过表达带电多泡体蛋白2B基因抑制肾透明细胞癌细胞的增殖

doi:10.12122/j.issn.1673-4254.2025.01.16

摘要/Abstract

摘要：

目的肾透明细胞癌中带电多泡体蛋白2B（CHMP2B）的表达水平与临床病理特征及预后的关系，探究CHMP2B 在肾透明细胞癌发生发展中的潜在作用机制。方法从TCGA数据库下载并整理肾透明细胞癌的RNAseq数据并提取FPKM格式的数据，比较肿瘤样本与癌旁样本 CHMP2B的表达差异，根据其表达中位值分为高、低表达组，分析CHMP2B表达水平与临床病理特征的相关性，采用Kaplan-Meier模型进行生存分析，COX风险回归模型用于临床预后因素分析；使用ESTIMATE算法分析CHMP2B与免疫和基质细胞浸润及肿瘤纯度的关系；使用Limma包对CHMP2B筛选共表达基因，并进一步行GSVA富集分析。使用ESTIMATE算法和Timer数据库分析CHMP2B表达与免疫浸润的相关性；使用cBioPortal数据库分析CHMP2B的基因突变对肾透明细胞癌免疫治疗的影响；从HPA数据库获取CHMP2B蛋白质在肾透明细胞癌组织和正常组织的免疫组化表达情况；通过收集健康与肾癌患者外周血标本评价CHMP2B作为临床标志物的可能。通过细胞实验验证CHMP2B对肾癌细胞增殖与迁移的影响。结果与癌旁组织相比较，CHMP2B在肾透明细胞癌组织中低表达，使其过表达会抑制肿瘤细胞增殖（P<0.01），其表达水平与年龄、性别、是否淋巴结转移以及分期等具有相关性（P<0.05）。生存分析结果显示CHMP2B低表达肾透明细胞癌患者生存预后更差（P<0.05），富集差异分析及共表达基因富集发现CHMP2B 在肾透明细胞癌中主要涉及病毒出芽、坏死性凋亡、内吞作用等，并涉及免疫调节过程。结论 CHMP2B在肾透明细胞癌组织中低表达，可有效影响肿瘤进展和肿瘤免疫过程，是一种有前途的预后分子标志物及治疗靶点。

关键词: 肾透明细胞癌, 带电多泡体蛋白2B, 内体分选复合物, 预后, 免疫

Abstract:

Objective To analyze the association of CHMP2B expression level of with clinicopathological characteristics and prognosis of clear cell renal cell carcinoma (CRCC) and the possible role of CHMP2B in tumorigenesis and progression of CRCC. Methods RNAseq data of CRCC were downloaded from the TCGA database for analysis of CHMP2B expression levels in tumor and adjacent tissues and their correlation with clinicopathological characteristics of the patients. Survival outcomes of the patients with high and low CHMP2B expressions were analyzed using the Kaplan-Meier model, and the COX risk regression model was used for identifying the prognostic factors of the patients. The correlation between CHMP2B expression and immune infiltration, its co-expressed genes, and the effect of CHMP2B gene mutations on immunotherapy responses, and its immunohistochemical expression in CRCC and normal tissues were analyzed. Clinical samples of CRCC were collected to examine CHMP2B expressions using RT-PCR, and cell experiment was carried out to test the effect of CHMP2B overexpression on biological behaviors of CRCC cells. Results CHMP2B was significantly under-expressed in renal cancer tissues, and its overexpression obviously inhibited the proliferation of CRCC cells in vitro. CHMP2B expression level was significantly correlated with age, gender, lymph node metastasis, and tumor stage, and the patients with low CHMP2B expression had poor survival outcomes. Enrichment and co-expression gene analyses suggested that CHMP2B was mainly involved in viral outgrowth, necrotic apoptosis, endocytosis, and immune-regulatory processes in kidney cancer. Conclusion CHMP2B is lowly expressed in renal cancer tissues to affect tumor progression and tumor immune processes, and may serve as a prognostic biomarker and therapeutic target for CRCC.

