features, cancer stages, and grades in PDAC. Additionally, an EPHX4-associated protein interaction network was constructed, and functional enrichment analysis was performed. Different algorithms and the GEPIA database were applied to predict EPHX4 expression in immune-infiltrating cells. The effects of EPHX4 knockdown on cell proliferation, survival, migration, and invasion were evaluated using CCK-8, colony formation, wound healing, and Transwell assays. The results demonstrated that EPHX4 was widely overexpressed in PDAC tissues(P<0.001), with significantly higher mRNA and protein expression levels in PDAC tissues compared to normal tissues. Clinical prognostic analysis revealed that patients with high EPHX4 expression had a significantly shorter median overall survival(HR=1.32, 95% CI 1.14～1.53, P=0.000 2). Furthermore, EPHX4 expression was closely associated with clinical-pathological features, cancer stage, and grade in PDAC patients, including age and pathological N stage. Gene set enrichment pathway analysis suggested that EPHX4 may be involved in regulating multiple biological pathways related to PDAC progression, such as MAPK, PI3K-Akt, and Wnt signaling. Results of immune infiltration and immune checkpoint analysis showed that the expression level of EPHX4 was significantly negatively correlated with the abundances of naive CD4~+T cells, M2 type macrophages, and activated CD8 T cells in the tumor microenvironment(P < 0.05). In vitro experiments confirmed that knockdown of EPHX4 in BxPC-3 and PANC-1 cells significantly inhibited the proliferation, migration, and invasion capabilities of PDAC cells, reducing these functions to less than 50% of the control group, with statistically significant differences(P < 0.05). This study for the first time reveals the clinical value of EPHX4 as an independent prognostic marker in pancreatic ductal adenocarcinoma. Its molecular mechanism, which multi-dimensionally regulates tumor malignant phenotypes and the immune microenvironment, provides a novel target for precision therapy.
ANBIL S,REISS K A,2024.Targeting BRCA and PALB2 in pancreatic cancer.Curr.Treat.Options Oncol.,25(3):346-363.
BEETHAM J K,TIAN T,HAMMOCK B D,1993.cDNA cloning and expression of a soluble epoxide hydrolase from human liver.Arch.Biochem.Biophys.,305(1):197-201.
BRAY F,LAVERSANNE M,SUNG H,et al.,2024.Global cancer statistics 2022:estimates of incidence and mortality worldwide for 36 cancers in 185 countries.CA Cancer J.Clin.,74(3):229-263.
CHEN Y X,WU W,JIN C X,et al.,2023.Integrating single-cell RNA-seq and bulk RNA-seq data to explore the key role of fatty acid metabolism in breast cancer.Int.J.Mol.Sci.,24(17):13209.
DE Sá S V,CORRêA-GIANNELLA M L,MACHADO M C,et al.,2007.Serpin peptidase inhibitor clade a member 1 as a potential marker for malignancy in insulinomas.Clin.Cancer Res.,13(18 pt 1):5322-5330.
DECKER M,ADAMSKA M,CRONIN A,et al.,2012.EH3 (ABHD9):the first member of a new epoxide hydrolase family with high activity for fatty acid epoxides.J.Lipid Res.,53(10):2038-2045.
FENG Z,LIU Z,PENG K,et al.,2021.A prognostic model based on nine DNA methylation-driven genes predicts overall survival for colorectal cancer.Front.Genet.,12:779383.
FERREIRA M R,SANTOS G A,BIAGI C A,et al.,2021.GSVA score reveals molecular signatures from transcriptomes for biomaterials comparison.J.Biomed.Mater Res.A,109(6):1004-1014.
FLEBBE H,HAMDAN F H,KARI V,et al.,2019.Epigenome mapping identifies tumor-specific gene expression in primary rectal cancer.Cancers (Basel),11(8):E1142.
HU Z I,O'REILLY E M,2024.Therapeutic developments in pancreatic cancer.Nat.Rev.Gastroenterol.Hepatol.,21(1):7-24.
KARAMITOPOULOU E,2023.Emerging prognostic and predictive factors in pancreatic cancer.Mod.Pathol.,36(11):100328.
KLEEFF J,KORC M,APTE M,et al.,2016.Pancreatic cancer.Nat.Rev.Dis.Primers.,2:16022.
