Clinical Significance of an m6A Reader Gene, IGF2BP2, in Head and Neck Squamous Cell Carcinoma.

The importance of N6-methyladenosine (m6A) in tumor recurrence and prognosis has been recognized in recent years. The role of m6A readers, such as insulin-like growth factor 2 mRNA-binding protein 2 (IGF2BP2), in regulating head and neck squamous cell carcinoma (HNSCC) remains unclear. We, therefore, assessed the prognostic role of IGF2BP2 in HNSCC using openly available data from The Cancer Genome Atlas (TCGA) in conjunction with HNSCC patient sample immunohistochemistry (n = 36). The correlation between IGF2BP2 expression and clinical characteristics was then examined. The role of IGF2BP2 in prognosis was assessed by Kaplan-Meier curves and Cox analysis. Finally, TCGA data was used to explore possible carcinogenic mechanisms with multi-GSEA (gene set enrichment analysis). Analysis of TCGA data and IHC results revealed that IGF2BP2 was upregulated in HNSCC tumor tissues, and the expression level was related to the T stage. Simultaneously, Kaplan-Meier curves and Cox analysis indicated that highly expressed IGF2BP2 correlated with poor prognosis and that IGF2BP2 was a potential prognostic factor for HNSCC. Gene set enrichment analysis revealed that scavenging and degradation, synthesis and metabolism, cell growth, death and motility, and cancer pathways were differentially enriched in patients with high IGF2BP2 expression. Our results demonstrate that IGF2BP2 plays an important role in tumor progression and may serve as an important biological prognostic factor for HNSCC.

Deng X ，Jiang Q ，Liu Z ，Chen W ... -  《Frontiers in Molecular Biosciences》

RNA N6-methyladenosine reader IGF2BP2 promotes lymphatic metastasis and epithelial-mesenchymal transition of head and neck squamous carcinoma cells via stabilizing slug mRNA in an m6A-dependent manner.

Lymph node metastasis is the main cause of poor prognosis of head and neck squamous carcinoma (HNSCC) patients. N6-methyladenosine (m6A) RNA modification is an emerging epigenetic regulatory mechanism for gene expression, and as a novel m6A reader protein, IGF2BP2 has been implicated in tumor progression and metastasis. However, not much is currently known about the functional roles of IGF2BP2 in HNSCC, and whether IGF2BP2 regulates lymphatic metastasis through m6A modification in HNSCC remains to be determined. The expression and overall survival (OS) probability of m6A-related regulators in HNSCC were analyzed with The Cancer Genome Atlas (TCGA) dataset and GEPIA website tool, respectively. The expression levels of IGF2BP2 were measured in HNSCC tissues and normal adjacent tissues. To study the effects of IGF2BP2 on HNSCC cell metastasis in vitro and in vivo, gain- and loss- of function methods were employed. RIP, MeRIP, luciferase reporter and mRNA stability assays were performed to explore the epigenetic mechanism of IGF2BP2 in HNSCC. We investigated 20 m6A-related regulators in HNSCC and discovered that only the overexpression of IGF2BP2 was associated with a poor OS probability and an independent prognostic factor for HNSCC patients. Additionally, we demonstrated that IGF2BP2 was overexpressed in HNSCC tissues, and significantly correlated to lymphatic metastasis and poor prognosis. Functional studies have shown that IGF2BP2 promotes both HNSCC cell migration as well as invasion via the epithelial-mesenchymal transition (EMT) process in vitro, and IGF2BP2 knockdown significantly inhibited lymphatic metastasis and lymphangiogenesis in vivo. Mechanistic investigations revealed that Slug, a key EMT-related transcriptional factor, is the direct target of IGF2BP2, and essential for IGF2BP2-regulated EMT and metastasis in HNSCC. Furthermore, we demonstrated that IGF2BP2 recognizes and binds the m6A site in the coding sequence (CDS) region of Slug and promotes its mRNA stability. Collectively, our study uncovers the oncogenic role and potential mechanism of IGF2BP2, which serves as a m6A reader, in controlling lymphatic metastasis and EMT in HNSCC, suggesting that IGF2BP2 may act as a therapeutic target and prognostic biomarker for HNSCC patients with metastasis.

Yu D ，Pan M ，Li Y ，Lu T ，Wang Z ，Liu C ，Hu G ... -  《JOURNAL OF EXPERIMENTAL & CLINICAL CANCER RESEARCH》

Molecular Characteristics, Prognostic Value, and Immune Characteristics of m(6)A Regulators Identified in Head and Neck Squamous Cell Carcinoma.

N6-methyladenosine (m6A) plays crucial roles in a diverse range of physiological and pathological processes, and it is believed that it tremendously promotes neoplasia and progression. However, knowledge of the molecular characteristics of m6A modification, its prognostic value, and the infiltration of immune cell populations in head and neck squamous cell carcinoma (HNSCC) is still insufficient. Therefore, a pan-cancer genomic analysis was systematically performed here by examining m6A regulators at the molecular level within 33 multiple cancer types, and the correlations between the expression of m6A molecules were researched using datasets from The Cancer Genome Atlas (TCGA). Based on the above analysis, insulin-like growth factor 2 mRNA-binding protein 2 (IGF2BP2) is upregulated in HNSCC and may serve as an independent prognostic factor of overall survival, thus showing potential as a prognostic biomarker in HNSCC. Genetic alteration analyses elucidated the reasons for the abnormal upregulation of IGF2BP2 in HNSCC. As a result, IGF2BP2 was selected for further univariate and multivariate analyses. The functions of the related genes were annotated through gene set enrichment analysis, and the activation states of multiple biological pathways were shown by gene set variation analysis. We found that LRRC59 and STIP1 may act as IGF2BP2-associated genes to have a regulatory function in the m6A modification. In addition, we found that the status of immune cell infiltration was correlated with the level of IGF2BP2 gene expression. Our results provide supplementation at the molecular level for epigenetic regulation in HNSCC and insight into effective immunotherapy targets and strategies.

