Paired Oral Squamous Cell Carcinoma Research Articles

Unidirectional alteration of methylation and hydroxymethylation at the promoters and differential gene expression in oral squamous cell carcinoma.

Background: Oral squamous cell carcinoma (OSCC) is one of the most common types of cancer worldwide. Although overall losses of 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) have been previously observed, a genome-wide, single-base-resolution, and simultaneous mapping of 5mC and 5hmC in OSCC is still unaccomplished. Similarly, the mechanism of how 5mC and 5hmC collectively lead to abnormal gene expression in OSCC is largely unexplored. Using parallel whole-genome bisulfite sequencing (WGBS) and whole-genome oxidative bisulfite sequencing (oxWGBS), we characterized 5mC- and 5hmC-profiles at single-nucleotide resolution in paired primary OSCC samples and their normal adjacent tissues (NATs). We also analyzed the effect of 5mC- and 5hmC-modifications on differential gene expression in OSCC using multi-omics analysis. Results: An overall reduction of both 5mC and 5hmC in various genomic regions have been observed in OSCC samples. At promoter regions, a total of 6,921 differentially methylated regions and 1,024 differentially hydroxymethylated regions were identified in OSCC. Interestingly, compared to bidirectional modification with 5mC and 5hmC, unidirectional modification with 5mC and 5hmC at the promoters is associated with bigger change in the gene expression. Additionally, genes bearing unidirectional modification with 5mC and 5hmC at the promoters are enriched in signaling pathways like cell proliferation, cell differentiation, and receptor tyrosine kinase pathway that are essential for the tumorigenesis. Finally, the grouped expression signature of top 20 genes bearing promoter-unidirectional-modification with 5mC and 5hmC tends to correlate with the clinical outcome of certain subtypes of head and neck squamous cell carcinoma. Conclusion: Using parallel WGBS and oxWGBS analyses, we observed an overall reduction of 5mC- and 5hmC-modifications at various genomic regions in OSCC. Unidirectional modification with 5mC and 5hmC at the promoters is associated with enhanced changes in gene expression in OSCC tissues. Furthermore, such differentially expressed genes bearing unidirectional modifications with 5mC and 5hmC at the promoters might have clinical relevance to the outcome of OSCC.

Systematic establishment and verification of an epithelial-mesenchymal transition gene signature for predicting prognosis of oral squamous cell carcinoma.

Objective: Epithelial-mesenchymal transition (EMT) is linked to an unfavorable prognosis in oral squamous cell carcinoma (OSCC). Here, we aimed to develop an EMT gene signature for OSCC prognosis. Methods: In TCGA dataset, prognosis-related EMT genes with p < 0.05 were screened in OSCC. An EMT gene signature was then conducted with LASSO method. The efficacy of this signature in predicting prognosis was externally verified in the GSE41613 dataset. Correlations between this signature and stromal/immune scores and immune cell infiltration were assessed by ESTIMATE and CIBERSORT algorithms. GSEA was applied for exploring significant signaling pathways activated in high- and low-risk phenotypes. Expression of each gene was validated in 40 paired OSCC and normal tissues via RT-qPCR. Results: A prognostic 9-EMT gene signature was constructed in OSCC. High risk score predicted poorer clinical outcomes than low risk score. ROCs confirmed the well performance on predicting 1-, 3- and 5-year survival. Multivariate cox analysis revealed that this signature was independently predictive of OSCC prognosis. The well predictive efficacy was validated in the GSE41613 dataset. Furthermore, this signature was distinctly related to stromal/immune scores and immune cell infiltration in OSCC. Distinct pathways were activated in two subgroups. After validation, AREG, COL5A3, DKK1, GAS1, GPX7 and PLOD2 were distinctly upregulated and SFRP1 was downregulated in OSCC than normal tissues. Conclusion: Our data identified and verified a robust EMT gene signature in OSCC, which provided a novel clinical tool for predicting prognosis and several targets against OSCC therapy.

Identification of Somatic Mutations in Plasma Cell-Free DNA from Patients with Metastatic Oral Squamous Cell Carcinoma.

