单细胞RNA测序与甲状腺癌中具有预后和免疫微环境的时钟昼夜节律调节因子的批量RNA测序分析相结合。

Single-cell RNA sequencing integrated with bulk RNA sequencing analysis of clock circadian regulator with prognostic and immune microenvironment in thyroid cancer.

作者信息

Shen Zhen, Zhao Yuelei, Xu Xinxin, Yang Huini, He Shuting, Ma Junchi, Zhang Shaoqiang, Hou Peng, Sui Fang

机构信息

Department of Otorhinolaryngology-Head and Neck Surgery, The First Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710061, Shaanxi Province, PR China.

Department of Endocrinology, The First Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710061, Shaanxi Province, PR China.

出版信息

Transl Oncol. 2025 Mar;53:102299. doi: 10.1016/j.tranon.2025.102299. Epub 2025 Jan 31.

DOI:10.1016/j.tranon.2025.102299

PMID:39892222

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC11833347/

Abstract

BACKGROUND

Disruption of circadian rhythm was found to be associated with immune infiltration and thyroid cancer. However, the role of clock circadian regulator (CLOCK) in the progression of thyroid cancer and its immune microenvironment remains largely unexplored. Therefore, our aim was to explore the role and potential mechanism of CLOCK in thyroid cancer.

METHODS

Single cell sequencing analysis and bulk RNA sequencing analysis was used for LASSO regression and Kaplan-Meier survival estimates. Potential mechanism analysis were gained through KEGG/GO analysis, GSEA analysis and PPI network. In vivo and in vitro experiment was used for further validation.

RESULTS

The result showed CLOCK protein was overexpressed in thyroid cancer compared with normal tissue in both thyroid specific mouse model and human sample. A prognostic model incorporating CLOCK and other related genes (FAT4, OR6K2, STK40, TMEM63A, HRCT1, SUPT5H, and OR2C3) was developed using LASSO regression. Functional assay and bioinformatics analysis indicated that CLOCK knockdown hindered tumor growth and the activity of MAPK signaling. Besides, analyses of gene enrichment, signaling pathways, and immune checkpoints suggested that CLOCK might inhibit immune infiltration within the tumor microenvironment. Confirmatory in vitro experiments and immunohistochemical assays in human samples further linked high CLOCK expression to reduced T cell cytotoxicity and infiltration.

CONCLUSION

These findings underscore the pivotal role of CLOCK in thyroid cancer prognosis and immune suppression, highlighting its potential as a target for therapeutic intervention and prognostic assessment in thyroid cancer management.

摘要

背景

昼夜节律紊乱被发现与免疫浸润和甲状腺癌有关。然而，生物钟昼夜调节因子（CLOCK）在甲状腺癌进展及其免疫微环境中的作用仍 largely 未被探索。因此，我们的目的是探讨 CLOCK 在甲状腺癌中的作用及潜在机制。

方法

单细胞测序分析和批量 RNA 测序分析用于 LASSO 回归和 Kaplan-Meier 生存估计。通过 KEGG/GO 分析、GSEA 分析和 PPI 网络进行潜在机制分析。体内和体外实验用于进一步验证。

结果

结果显示，在甲状腺特异性小鼠模型和人类样本中，与正常组织相比，甲状腺癌中 CLOCK 蛋白均过表达。使用 LASSO 回归建立了一个包含 CLOCK 和其他相关基因（FAT4、OR6K2、STK40、TMEM63A、HRCT1、SUPT5H 和 OR2C3）的预后模型。功能测定和生物信息学分析表明，敲低 CLOCK 可阻碍肿瘤生长和 MAPK 信号通路的活性。此外，基因富集、信号通路和免疫检查点分析表明，CLOCK 可能抑制肿瘤微环境中的免疫浸润。在体外进行的验证实验和在人类样本中进行的免疫组织化学分析进一步将高 CLOCK 表达与降低的 T 细胞细胞毒性和浸润联系起来。