Elsevier

Advances in Medical Sciences

Volume 65, Issue 2, September 2020, Pages 442-456
Advances in Medical Sciences

Original research article
Clinical significance of CCNE2 protein and mRNA expression in thyroid cancer tissues

https://doi.org/10.1016/j.advms.2020.09.001Get rights and content

Abstract

Purpose

Thyroid carcinoma (TC) is the most common endocrinal malignancy worldwide. Cyclin E2 (CCNE2), a member of the cyclin family, acts as a regulatory subunit of cyclin-dependent kinases (CDKs). It controls the transition of quiescent cells into the cell cycle, regulates the G1/S transition, promotes DNA replication, and activates CDK2. This study explored the role and potential molecular mechanisms of CCNE2 expression in TC tissues.

Material/methods

Immunohistochemistry was used to evaluate the CCNE2 protein expression levels in TC. High-throughput data on CCNE2 in TC were obtained from RNA sequencing (RNA-seq), microarray, and literature data. The CCNE2 expression levels in TC were comprehensively assessed through an integrated analysis. Analyses of Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and protein–protein interaction (PPIs) data facilitated the investigation of the relative molecular mechanisms of CCNE2 in TC.

Results

The immunohistochemical experiment showed a significant increase in the expression of CCNE2 in the TC tissues. For 505 TC and 59 non-cancerous samples from RNA-seq data, the area under the curve (AUC) was 0.8016 (95% confidence interval [CI] 0.742–0.8612; p<0.001). With another 14 microarrays, the pool standard mean difference [SMD] was 1.01 (95% CI [0.82–1.19]). The pooled SMD of CCNE2 was 1.12 (95% CI [0.60–1.64]), and the AUC was 0.87 (95% CI [0.84–0.90]) for 1157 TC samples and 366 non-cancerous thyroid samples from all possible sources. Nine hub genes were upregulated in TC.

Conclusions

A high expression of CCNE2 may lead to carcinogenesis and the development of TC.

Introduction

The incidence of thyroid cancer (TC) has increased dramatically worldwide in recent decades [[1], [2], [3]]. On the basis of the cell type of origin and growth patterns, four types of TCs have been identified: differentiated papillary, follicular, medullary, and undifferentiated anaplastic carcinomas [4]. With developments in modern medicine, TC is usually curable. Nevertheless, an effective diagnostic approach with high accuracy and a risk evaluation method is needed. A recent study demonstrated that TC could exhibit divergent clinical behavior, with some cases exceeding expectations [5]. Ultrasound-guided fine needle aspiration biopsy, which is widely used in the diagnosis of TC, has limitations, especially in terms of the follicular lesions labeled “indeterminate” or “nondiagnostic” and those suspicious for malignancy according to the Bethesda system of thyroid cytology classification [5,6]. The surgical strategies and prognosis evaluations in TC depend on the diagnosis and risk estimations. An understanding of the molecular mechanisms of TC would facilitate improvements in diagnosis and therapy [7].

The cyclin E2 (CCNE2) gene, located at 8q22.1, encodes CCNE2 in humans and regulates the G1/S transition [[8], [9], [10]]. In addition to playing a role in cell division, CCNE2 promotes DNA replication and activates cyclin-dependent kinase 2 (CDK2), which has important functions in cellular biological processes [[8], [9], [10]]. The accumulating evidence suggests that CCNE2 can act as a proto-oncogene in cancers. For example, in castration-resistant prostate cancer, CCNE2 expression is downregulated, leading to cancer suppression [11]. Aberrant CCNE2 expression has been found in a variety of cancers [[11], [12], [13]]. However, the specific roles and underlying mechanisms of CCNE2 in the development and progression of TC remain elusive.

Therefore, to verify the changes in CCNE2 expression in TC and to investigate the underlying molecular mechanisms, immunohistochemical (IHC), RNA-sequencing (RNA-seq), and microarray data were analyzed to determine the clinical value of CCNE2 expression in TC. The co-expressed genes of CCNE2 were also identified. The enrichment pathway of the CCNE2 co-expressed and hub genes was assessed through Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment, as well as protein–protein interaction (PPI) network analysis. The discoveries of this study could increase the understanding of the role of CCNE2 in the development and progression of TC.

Section snippets

Immunohistochemical detection of cyclin E2 protein levels

Two tissue microarrays, THC961 and THC1021, were obtained from Fanpu, Inc. (Guilin, China). They consisted of 23 non-cancerous thyroid tissues and 125 TC tissues. The TC tissues included papillary carcinoma (PTC), follicular carcinoma (FTC), and anaplastic carcinoma (ATC)/undifferentiated thyroid carcinoma (UTC) tissues. In addition, 64 non-cancerous thyroid tissues and 46 TC tissues from patients who came to the Department of Pathology at the First Affiliated Hospital of Guangxi Medical

Immunohistochemical investigation of cyclin E2 protein levels

A total of 171 TC and 87 non-cancerous tissue samples were obtained from the First Affiliated Hospital of Guangxi Medical University and the Human Protein Atlas (Table 1; Fig. 1, Fig. 2). The CCNE2 expression levels in the 171 TC samples (7.21 ± 1.631) were statistically significantly higher than those in the 87 non-cancerous tissue samples (1.22 ± 1.497; p < 0.001; Fig. 3a). The AUC, which reflected the capacity of CCNE2 expression to make a distinction between the TC and non-cancerous

Discussion

This study had two goals: (1) the exploration and verification of the changes in CCNE2 expression in TC and (2) the preliminary investigation of the possible underlying mechanisms of CCNE2 in TC by means of the CCNE2 co-expression genes. The 1157 TC and 366 non-cancerous thyroid samples, which were combined from the IHC experiment, and the RNA-seq and microarray data were considered. The expression level of CCNE2 in the TC tissues was significantly higher than that in the non-cancerous tissues.

Conclusion

In summary, the over-expression of CCNE2 in TC was demonstrated through the analysis of TC and normal samples from all possible sources, including immunohistochemistry, TCGA, microarrays, and the literature. CCNE2 was found to be highly correlated with the cell cycle. Nevertheless, the specific molecular mechanisms in CCNE2, which promotes the biological aggressiveness of TC, need further study.

Data statement

Data used for meta-analysis and gene expression analysis was extracted from previously published papers and publicly available databases.

Financial disclosure

The current study was supported by Funds of Guangxi Zhuang Autonomous Region Health and Family Planning Commission Self-Financed Scientific Research Project (Z2012053), National Natural Science Foundation of China (NSFC81960329), Natural Science Foundation of Guangxi, China (2017GXNSFAA198253) and the Guangxi Zhuang Autonomous Region University Student Innovative Plan (201910598148).

The Author Contribution

Study Design: Cui-Zhen Liu, Jin-Bo Peng, Gang Chen, Peng Lin, Hong Yang, Yun He, Yu-Yan Pang, Wei Ma.

Data Collection: Wan-ping Guo, Peng Lin, Xiao-Li Huang, Xiao-Fan Liu, Yu-Yan Pang, Wei Ma.

Statistical Analysis: Wan-ping Guo, Peng Lin, Xiao-Li Huang, Xiao-Fan Liu.

Data Interpretation: Wan-ping Guo, Peng Lin, Xiao-Li Huang, Xiao-Fan Liu, Gang Chen, Peng Lin, Hong Yang, Yun He.

Manuscript Preparation: Cui-Zhen Liu, Wan-Ping Guo, Xiao-Li Huang, Xiao-Fan Liu, Peng Lin, Yu-Yan Pang, Wei Ma.

Literature

Declaration of competing interest

The authors declare no conflict of interests.

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