The accurate diagnosis and treatment of oral squamous cell carcinoma (OSCC) requires an understanding of its genomic alterations. Liquid biopsies, especially cell-free DNA (cfDNA) analysis, are a minimally invasive technique used for genomic profiling. We conducted comprehensive whole-exome sequencing (WES) of 50 paired OSCC cell-free plasma with whole blood samples using multiple mutation calling pipelines and filtering criteria. Integrative Genomics Viewer (IGV) was used to validate somatic mutations. Mutation burden and mutant genes were correlated to clinico-pathological parameters. The plasma mutation burden of cfDNA was significantly associated with clinical staging and distant metastasis status. The genes TTN, PLEC, SYNE1, and USH2A were most frequently mutated in OSCC, and known driver genes, including KMT2D, LRP1B, TRRAP, and FLNA, were also significantly and frequently mutated. Additionally, the novel mutated genes CCDC168, HMCN2, STARD9, and CRAMP1 were significantly and frequently present in patients with OSCC. The mutated genes most frequently found in patients with metastatic OSCC were RORC, SLC49A3, and NUMBL. Further analysis revealed that branched-chain amino acid (BCAA) catabolism, extracellular matrix-receptor interaction, and the hypoxia-related pathway were associated with OSCC prognosis. Choline metabolism in cancer, O-glycan biosynthesis, and protein processing in the endoplasmic reticulum pathway were associated with distant metastatic status. About 20% of tumors carried at least one aberrant event in BCAA catabolism signaling that could possibly be targeted by an approved therapeutic agent. We identified molecular-level OSCC that were correlated with etiology and prognosis while defining the landscape of major altered events of the OSCC plasma genome. These findings will be useful in the design of clinical trials for targeted therapies and the stratification of patients with OSCC according to therapeutic efficacy.

Fibroblast growth factor receptors 1 and 4 combined with lymph node metastasis predicts poor prognosis in oral cancer.

The fibroblast growth factor receptor (FGFR) members including FGFR1-4 have been identified as promising novel therapeutic targets and prognostic markers in multiple solid tumors. However, the predictive role of the expression of FGFR proteins in oral squamous cell carcinoma (OSCC) requires further exploration. Immunohistochemical evaluation of FGFR1-4 was performed on 161 paired OSCC samples. The associations of FGFRs with clinicopathologic and prognostic parameters were analyzed. To further assess the contribution of FGFRs to OSCC proliferation, cell lines, and one PDX model was utilized to examine the anti-tumor effect of the pan-FGFR inhibitor AZD4547. All FGFR members were found to be overexpressed in OSCC tumors when compared to normal tissues, and their expression was significantly associated with poor overall survival and disease-free survival. Multivariate Cox regression analysis revealed high expression of FGFR1 (p = 0.014) and FGFR4 (p = 0.009) were independent prognostic factors and co-overexpression of FGFR1 and FGFR4 with lymph node metastasis increased HR for death (p = 0.02). The pan-FGFR inhibitor AZD4547 showed anti-tumor activity in cell lines and in a patient-derived xenograft of OSCC. This study highlights the co-overexpression of FGFR1 and FGFR4 as a significantly poor prognosis indicator in OSCC when combined with lymph node metastasis.

Abnormal downregulation of 10-formyltetrahydrofolate dehydrogenase promotes the progression of oral squamous cell carcinoma by activating PI3K/Akt/Rb pathway.

10-formyltetrahydrofolate dehydrogenase (ALDH1L1) is a major folate enzyme, which is usually underexpressed in malignant tumors and competes with tumors for the same folate substrate. However, the specific role and mechanisms of ALDH1L1 in oral squamous cell carcinoma (OSCC) remainsobscure. The expression level of ALDH1L1 in paired OSCC tissues and adjacent noncancerous tissues were detected by quantitative realtime PCR, Western blot and immunohistochemistry. The relationship between ALDH1L1 expression and clinical characteristics was analyzed. Besides, CCK8, EdU staining, colony formation, wound healing, transwell invasion, apoptosis, cell cycle assays and nude mice tumor bearing experiments were employed to assess the role of ALDH1L1 in OSCC. To explore the underlying mechanisms of these effects, cell cycle-related markers were examined. In this study, we revealed that ALDH1L1 expression was significantly reduced in OSCC, and its downregulation was associated with the malignancy of the tumor and poor prognosis of patients. In vivo and in vitro experiments, downregulation of ALDH1L1 in OSCC significantly inhibited the occurrence of NADP+ -dependent catalytic reactions and facilitated tumor cell growth, migration, invasion, survival, cell cycle progression, and xenograft tumor growth. On the contrary, re-expression of ALDH1L1 plays a similar role to anti-folate therapy, promoting NADPH production and suppressing the progression of OSCC. Furthermore, ALDH1L1 overexpressing obviously inhibited the expression of PI3K, p-Akt, CDK2, CDK6, Cyclin D1, Cyclin D3, and Rb in OSCC cells, and promoted the expression of p27. LY294002 and 740 Y-P were used to confirm the inhibitory effects of ALDH1L1 on OSCC progression through PI3K/Akt/Rb pathway. Our findings highlight the clinical value of ALDH1L1 as a prognostic marker and the potential of a new target for anti-folate therapy.

