Gangcai Zhu

MicroRNA-324-3p regulates nasopharyngeal carcinoma radioresistance by directly targeting WNT2B.

PURPOSE Radioresistance severely restricts the clinical treatment of nasopharyngeal carcinoma (NPC). Accumulating evidence demonstrates that aberrant expression of microRNAs (miRNAs) contributes to cancer progression and sensitivity to radiation. Therefore, we aimed to identify miRNAs associated with radioresistance in NPC. METHODS Aberrant miRNA-324-3p expression in NPC CNE-2 cells with radioresistance (CNE-2-Rs), compared to its parental cells, was screened by high-throughput sequencing technology and determined by quantitative reverse transcription-polymerase chain reaction analysis (qRT-PCR) analysis. Bioinformatic analysis was used to predict the downstream target genes of miRNA-324-3p. Then, functional and mechanical analyses of miRNA-324-3p in NPC radioresistance were performed by overexpression and down-regulation of miRNA-324-3p in CNE-2-Rs cells and its parental cells. Finally, the clinical significance of miRNA-324-3p and WNT2B was investigated in NPC tissues. RESULTS Our data reveal that the expression of miRNA-324-3p is significantly decreased in CNE-2-Rs cells compared to its parental cells, and WNT2B is predicted to be the downstream target of miRNA-324-3p. Both overexpression and down-regulation of miRNA-324-3p following irradiation result in radiosensitivity alterations and protein changes of WNT2B signalling pathway in CNE-2-Rs cells and its parental cells. Importantly, down-regulation of miRNA-324-3p and up-regulation of WNT2B are significantly correlated with advanced clinical stages of NPC and this inverse expression pattern is also observed in NPC tissues before and after irradiation. CONCLUSIONS The present study reveals that miRNA-324-3p contributes to the radioresistance of NPC by regulating the WNT2B signalling pathway. Both miRNA-324-3p and WNT2B are potential biomarkers for radioresistance in NPC, which may serve as valuable targets for reversing radioresistance in the management of NPC.

Metadherin regulates metastasis of squamous cell carcinoma of the head and neck via AKT signalling pathway-mediated epithelial-mesenchymal transition.

Our recent study suggested that metadherin (MTDH) is overexpressed in laryngeal squamous cell carcinoma. Here, we further investigated its role in promoting metastasis of squamous cell carcinoma of the head and neck (SCCHN). An immunohistochemistry analysis demonstrated that MTDH is elevated and positively correlated with metastasis in 189 primary SCCHN tissues. In vitro experiments demonstrated that MTDH overexpression enhanced the migratory and invasive ability of SCCHN cells. Moreover, MTDH induced epithelial-mesenchymal transition (EMT) by both regulating morphological changes and mediating the expression of the biomolecular makers E-cadherin and vimentin. In addition, MTDH mediated AKT activation, and all of the above effects were nearly completely blocked by the inhibition of AKT. Our results suggested that MTDH might promote the metastasis of SCCHN through AKT signalling pathway mediated-EMT.

Metadherin regulation of vascular endothelial growth factor expression is dependent upon the PI3K/Akt pathway in squamous cell carcinoma of the head and neck.

AbstractOur previous study indicated overexpression of metadherin (MTDH) is an adverse prognostic factor in squamous cell carcinoma of the head and neck (SCCHN) and promotes SCCHN cell proliferation and invasion. However, its mechanism remains unclear. Recent studies have indicated that MTDH is a cancer-metastasis-associated molecule that participates in the process of angiogenesis. Therefore, the study is aimed to investigate that whether vascular endothelial growth factor (VEGF), as one of the most potent proangiogenic cytokines, is regulated by MTDH and the role of the phosphatidylinositide 3-kinases/Protein Kinase B (PI3K/Akt) pathway in this process of regulation and the clinical significance of both MTDH and VEGF in SCCHN.Immunohistochemistry was used to assay the expression of MTDH and VEGF in a cohort of 189 SCCHN patients with intact follow-up information. The expression of MTDH was then upregulated or inhibited by lentivirus-mediated MTDH Complementary deoxyribonucleic acid or MTDH short hairpin ribonucleic acid (shRNA) to observe the resulting alterations in VEGF expression and the PI3K/Akt signaling pathway in SCCHN cell lines. In addition, the PI3K/Akt pathway was modulated to observe the resulting changes in the MTDH-mediated expression of VEGF.The immunohistochemistry data showed that MTDH expression is positively correlated with VEGF expression in SCCHN tissues. Moreover, the overexpression of MTDH in SCCHN Tu686 and 5-8F cells led to increases in the expression of VEGF, and this effect was accompanied by activation of the PI3K/Akt pathway. Conversely, shRNA-mediated knockdown of MTDH led to decreased VEGF expression. In addition, inhibition of the Akt signaling pathway reversed the upregulation of VEGF resulting from MTDH overexpression. Moreover, the survival analysis revealed that VEGF is an independent prognostic factor, and a combined survival analysis based on both MTDH and VEGF showed synergistic effects in the prognosis evaluation of SCCHN patients.The findings of the present study demonstrate that MTDH regulates the expression of VEGF via the PI3K/Akt signaling pathway, indicating the potential role of the MTDH-mediated activation of VEGF signaling pathway in SCCHN angiogenesis and metastasis.

