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. 2023 Mar 30:13:1113781.
doi: 10.3389/fonc.2023.1113781. eCollection 2023.

Comprehensive analysis reveals CCDC60 as a potential biomarker correlated with prognosis and immune infiltration of head and neck squamous cell carcinoma

Zhixin Liu  1 , Shuai Chen  2   3 , Wenming Jia  2   3 , Ye Qian  2   3 , Xiaoqi Yang  2   3 , Minfa Zhang  2   3 , Tianhe Fang  4 , Heng Liu  2   3
Affiliations

Comprehensive analysis reveals CCDC60 as a potential biomarker correlated with prognosis and immune infiltration of head and neck squamous cell carcinoma

Zhixin Liu et al. Front Oncol. .

Abstract

Background: Coiled-coil domain containing 60 (CCDC60) is a member of the CCDC family, which participates in the progression of many types of cancer. However, the prognostic value of CCDC60 in head and neck squamous cell carcinoma (HNSC) and its function in tumor immunity remain unclear.

Methods: CCDC60 expression and its prognostic potential in HNSC were evaluated by bioinformatics approaches, which was validated in human HNSC samples. Genetic alteration analysis of CCDC60 and the underlying biological function of CCDC60 related co-expressed genes in HNSC were analyzed. The impact of CCDC60 on the regulation of immune infiltration in HNSC was comprehensively investigated. In vitro, a series of functional assays on CCDC60 were performed in HNSC cells.

Results: Our study has indicated that compared with the adjacent normal tissues, CCDC60 expression was considerably downregulated in HNSC tissues. High CCDC60 expression was connected with favorable outcome of HNSC patients, and its prognostic significance was examined by distinct clinical characteristics. We identified the CCDC60-related co-expression genes, which were mainly enriched in the NOD-like receptor signaling pathway associated with the inhibition of tumor growth, leading to a better prognosis of HNSC patients. In vitro, CCDC60 overexpression significantly inhibited the growth, migration and invasiveness but regulated cell cycle progression, and promoted cell adhesion of Fadu and Cal27 cells. Additionally, high CCDC60 expression had strong connections with the infiltrating levels of immune cells, immune marker sets, immunomodulators and chemokines in HNSC, suggesting that targeting CCDC60 could be a promising strategy to enhance the efficacy of immunotherapy for HNSC patients.

Conclusion: Tumor suppressor CCDC60 may be identified as a prognostic and immune-related indicator in HNSC, which had the potential functions in regulating the immune infiltration of HNSC and improving the response to immunotherapy for HNSC patients.

Keywords: CCDC60; hNSC; immune infiltration; immunotherapy; prognosis.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
The expression level of CCDC60 in HNSC. (A) The mRNA expression of CCDC60 in different types of human cancers via the TIMER2.0 database. (B) Decreased CCDC60 expression in HNSC samples from the TCGA database (n= 546). (C) ROC curve analysis of CCDC60 gene for HNSC diagnosis. (D) CCDC60 mRNA expression levels in 15 pairs of HNSC tissues without lymph node metastasis (E) CCDC60 mRNA expression levels in 8 pairs of HNSC tissues with lymph node metastasis. Box-whisker plots showing the CCDC60 transcription in subgroups of HNSC patients, stratifed based on individual cancer stage (F), node metastasis status (G), tumor grade (H), patient gender (I), age (J) and TP53 mutation (K). *P < 0.05, **P < 0.01, ***P < 0.001.
Figure 2
Figure 2
Genetic Alteration and Survival Analysis of CCDC60 in HNSC. (A) The alteration frequency with mutational types of CCDC60. (B) CCDC60 copy number variations in HNSC. (C) Survival analysis of CCDC60 altered group and unaltered group. (D, E) Survival curves of overall survival (OS) and disease-specific survival (DSS) of CCDC60 in HNSC patients from the DriverDBv3 database. (F, G) Survival curves of OS and relapse-free survival (RFS) of CCDC60 in the HNSC cohort by Kaplan-Meier plotter.
Figure 3
Figure 3
Functional analysis of CCDC60 related co-expressed genes in HNSC. (A) The volcano plot of CCDC60 co-expression genes. The red dots represented genes that are positively related to CCDC60, while the green dots are the opposite. (B) Protein-protein interaction (PPI) network of CCDC60-related genes by the GeneMANIA databse. (C, D) The top 50 significantly positive or negative genes which linked to CCDC60 in HNSC. Functional analysis of CCDC60 co-expression genes in HNSC samples are available at the LinkedOmics database, including Biological processes, BP (E); Cellular components, CC (F); Molecular functions, MF (G); KEGG pathway (H).
Figure 4
Figure 4
CCDC60 inhibited the proliferation, regulated cell cycle progression of HNSC cells. (A) Colony formation assay and the related analysis in Fadu and Cal27 cells treated as indicated. (B) Growth curves of Fadu and Cal27 cells with indicated treatment were examined by CCK-8 assay. (C) Apoptotic rate of Fadu and Cal27 cells with indicated treatment were detected via flow cytometry. (D) The cell cycle distribution of Fadu and Cal27 cells treated as indicated. *P < 0.05, **P < 0.01.
Figure 5
Figure 5
CCDC60 suppressed the migration and invasion, promoted cell adhesion of HNSC cells. (A) Wound-healing assay and the related analysis of Fadu and Cal27 cells treated as indicated. (B) Transwell assay and the related analysis of Fadu and Cal27 cells with indicated treatment. (C) Cell adhesion assay and the related analysis of Fadu and Cal27 cells treated as indicated. *P < 0.05, **P < 0.01, ***P < 0.001.
Figure 6
Figure 6
Relationship between CCDC60 expression and tumor-infiltrating lymphocytes in HNSC via the TIMER2.0 database.
Figure 7
Figure 7
Correlation analysis of immune markers with CCDC60 expression in HNSC. Scatterplots of relationships between CCDC60 and gene markers of M1 Macrophage (A), T Cell Exhaustion (B), TAM (C), Treg (D) and Dendritic Cell (E) in HNSC (n = 520).
Figure 8
Figure 8
CCDC60 expression was correlated with immunomodulators in HNSC. (A) Relationship of CCDC60 expression with immunoinhibitors in HNSC samples using the TISIDB database. (B) Relationship of CCDC60 expression with immunostimulators in HNSC samples using the TISIDB database.
Figure 9
Figure 9
CCDC60 expression was correlated with chemokines in HNSC. (A) Relationship of CCDC60 expression with chemokines in HNSC samples using the TISIDB database. (B) Relationship of CCDC60 expression with chemokine receptors in HNSC samples using the TISIDB database.
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