LncRNA HOXD-AS1 promotes oral squamous cell carcinoma by sponging miR-203a-5p.

In the present study, we aimed to explore lncRNA HOXD cluster antisense RNA 1 (HOXD-AS1) expression in oral squamous cell carcinoma (OSCC) tissues, its biological roles, and the underlying potential mechanisms in OSCC progression. HOXD-AS1 expression in paired OSCC and non-tumor tissues from 60 OSCC patients was determined by RT-qPCR. The effects of HOXD-AS1 and miR-203a-5p overexpression on expression of Annexin A4, a validated target of miR-203a-5p, were analyzed by RT-qPCR and Western blot. The roles of HOXD-AS1, miR-203a-5p, and Annexin A4 in the invasion and migration of OSCC cells were analyzed by Transwell assays. HOXD-AS1 overexpression in OSCC predicted poor survival. HOXD-AS1 was predicted to interact with miR-203a-5p, but its expression was not significantly correlated with miR-203a-5p. HOXD-AS1 overexpression increased Annexin A4 expression, while miR-203a-5p overexpression decreased Annexin A4 expression in OSCC cells. Transwell assays showed that invasion and migration of OSCC cells were enhanced by HOXD-AS1 and Annexin A4 overexpression but were reduced by miR-203a-5p overexpression. In addition, miR-203a-5p overexpression suppressed the role of HOXD-AS1 in cell movement and Annexin A4 expression. HOXD-AS1may upregulate Annexin A4 by sponging miR-203a-5p in OSCC to promote cancer cell invasion and migration.

Comprehensive analysis of circRNA-miRNA-mRNA in oral squamous cell carcinoma

ObjectivesThe aim of the present study was to explore expression profiles of circular RNAs (circRNAs) and potential molecular mechanisms in oral squamous cell carcinoma (OSCC). DesignIn this study, high-throughput microarray was used to detect the expression profiles of circRNAs in OSCC. Reverse transcription quantitative PCR (RT-qPCR) was used to quantify the expression of dysregulated circRNAs in microarray. The validated microRNAs sponged by circRNAs and targeted mRNA were collected from literatures and miRTarBase database. and Gene Ontologypathways were analyzed on DAVID platform. Furthermore, network of circRNA-miRNA-mRNA and protein-protein interaction were constructed by Cytoscape 6.0 software. ResultsThe results revealed that 1540 circRNAs were differentially expressed in 4 paired OSCC tissues and adjacent tissues significantly. Among them, 6 abundant circRNAs which have been reported in literatures were identified through RT-qPCR. According to the hypothesis of competing endogenous RNAs, 14 miRNAs sponged by the validated circRNAs and 62 downstream target protein coding genes were identified. Upon Gene Ontologyanalysis, 62 target genes were found to play roles in positively regulation of cell migration, cell division, angiogenesis and response to hypoxia. For Kyoto Encyclopedia of Genes and Genomes pathways, target genes were found to be involved in oncogenic signaling pathways (hsa05200), PI3K-Akt signaling pathway (hsa04151), focal adhesion (hsa04510), microRNAs in cancer (hsa05206), ECM-receptor interaction (hsa04512). These results suggested that circRNAs may participate in the tumorigenesis and development of OSCC via sponging miRNAs. ConclusionsThis study indicated that circRNA may play vital role in tumorigenesis and metastasis in OSCC.