Quantitative iTRAQ LC-MS/MS Proteomics Reveals Transcription Factor Crosstalk and Regulatory Networks in Hypopharyngeal Squamous Cell Carcinoma

To date, no effective therapeutic treatments have been developed for hypopharyngeal squamous cell carcinoma (HPSCC), a disease that has a five-year survival rate of approximately 31% because of its late diagnosis and aggressive nature. Despite recent improvements in diagnostic methods, there are no effective measures to prevent or detect HPSCC in an early stage. The goal of the current study was to identify molecular biomarkers and networks that can facilitate the speedy identification of HPSCC patients who could benefit from individualized treatment. Isobaric tags for relative and absolute quantification (iTRAQ) labeling was employed with two-dimensional liquid chromatography-tandem mass spectrometry to identify quantitatively the differentially expressed proteins among three types of HPSCC disease stages. The iTRAQ results were evaluated by literature searches and western blot analysis. For example, FUBP1, one of 412 proteins with significantly altered expression profiles, was confirmed to have elevated expression in fresh HPSCC tissues. Integrin-mediated cell matrix adhesion and actin filament-inducing cytoskeleton remodeling were the cellular events that were the most relevant to HPSCC tumorigenesis and the metastatic process. The construction of transcriptional regulation networks led to the identification of key transcriptional regulators of tumor development and lymph node metastasis of HPSCC, including Sp1, c-Myc and p53. Additionally, our study indicated that the interactions among Sp1, c-Myc and p53 may play vital roles in the carcinogenesis and metastasis of HPSCC.

Elevated expression of histone demethylase PHF8 associates with adverse prognosis in patients of laryngeal and hypopharyngeal squamous cell carcinoma

AIM Overexpression of histone demethylase PHF8 has been reported to function as an oncoprotein in many cancers; however, the implications of PHF8 involvement in laryngeal and hypopharyngeal squamous cell carcinoma (LHSCC) remain unclear. This study aims to explore the expression of PHF8 and its clinical significance in LHSCC. MATERIALS & METHODS Western blotting and immunohistochemistry were performed to evaluate PHF8 protein expression in fresh and archived LHSCC samples. Global expressions of H3K27 and H3K9 methylation were analyzed in a cell line with PHF8 siRNA treatment. RESULTS & CONCLUSION In our study, PHF8 was upregulated in fresh LHSCC tissues. Immunohistochemical staining revealed that the expression of PHF8 was positively associated with T classification, clinical stage, primary tumor position and tumor relapse. Survival analysis demonstrated that high PHF8 expression was significantly associated with shorter overall survival and disease-free survival. Moreover, PHF8 regulates the levels of H3K9me2 and H3K27me2 in LHSCC. Taken together, PHF8 might be a novel prognostic marker for this disease.

KDM5B overexpression predicts a poor prognosis in patients with squamous cell carcinoma of the head and neck

Purpose: Lysine demethylase (KDM) 5B, as a member of the histone lysine demethylase family, is overexpressed and functions abnormally in various human cancers. However, its expression in the squamous cell carcinoma of the head and neck (SCCHN) remains unclear. Methods: KDM5B expression was analyzed by immunohistochemistry and correlated with clinicopathological parameters in 103 archival SCCHN tissue samples and 24 adjacent noncancerous epithelial tissues. Results: We found that KDM5B expression was higher in SCCHN than that in adjacent noncancerous tissues. This was closely associated with lymph node metastasis and tumor recurrence. In addition, Kaplan-Meier analysis revealed that patients with high KDM5B expression had shorter disease-free and overall survival times than those with low KDM5B expression. Importantly, both univariate and multivariate analysis demonstrated that KDM5B level was an independent prognostic factor in SCCHN patients. Conclusions: These results indicate that KDM5B is a valuable biomarker that can be used to predict SCCHN patient outcome.