Geng X ，Zhang Y ，Zeng Z ，Zhu Z ，Wang H ，Yu W ，Li Q ... -  《Frontiers in Oncology》

Identification of Two m6A Readers YTHDF1 and IGF2BP2 as Immune Biomarkers in Head and Neck Squamous Cell Carcinoma.

Background: N6-methyladenosine (m6A) is the most abundant internal modification pattern in mammals that a plays critical role in tumorigenesis and immune regulations. However, the effect of m6A modification on head and neck squamous cell carcinoma (HNSCC) has not been clearly studied. Methods: We screened m6A regulators that were significantly correlated with tumor immune status indicated by ImmuneScore using The Cancer Genome Atlas (TCGA) dataset and obtained distinct patient clusters based on the expression of these m6A regulators with the R package "CensusClusterPlus." We then performed gene set enrichment analysis (GSEA), CIBERSORT, and single-sample gene set enrichment analysis (ssGSEA) to assess the differences in gene function enrichment and tumor immune microenvironment (TIME) among these clusters. We further conducted differently expressed gene (DEG) analysis and weighted gene co-expression network analysis (WGCNA) and constructed a protein-protein interaction (PPI) network to determine hub genes among these clusters. Finally, we used the GSE65858 dataset as an external validation cohort to confirm the immune profiles related to the expression of m6A regulators. Results: Two m6A readers, YTHDF1 and IGF2BP2, were found to be significantly associated with distinct immune status in HNSCC. Accordingly, patients were divided into two clusters with Cluster 1 showing high expression of YTHDF1 and IGF2BP2 and Cluster 2 showing low expression levels of both genes. Clinicopathologically, patients from Cluster 1 had more advanced T stage and pathological grades than those from Cluster 2. GSEA showed that Cluster 1 was closely related to the RNA modification process and Cluster 2 was significantly correlated with immune regulations. Cluster 2 had a more active TIME characterized by a more relative abundance of CD8+ T cells and CD4+ T cells and higher levels of MHC I and MHC II molecules. We constructed a PPI network composed of 16 hub genes between the two clusters, which participated in the T-cell receptor signaling pathway. These results were externally validated in the GSE65858 dataset. Conclusions: The m6A readers, YTHDF1 and IGF2BP2, were potential immune biomarkers in HNSCC and could be potential treatment targets for cancer immunotherapy.

Li S ，Wu Q ，Liu J ，Zhong Y ... -  《Frontiers in Genetics》

Identification of m6A methyltransferase-related genes predicts prognosis and immune infiltrates in head and neck squamous cell carcinoma.

Head and neck squamous cell carcinoma (HNSCC) accounts for 90% of head and neck malignant tumors. As the early symptoms of HNSCC are not obvious, and it is prone to recurrence and metastasis, making the overall survival (OS) rate of patients very low. Existing studies have shown m6A methylation plays a crucial role in various cancers, but it is rarely studied in HNSCC. This study aimed to explore the expression of m6A methylation-related genes in HNSCC and its correlation with prognosis, and to explore its relationship with immune infiltration. The gene expression data of HNSCC patient tumor samples (tumor =510) and adjacent normal tissue samples (normal =50) were extracted from The Cancer Genome Atlas (TCGA) database, and the expression characteristics of m6A regulatory factors were described. Kaplan-Meier survival analysis was used to analyze the relationship between m6A regulatory factors and OS and disease-specific survival (DSS). Least absolute shrinkage and selection operator (LASSO) regression was used to construct the m6A regulatory factor-HNSCC risk prediction model. In addition, the relationship between m6A methylation-related genes and tumor immune infiltration were discussed. The differential expression of 20 genes were identified by TCGA, and 18 genes (IGF2BP2, IGF2BP1, IGF2BP3, VIRMA, YTHDF1, YTHDF2, YTHDF3, ZC3H13, METTL14, ALKBH5, METTL3, RBMX, WTAP, YTHDC1, FTO, HNRNPC, HNRNPA2B1, and RBM15) were overexpressed in HNSCC. The survival rate of different gene expression levels was different. The high expression of YTHDC1 and YTHDC2 indicated better OS. Furthermore, for DSS, increased expression of YTHDC2 was also correlated with better clinical outcomes (P<0.05). At the same time, we drew a 3-gene risk score model in the TCGA-HNSCC cohort, and the survival curve showed compared with low-risk patients, high-risk patients had significantly worse OS (P<0.05). Gene enrichment analysis showed EPITHELIAL_MESENCHYMAL_TRANSITIO, MTORC1_SIGNALING, MYC_TARGETS_V1, MYC_TARGETS_V2, MYOGENESIS pathways, high TP53 mutations, and suppressive immunity were related to the high-risk group. The low-risk group was related to ALLOGRAFT_REJECTION, COMPLEMENT, IL6_JAK_STAT3_SIGNALING, INTERFERON_ALPHA_RESPONSE, INTERFERON_GAMMA_RESPONSE pathways, low TP53 mutations, and active immunity. The m6A methyltransferase-related genes can predict the prognosis of HNSCC and are related to immune infiltration.

Zhang Y ，Li L ，Ye Z ，Zhang L ，Yao N ，Gai L ... -  《-》