Precise Identification of Recurrent Somatic Mutations in Oral Cancer Through Whole-Exome Sequencing Using Multiple Mutation Calling Pipelines

Understanding the genomic alterations in oral carcinogenesis remains crucial for the appropriate diagnosis and treatment of oral squamous cell carcinoma (OSCC). To unveil the mutational spectrum, in this study, we conducted whole-exome sequencing (WES), using six mutation calling pipelines and multiple filtering criteria applied to 50 paired OSCC samples. The tumor mutation burden extracted from the data set of somatic variations was significantly associated with age, tumor staging, and survival. Several genes (MUC16, MUC19, KMT2D, TTN, HERC2) with a high frequency of false positive mutations were identified. Moreover, known (TP53, FAT1, EPHA2, NOTCH1, CASP8, and PIK3CA) and novel (HYDIN, ALPK3, ASXL1, USP9X, SKOR2, CPLANE1, STARD9, and NSD2) genes have been found to be significantly and frequently mutated in OSCC. Further analysis of gene alteration status with clinical parameters revealed that canonical pathways, including clathrin-mediated endocytotic signaling, NFκB signaling, PEDF signaling, and calcium signaling were associated with OSCC prognosis. Defining a catalog of targetable genomic alterations showed that 58% of the tumors carried at least one aberrant event that may potentially be targeted by approved therapeutic agents. We found molecular OSCC subgroups which were correlated with etiology and prognosis while defining the landscape of major altered events in the coding regions of OSCC genomes. These findings provide information that will be helpful in the design of clinical trials on targeted therapies and in the stratification of patients with OSCC according to therapeutic efficacy.

CircLPAR3 Acts as an Oncogene in Oral Squamous Cell Carcinoma Through Regulating the miR-643/HMGB2 Network.

The malignant progression of oral squamous cell carcinoma (OSCC) has been confirmed to be mediated by a variety of factors, including circular RNA (circRNA). However, the role of circLPAR3 in OSCC development is still unclear. 70 paired OSCC tissues and normal control tissues were obtained from 70 OSCC patients. Quantitative real-time PCR was used to detect the expression of circLPAR3, microRNA (miR)-643, and high-mobility group box 2 (HMGB2). Cell proliferation, apoptosis, metastasis and stemness were assessed using cell counting kit 8 assay, colony-formation assay, flow cytometry, transwell assay and sphere formation assay. Marker protein expression and HMGB2 protein expression were determined by western blot analysis. The interaction between miR-643 and circLPAR3 or HMGB2 was confirmed by RNA pull-down assay, dual-luciferase reporter and RIP assay. The role of circLPAR3 in OSCC tumorigenesis was explored by constructing the xenograft models. Our data showed that circLPAR3 was highly expressed in OSCC tissues and cells. CircLPAR3 silencing suppressed OSCC cell proliferation, metastasis and stemness, while promoted apoptosis. On the mechanism, we discovered that circLPAR3 could sponge miR-643 to positive regulate HMGB2. MiR-643 overexpression had an inhibition effect on OSCC progression, and its inhibitor could reverse the negative regulation of circLPAR3 knockdown on OSCC progression. In addition, overexpressed HMGB2 also reversed the suppressive effect of circLPAR3 silencing on OSCC progression. Animal experiments results showed that downregulated circLPAR3 repressed OSCC tumorigenesis in vivo. Taken together, our data showed that circLPAR3 contributed to OSCC malignant progression through regulating the miR-643/HMGB2 axis.

A Five-mRNA Expression Signature to Predict Survival in Oral Squamous Cell Carcinoma by Integrated Bioinformatic Analyses.

Objectives: This study was designed to identify a messenger RNA (mRNA) expression signature to predict survival in patients with oral squamous cell carcinoma (OSCC).Methods: mRNA expression profiles were integrated with clinical data from 280 samples, including 19 normal tissues and 261 OSCC tissues in The Cancer Genome Atlas. We identified differentially expressed mRNAs (DEmRNAs) between the OSCC and normal tissue samples and developed a novel mRNA-focused expression signature using a Cox regression analysis and other bioinformatic methods. The prognostic value of this signature was evaluated by Kaplan–Meier analysis, multivariable COX regression, and receiver operating characteristic (ROC) curve analysis. Protein–protein interaction (PPI) network, gene ontology, and Kyoto Encyclopedia of Genes and Genomes enrichment analysis were performed to predict the function of the DEmRNAs. Signature-related mRNAs were analyzed by gene set enrichment analyses (GSEA) and validated by quantitative real-time polymerase chain reaction (qRT-PCR) in 20 paired OSCC and adjacent healthy tissues.Results: We identified a novel 5-mRNA expression signature (HOXA1, CELSR3, HIST1H3J, ZFP42, and ASCL4) that could predict patient outcomes in OSCC. The risk score based on the signature was able to separate OSCC patients into high- and low-risk groups that showed significantly different overall survival (p < 0.001, log-rank test). The signature was further validated as an effective independent prognostic predictor of OSCC by multivariate Cox regression analysis (hazard ratio = 3.747, confidence interval: 2.279–5.677, p < 0.001) and ROC curve of the third year (area under the curve = 0.733). Functional analysis demonstrated that the key hub genes in the PPI network were mainly enriched in cell division, cell proliferation, and the p53 signaling pathway. GSEA results showed that the 5 mRNAs were significantly enriched in mismatch repair, DNA replication, and the NOTCH signaling pathway. Finally, qRT-PCR results showed that the 5 mRNAs were upregulated in OSCC tissue in agreement with the predictions from our bioinformatics analysis.Conclusions: We identified a novel 5-mRNA signature that could predict the survival of patients with OSCC and may be a promising biomarker for personalized cancer treatments.