Hypoxia promotes migration/invasion and glycolysis in head and neck squamous cell carcinoma via an HIF-1α-MTDH loop

Hypoxia is a hallmark of progressive cancer. Hypoxic cancer cells trigger glycolysis in response to a decreased O2 supply to meet metabolic and bioenergetic demands. Meanwhile, these responses to hypoxia and alterations of the microenvironment promote cancer cell metastasis by increasing transcription of hypoxia-inducible factor (HIF)-regulated genes. However, the detailed mechanism by which hypoxia regulates cancer cell metastasis and glycolysis remains to be investigated. In the present study, we identified that metadherin (MTDH), a multifaceted oncogene, is involved in the regulation of head and neck squamous cell carcinoma (HNSCC) metastasis and invasion under hypoxic conditions. Furthermore, the study indicated that there is a positive feedback loop between HIF-1α and MTDH in HNSCC cells, and that hypoxia promotes HNSCC cell metastasis and epithelial-mesenchymal transition by mediating the HIF-1α-MTDH loop. These findings implicate HIF-1α-MTDH as a promising target for anticancer drugs in solid tumors, and help to explain the pro-tumorigenic and unfavorable effect of MTDH on HNSCC observed in our previous studies.

A Prognostic 5-lncRNA Expression Signature for Head and Neck Squamous Cell Carcinoma

Head and neck squamous cell carcinoma (HNSCC) is a common malignant cancer that accounts for 5–10% of all cancers. This study aimed to identify essential genes associated with the prognosis of HNSCC and construct a powerful prognostic model for the risk assessment of HNSCC. RNAseq expression profile data for the patients with HNSCC were obtained from the TCGA database (GEO). A total of 500 samples with full clinical following-up were randomly divided into a training set and a validation set. The training set was used to screen for differentially expressed lncRNAs. Single-factor survival analysis was performed to obtain lncRNAs that associated with prognosis. A robust likelihood-based survival model was constructed to identify the lncRNAs that are essential for the prognosis of HNSCC. A co-expression network between genes and lncRNAs was also constructed to identify lncRNAs co-expressed with genes to serve as the final signature lncRNAs for prognosis. Finally, the prognostic effect of the signature lncRNAs was tested by multi-factor survival analysis and a scoring model for the prognosis of HNSCC was constructed. Moreover, the results of the validation set and the relative expression levels of the signature lncRNAs in the tumour and the adjacent tissue were consistent with the results of the training set. The 5 lncRNAs were distributed among 3 expression modules. Further KEGG pathway enrichment analysis showed that these 3 co-expressed modules participate in different pathways, and many of these pathways are associated with the development and progression of disease. Therefore, we proposed that the 5 validated lncRNAs can be used to predict the prognosis of HNSCC patients and can be applied in postoperative treatment and follow-up.

Tumor-associated macrophages derived CCL18 promotes metastasis in squamous cell carcinoma of the head and neck

BackgroundAlternatively activated macrophages in tumor microenvironment is defined as M2 tumor-associated macrophages (M2 TAMs) that promote cancer progression. However, communicative mechanisms between M2 TAMs and cancer cells in squamous cell carcinoma of head and neck (SCCHN) remain largely unknown.MethodsQuantitative real-time PCR, western blotting, enzyme-linked immunosorbent assay and flow cytometry were applied to quantify mRNA and protein expression of genes related to M2 TAMs, epithelial–mesenchymal transition (EMT) and stemness. Wounding-healing and Transwell invasion assays were performed to detect the invasion and migration. Sphere formation assay was used to detect the stemness of SCCHN cells. RNA-sequencing and following bioinformatics analysis were used to determine the alterations of transcriptome.ResultsTHP-1 monocytes were successfully polarized into M2-like TAMs, which was manifested by increased mRNA and protein expression of CCL18, IL-10 and CD206. Conditioned medium from M2-like TAMs promoted the migration and invasion of SCCHN cells, which was accompanied by the occurrence of EMT and enhanced stemness. Importantly, CCL18 neutralizing antibody partially abrogated these effects that caused by conditional medium from M2-like TAMs. In addition, recombinant human CCL18 (rhCCL18) correspondingly promoted the malignant biological behaviors of SCCHN in vitro. Finally, RNA-sequencing analysis identified 331 up-regulated and 363 down-regulated genes stimulated by rhCCL18, which were statistically enriched in 10 cancer associated signaling pathways.ConclusionThese findings indicate that CCL18 derived from M2-like TAMs promotes metastasis via inducing EMT and cancer stemness in SCCHN in vitro.

an individual drug-therapy and genetic testing report of temporal bone verrucous carcinoma

Objective To investigate the pathology and pathogenesis of and treatment methods for temporal bone verrucous carcinoma. Materials and methods A single-patient report of verrucous carcinoma on the left external auditory canal is presented and analyzed along with all cases of temporal bone verrucous carcinoma that have been documented in the English literature. Results Most of the patients with verrucous carcinoma of the temporal bone have histories of surgery, trauma, or infection, and verrucous carcinomas are sensitive to antimicrotubule chemotherapeutic medicines. Adjuvant radiation therapy is not effective, but surgical treatment might be relatively more effective. Conclusion Temporal bone verrucous carcinoma has a poor prognosis; therefore, the preferred treatment is surgical resection facilitated with antimicrotubule chemotherapeutic treatment. Adjuvant radiation therapy is not a preferred treatment for temporal bone verrucous carcinoma.