Key words: renal clear cell carcinoma, charged multivesicular body protein 2B, endosomal sorting complex required for transport, prognosis, immunity

陈晓睿, 魏青政, 张宗亮, 原江水, 宋卫青. 过表达带电多泡体蛋白2B基因抑制肾透明细胞癌细胞的增殖[J]. 南方医科大学学报, 2025, 45(1): 126-136.

Xiaorui CHEN, Qingzheng WEI, Zongliang ZHANG, Jiangshui YUAN, Weiqing SONG. Overexpression of CHMP2B suppresses proliferation of renal clear cell carcinoma cells[J]. Journal of Southern Medical University, 2025, 45(1): 126-136.

图/表 12

图1 CHMP2B在KIRC中的差异表达

Fig.1 Differential expression of CHMP2B in KIRC. A: Unpaired difference analysis. B: Paired difference analysis.

图2 CHMP2B生存分析

Fig.2 CHMP2B survival analysis in CRCC patients. A: CHMP2B ROC curve for predicting overall survival (The x axis is the false positive rate, and the x axis the true positive rate). B: Effect of high and low CHMP2B expression on overall survival of KIRC patients. C: Effect of high and low CHMP2B expression on disease-specific survival of KIRC.

图3 CHMP2B表达与KIRC临床特征的相关性分析

Fig.3 Correlation analysis between CHMP2B expression and clinical characteristics of CRCC patients. A: Relationship between CHMP2B expression and T stage. B: Relationship between CHMP2B expression and N stage. C: Relationship between CHMP2B expression and M stage. D: Relationship between CHMP2B expression and grade. E: Relationship between CHMP2B expression and gender. F: Relationship between CHMP2B expression and age. ***P<0.001.

图4 COX回归分析

Fig.4 COX regression analysis. A: Univariate analysis. B: Multivariate analysis. C: Nomogram of survival rate.

图5 CHMP2B在KIRC中与ESCRT家族相关性分析、GO分析以及KEGG富集分析

Fig.5 CHMP2B was correlated with ESCRT family analysis, GO analysis and KEGG enrichment analysis in KIRC.. A: Relationship between CHMP2B and ESCRT family genes. B: GO analysis of CHMP2B. C: KEGG analysis of CHMP2B.

图6 GSVA富集分析

Fig.6 GSVA enrichment analysis. A: KEGG analysis of CHMP2B. B: GO analysis of CHMP2B.

图7 CHMP2B在泛癌中的差异表达

Fig.7 Differential expression of CHMP2B in pan-cancer. *P<0.05, **P<0.01, ***P<0.001.

图8 CHMP2B与肿瘤微环境相关性分析

Fig.8 Correlation analysis between CHMP2B and tumor microenvironment. A: CHMP2B immune score. B: Relationship between CHMP2B expression and immune cells. C: Differential expression of CHMP2B in immune cells. *P<0.05, **P<0.01, ***P<0.001.

图9 CHMP2B免疫相关分析

Fig.9 CHMP2B immune correlation analysis. A: Heatmap of related immune checkpoint. B: Scatter plot of immune mutation. C: Relationship between high and low CHMP2B expression and treatment with ctla 4 and pd 1.

图10 CHMP2B免疫组化

Fig.10 CHMP2B immunohistochemistry. A: Renal clear cell carcinoma tissue. B: Normal kidney tissue.

图11 RT-PCR分析CHMP2B差异表达

Fig.11 Differential CHMP2B expression analyzed using RT-PCR. ****P<0.0001 vs NC.

图12 CHMP2B对透明细胞癌细胞786-O生物学行为的影响

Fig.12 Effects of CHMP2B on the biological behaviors of clear cell carcinoma cells 786-O. A: Expression level of CHMP2B gene in the transfected cells detected with real-time PCR. B: Change of cell proliferation rate assessed with CCK-8 experiment. C: Effect of overexpression of CHMP2B on horizontal migration ability of the cells (Original magnification: ×100). D: Effect of overexpression CHMP2B on vertical migration ability of cells in Transwell experiment (crystal violet staining, ×100). ****P<0.0001 vs Vector-NC.

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