Hsa_circ_0003829 serves as a potential diagnostic predictor for oral squamous cell carcinoma.

ObjectiveIncreasing evidence suggests that circular RNAs (circRNAs) play a major role in tumorigenesis and cancer progression. This study aimed to identify aberrant expression of hsa_circ_0003829 in oral squamous cell carcinoma (OSCC) and to explore its clinical significance.MethodsWe conducted a prospective clinical study to examine the expression pattern of hsa_circ_0003829 in 60 paired OSCC and normal clinical samples and in cell lines using real-time quantitative polymerase chain reaction. We also evaluated the diagnostic value of hsa_circ_0003829 in OSCC based on receiver operating characteristic (ROC) curve analysis, and examined the relationships between hsa_circ_0003829 expression and clinicopathological features in patients with OSCC. We further used bioinformatics software CircInteractome (https: //Circinteractome.nia.nih.gov/) to predict circRNA–microRNA interactions.ResultsHsa_circ_0003829 was significantly downregulated in OSCC compared with adjacent normal tissues. The area under the ROC curve was 0.81. Low expression levels of hsa_circ_0003829 in OSCC tissues were negatively correlated with lymph node metastasis status and TNM stage.ConclusionsDownregulated expression of has_circ_0003829 suggests that this may be a key circRNA in OSCC, and may serve as a prospective biomarker for the diagnosis of OSCC.

LncRNA LUADT1 Promotes Oral Squamous Cell Carcinoma Cell Proliferation by Regulating miR-34a/GAS1 Axis.

IntroductionThe oncogenic role of lncRNA LUADT1 has been investigated only in lung cancer. This study aimed to investigate the role of LUADT1 in oral squamous cell carcinoma (OSCC).Patients and MethodsThe expression levels of LUADT1 in paired OSCC and non-tumor tissues from OSCC patients were determined by RT-qPCR. A 5-year follow-up study was performed to analyze the prognostic value of LUADT1 for OSCC. Dual-luciferase assay and overexpression experiments were performed to assess the interactions among LUADT1, miR-34a and GASL1. Cell proliferation was analyzed by cell proliferation assay.ResultsIn this study, we found that LUADT1 was upregulated in OSCC and predicted poor survival. LUADT1 was predicted to interact with miR-34a, which was confirmed by dual-luciferase activity assay. However, overexpression experiments showed that they did not affect the expression of each other. Interestingly, overexpression of LUADT1 resulted in upregulation of GAS1, a target of miR-34a. Cell proliferation assay revealed that overexpression of LUADT1 and GAS1 resulted in promoted cell proliferation. MiR-34a played an opposite role and reversed the effects of LUADT1 overexpression.ConclusionLUADT1 may promote OSCC proliferation by regulating miR-34a/GAS1 axis.

KRT84 is a potential tumor suppressor and good prognosis signature of oral squamous cell carcinoma.

Aims: Oral squamous cell carcinoma (OSCC) is a common oral cancer; however, current therapeutic approaches still show limited efficacy. Our research aims to explore effective biomarkers related to OSCC.Main methods: Gene expression profiles of paired OSCC tumor and paracancerous samples from The Cancer Genome Atlas (TCGA) were analyzed. mRNA and protein levels of KRT84 in OSCC cell line HSC-3 were measured by real-time quantitative polymerase chain reaction (RT-qPCR) and Western blot. KRT84 protein levels in OSCC tumor samples of different stages were determined by immunohistochemistry. Overall survival (OS) of OSCC samples was evaluated and association of multiple factors with OS was assessed.Key findings: Compared with paracancerous samples, 4642 DEGs were identified in OSCC tumor samples. Among them, KRT84 expression level in OSCC tumor tissues was obviously decreased, which was validated in HSC-3 cells. KRT84 expression level showed decreasing tendency with the increase of tumor grade and stage. Patients with low KRT84 expression level had inferior OS independently of multiple factors. Besides, antigen processing and presentation pathway were significantly activated in OSCC samples with high KRT84 expression. Elevated KRT84 mRNA as well as protein levels were confirmed by RT-qPCR and Western blot in OSCC and normal cell lines, and immunohistochemistry in OSCC tumor and paracancerous tissues.Significance: Our study suggests KRT84 as a tumor suppressor and good prognostic indicator for OSCC, which might be significant for OSCC diagnosis and treatment.