LI A Q,ZHANG K,ZHOU J J,et al.,2024a.Bioinformatics and experimental approach identify lipocalin 2 as a diagnostic and prognostic indicator for lung adenocarcinoma.Int.J.Biol.Macromol.,272:132797.
LI C W,TANG Z F,ZHANG W J,et al.,2021.GEPIA2021:integrating multiple deconvolution-based analysis into GEPIA.Nucleic Acids Res,49(W1):W242-W246.
LI Y L,WU X Y,SHENG C B,et al.,2024b.IGSF8 is an innate immune checkpoint and cancer immunotherapy target.Cell,187(11):2703-2716.e23.
OESCH F,JERINA D M,DALY J W,1971.Substrate specificity of hepatic epoxide hydrase in microsomes and in a purified preparation:evidence for homologous enzymes.Arch.Biochem.Biophys.,144(1):253-261.
OESCH F,JERINA D M,DALY J W,et al.,1973.Induction,activation and inhibition of epoxide hydrase:an anomalous prevention of chlorobenzene-induced hepatotoxicity by an inhibitor of epoxide hydrase.Chem.Biol.Interact.,6(3):189-202.
OHTSUKA T,FERNANDEZ-DEL CASTILLO C,FURUKAWA T,et al.,2024.International evidence-based Kyoto guidelines for the management of intraductal papillary mucinous neoplasm of the pancreas.Pancreatology,24(2):255-270.
PARK W,CHAWLA A,O’REILLY E M,2021.Pancreatic cancer:a review.JAMA,326(9):851.
PERGOLIZZI R G,BROWER S T,2024.Molecular targets for the diagnosis and treatment of pancreatic cancer.Int.J.Mol.Sci.,25(19):10843.
QADIR R M A B,UMAIR M B,TARIQ U B,et al.,2024.Unraveling pancreatic cancer:epidemiology,risk factors,and global trends.Cureus,16(11):e72816.
REN Q,MA M,YANG J,et al.,2018.Soluble epoxide hydrolase plays a key role in the pathogenesis of Parkinson’s disease.Proc.Natl.Acad.Sci.USA,115(25):E5815-E5823.
ROBERTS D L,O'DWYER S T,STERN P L,et al.,2015.Global gene expression in pseudomyxoma peritonei,with parallel development of two immortalized cell lines.Oncotarget,6(13):10786-10800.
SHEN N M,GAO G,LU X H,et al.,2024.Comprehensive analysis of the immune implication of EPHX4 gene in laryngeal squamous cell carcinoma.Braz.J.Otorhinolaryngol.,90(4):101411.
SIEGEL R L,GIAQUINTO A N,JEMAL A,2024.Cancer statistics,2024.CA Cancer J.Clin.,74(1):12-49.
SKODA R C,DEMIERRE A,MCBRIDE O W,et al.,1988.Human microsomal xenobiotic epoxide hydrolase.Complementary DNA sequence,complementary DNA-directed expression in COS-1 cells,and chromosomal localization.J.Biol.Chem.,263(3):1549-1554.
SU P F,LI O,KE K,et al.,2024.Targeting tumor-associated macrophages:critical players in tumor progression and therapeutic strategies (review).Int.J.Oncol.,64(6):60.
TIMOFEEVA M,KROPP S,SAUTER W,et al.,2010.Genetic polymorphisms of MPO,GSTT1,GSTM1,GSTP1,EPHX1 and NQO1 as risk factors of early-onset lung cancer.Int.J.Cancer,127(7):1547-1561.
ZHAO Z,LIU M,XU Z,et al.,2022.Identification of ACSF gene family as therapeutic targets and immune-associated biomarkers in hepatocellular carcinoma.Aging (Albany NY),14(19):7926-7940.
ZHONG X B,LAI Y,2024.Special section on cytochrome P450 enzymes in toxicology and as drug targets-editorial.Drug Metab.Dispos.,52(6):476-478.
基本信息:
DOI：10.13417/j.gab.044.000633
中图分类号:R735.9
引用信息:
[1]李祎涵,贺锦涛,黄少阳等.环氧化物水解酶4基因(EPHX4)在胰腺导管腺癌中的表达及其临床意义[J].基因组学与应用生物学,2025,44(06):633-646.DOI:10.13417/j.gab.044.000633.
基金信息:
云南省科学技术厅基础研究专项(202301AT070023)资助