Long noncoding RNA UCA1 promotes cell growth, migration, and invasion by targeting miR-143-3p in oral squamous cell carcinoma.

BackgroundThe long noncoding RNA (lncRNA) urothelial carcinoma‐associated 1 (UCA1) is dysregulated in many types of tumors; however, its role in oral squamous cell carcinoma (OSCC) remains unclear. This study aims to determine the effect of lncRNA UCA1 on OSCC.MethodsFifty‐six paired OSCC and adjacent nontumorous tissues were collected and the levels of UCA1, miR‐143‐3p, and MYO6 in the tissues were evaluated by qRT‐PCR. In in vitro experiments, cell viability, migration, and invasion were measured by, respectively, performing CCK‐8, wound healing, and transwell assays. The target relationships among UCA1, miR‐143‐3p, and MYO6 were verified by dual‐luciferase assay. Western blot and immunohistochemistry were carried out to determine the protein levels. Xenograft mouse model was established to explore the effects of UCA1 in vivo.ResultsLevels of UCA1 and MYO6 were increased significantly in OSCC, while the level of miR‐143‐3p was decreased compared with the adjacent nontumorous tissues. UCA1 promoted OSCC cell growth, migration, and invasion both in vitro and in vivo, while miR‐143‐3p reversed the progression. MYO6 was validated as a target for miR‐143‐3p, and MYO6 overexpression reversed the effects of miR‐143‐3p mimic on OSCC cells.ConclusionLncRNA UCA1 contributes to the proliferation and metastasis of OSCC cells by targeting miR‐143‐3p and upregulating its downstream gene MYO6. UCA1 could serve as a promising novel target therapy for treatment of OSCC.

Fibroblast Activation Protein (FAP) Overexpression Induces Epithelial-Mesenchymal Transition (EMT) in Oral Squamous Cell Carcinoma by Down-Regulating Dipeptidyl Peptidase 9 (DPP9).

PurposeFibroblast activation protein (FAP) acts as a tumor promoter via epithelial–mesenchymal transition (EMT) in human oral squamous cell carcinoma (OSCC). The present study was designed to investigate the FAP targeting proteins and explore the precise mechanism by which FAP promotes EMT in OSCC.Patients and MethodsProteins interacting with FAP were found and filtered by immunoprecipitation-mass spectrometry (IP-MS). Both DPP9 protein and mRNA were examined in 90 paired OSCC samples and matched normal tissue. DPP9 knockdown was conducted to determine its function in OSCC in vitro and in vivo.ResultsDipeptidyl peptidase 9 (DPP9) was identified as interacting with FAP intracellularly by IP-MS. The levels of both DPP9 protein and mRNA were down-regulated in OSCC tissue. Lower DPP9 expression was correlated with unfavorable survival rates of OSCC patients. DPP9 knockdown accelerates the proliferation of OSCC cells in vitro and in vivo. Overexpression of FAP leads to a reduction in DPP9 expression. Likewise, DPP9 overexpression reverses the proliferation, migration, invasion and EMT induced by FAP during OSCC.ConclusionOur study finds that FAP promotes EMT of OSCC by down-regulating DPP9 in a non-enzymatic manner. FAP-DPP9 pathway could be a potential therapeutic target of OSCC.

Hsa_circ_0086414 Might Be a Diagnostic Biomarker of Oral Squamous Cell Carcinoma.

BackgroundCircular RNAs (circRNAs), a newly-discovered class of non-coding RNAs, have a significant role in the progression of cancers, but the effect of hsa_circ_0086414 in human oral squamous cell carcinoma (OSCC) is still unclear.Material/MethodsThe circRNAs expression profile in OSCC tissue samples was assessed by high-throughput sequencing. The hsa_circ_0086414 expression level in 55 paired OSCC tissue samples and 2 kinds of OSCC cells was evaluated by quantitative real-time polymerase chain reaction (qRT-PCR). Additionally, the correlation between the hsa_circ_0086414 expression and clinicopathological characteristics of individuals with OSCC was studied. We used receiver operating characteristic (ROC) curves to observe the hsa_circ_0086414 value of diagnosis in OSCC. The network of hsa_circ_0086414-miRNAs-mRNAs was constructed. Gene Ontology (GO), Disease Oncology (DO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were carried out based on sequencing data and bioinformatics predictions.ResultsHsa_circ_0086414 expression in OSCC tissue samples and OSCC cells was first discovered to be significantly downregulated compared with the adjacent healthy tissues (AHTs) and normal (HaCaT) cells, respectively. Moreover, its expression level was significantly correlated with stage in TNM, size of tumor, and lymph node metastasis. The area below the ROC curve was 0.749. Hsa_circ_0086414-miRNAs-mRNAs network analysis and GO, DO, and KEGG analyses all demonstrated that hsa_circ_0086414 is correlated with cancer progression to a certain extent.ConclusionsWe discovered that hsa_circ_0086414 might be an essential diagnostic biomarker in OSCC. Furthermore, hsa_circ_0086414 could be a target for OSCC therapy.

CCL18 overexpression predicts a worse prognosis in oral squamous cell carcinoma (OSCC).

Oral squamous cell carcinoma (OSCC) presents severe morbidity and high mortality owing to local recurrence or remote metastasis. Molecular markers, including chemokines, might provide more efficient prognostic information or even therapeutic targets for the treatment of OSCC. Using quantitative RT-qPCR, we found that CCL18 was dramatically overexpressed in 30 OSCC tissues at the mRNA level in comparison with their adjacent non-cancerous oral mucosa tissues and 15 oral mucosa tissues from non-malignant patients. We then analyzed the relationship between CCL18 overexpression and patient clinical characters and outcomes using immunohistochemistry staining (IHC) in 102 paired OSCC cancerous and adjacent non-cancerous tissues; the increase in CCL18 expression was significantly higher in male patients (p=0.047), tumors of the palate and floor of the mouth (p=0.014), patients with positive lymph node metastasis (p=0.007), and patients with poor tumor differentiation (p=0.029). The median overall survival time and time-to-recurrence were 80.6 and 61.4 months in patients with high CCL18 expression, respectively, as against 93.4 and 81.6 months in patients with comparatively lower CCL18 expression, respectively (p=0.033 and 0.012, respectively; log-rank test). Multivariate analyses indicated age, poor differentiation, and CCL18 levels to be independent prognostic factors for predicting both overall and disease-free survival time. Our study suggests that CCL18 is a novel candidate marker for the OSCC malignancy and prognosis, including lymph node metastasis, time-to-recurrence, and disease-free survival time.

ALDH3A1 acts as a prognostic biomarker and inhibits the epithelial mesenchymal transition of oral squamous cell carcinoma through IL-6/STAT3 signaling pathway.

Objectives: Aldehyde dehydrogenase 3A1 (ALDH3A1) is a member of the ALDH superfamily and its relationship with oral squamous cell carcinoma (OSCC) still unknown. In our subject, we aimed to reveal the expression pattern and clinical value of ALDH3A1 in OSCC and its biological function in OSCC cell lines.Materials and methods: The expression level of ALDH3A1 in paired OSCC tissues and adjacent noncancerous tissues were detected by quantitative real-time PCR, Western blot and immunohistochemistry. The relationship between ALDH3A1 expression and clinical characteristics was analyzed. Besides, cell-counting kit 8, colony formation, wound healing, transwell invasion, apoptosis and cell cycle assays were employed to assess the role of ALDH3A1 in OSCC cells. To explore the influence of ALDH3A1 on OSCC epithelial-to-mesenchymal transition (EMT), the expression of EMT markers (E-cadherin, vimentin, snail, MMP3) on OSCC cells were detected, and possible mechanisms were analyzed.Results: In OSCC tissues, ALDH3A1 was significantly decreased compared to the adjacent normal tissues. Lower ALDH3A1 expression in OSCC tissues was associated with a higher incidence of lymph node metastasis (LNM). Moreover, the overall survival of OSCC with low ALDH3A1 expression was significantly worse compared to that of OSCC with high ALDH3A1 expression. Restored expression of ALDH3A1 suppressed cell proliferation, migration and invasion in OSCC cells. Further experiments showed that ALDH3A1 might inhibit EMT in OSCC via a regulation of the IL-6/STAT3 signal pathway.Conclusion: These data indicate that ALDH3A1 may serve as a biomarker and may be developed into a novel treatment for OSCC.

Guanylate-binding protein 6 is a novel biomarker for tumorigenesis and prognosis in tongue squamous cell carcinoma.

Guanylate-binding protein 6 (GBP6) is a member of the guanylate-binding protein family, and its role in cancer has not yet been reported. We aimed to investigate the clinical significance of GBP6 in oral squamous cell carcinoma (OSCC). Next-generation sequencing was applied for analyzing differential gene expression profiling between corresponding tumor adjacent normal (CTAN) and tumor tissue from two paired OSCC patients. Real-time PCRs (RT-PCRs) were used to investigate the gene expression level of GBP6 of CTAN and tumor tissue samples from 14 TSCC patients. Immunohistochemistry was used to investigate the protein expression level of GBP6 in tumor tissues and paired CTAN tissues from 488 OSCC patients, including 183 buccal mucosa squamous cell carcinoma (BMSCC), 245 tongue squamous cell carcinoma (TSCC), and 60 lip squamous cell carcinoma (LSCC) patients. Compared with CTAN tissues of OSCC patients, GBP6 is identified as a downregulated gene using the NGS platform, which was confirmed in 14 OSCC patients by RT-PCR. Moreover, protein expression level of GBP6 in tumor tissues was lower than that in CTAN tissues and the low GBP6 expression was correlated with poor cell differentiation/lymph node metastasis in TSCC patients. In addition, TSCC patients with low expression levels of GBP6 had poor disease-specific survival rate. The low expression of GBP6 was associated with tumorigenesis and poor prognosis in OSCC patients, especially in TSCC patients. GBP6 may serve as a novel favorable diagnostic and prognostic biomarker in TSCC patients.

Long non-coding RNA CCAT1 is a prognostic biomarker for the progression of oral squamous cell carcinoma via miR-181a-mediated Wnt/β-catenin signaling pathway

ABSTRACT Oral squamous cell carcinoma (OSCC) ranks as the sixth most common carcinoma worldwide, and the third most common carcinoma in developing countries as well. Recently, the aberrant expression of lncRNA CCAT1 has been revealed to play an important role in the development of several cancers. However, its role in OSCC remains unknown. The expression levels of CCAT1 and miR-181a were determined in 15 paired primary OSCC tissues and their adjacent noncancerous tissues and cell lines with qPCR. shRNA against CCAT1 was employed to investigate the impact of CCAT1 on proliferation and metastasis. Then dual luciferase reporter and RIP assays were utilized to study the interaction between CCAT1 and miR-181a. Cells transfected with sh-CCAT1 or treated with miR-181a inhibitor were subjected to western blot to investigate the role of Wnt/β-catenin signaling in CCAT1-mediated proliferation and metastasis. Finally, the role of CCAT1 in OSCC was confirmed with tumor xenografts mice model. CCAT1 was upregulated in OSCC tissues and cell lines. Knockdown of CCAT1 inhibited the proliferation, migration and invasion of OSCC cells, while the cell apoptosis was enhanced. Luciferase and RIP assays revealed that miR-181a was a direct target of CCAT1. Inhibition of miR-181a partially reversed the efficacy of sh-CCAT1. Moreover, sh-CCAT1 inhibited OSCC tissues growth through inhibiting Wnt signaling in a miR-181a-dependent manner in vivo. lncRNA CCAT1 activated Wnt/β-catenin signaling via inhibiting miR-181a, resulting in the cell proliferation, migration and invasion of OSCC, suggesting that CCAT1 might serve as a potential target of OSCC treatment. Abbreviation: LncRNA: long non-coding RNA; OSCC: oral squamous cell carcinoma; 3’ UTR: 3’ untranslated region; ANOVA: one-way analysis of variance; CDK: cyclin-dependent kinase; ceRNA: competing endogenous RNA; FBS: fetal bovine serum; HGF: human gingival fibroblasts; MAPK: mitogen-activated protein kinase; miRNA: micro RNA; ncRNA: noncoding RNAs; PBS: phosphate-buffered saline; PI3K: phosphatidylinositol 3-kinase