Alternatively spliced ANLN isoforms synergistically contribute to the progression of head and neck squamous cell carcinoma.

Head and neck squamous cell carcinoma (HNSCC) is a common cancer with high mortality. Anilin actin-binding protein (ANLN) has been reported to be associated with carcinogenesis in multiple tumors. However, the expression pattern and functional effects of ANLN in HNSCC remain to be unclear. Clinical data and online databases were used to analyze the expression of ANLN and its relationship with HNSCC patient survival. Expression of two major splice variants of ANLN was assessed in HNSCC tissues and cell lines. The functional effects and related mechanisms of ANLN isoforms were investigated in HNSCC in vitro and in vivo. Our study showed that patients with high expression of ANLN had a poor prognosis. The two primary isoforms of ANLN transcripts ANLN-201 and ANLN-210 were highly expressed in HNSCC tissues and cell lines. Knockout of ANLN restrained cell proliferation, migration, and invasion of SCC-9 cells. Mechanically, ANLN-201 could interact with c-Myc to keep its protein stability, thereby playing a oncogenic role in HNSCC. ANLN-210 could be transferred to macrophages via exosomes by binding to RNA-binding protein hnRNPC. Exosomal ANLN-210 promoted macrophage polarization via PTEN/PI3K/Akt signaling pathway, thus stimulating tumor growth of HNSCC. ANLN was an independent prognostic factor in patients with HNSCC. Alternatively spliced ANLN isoforms collaboratively promote HNSCC tumorigenesis in vitro and in vivo, which might provide the in-depth role and mechanism of ANLN in HNSCC development.

The LncRNA FGD5-AS1/miR-497-5p axis regulates septin 2 (SEPT2) to accelerate cancer progression and increase cisplatin-resistance in laryngeal squamous cell carcinoma.

Aberrant expression or mutation of the Septin gene family is closely associated with cancer progression, and septin 2 (SEPT2) exerts its tumor-promoting effects in multiple cancers, but its role in regulating laryngeal squamous cell carcinoma (LSCC) progression and drug resistance has not been investigated. Based on the published data, the present study identified that SEPT2 promoted cancer progression and increased cisplatin-resistance in LSCC, and a novel LncRNA FGD5-AS1/miR-497-5p axis was crucial for this process. Mechanistically, SEPT2 tended to be enriched in LSCC tissues and cells, and knock-down of SEPT2 inhibited cell proliferation, viability, migration, and tumorigenesis in LSCC cells in vitro and in vivo. Aside from that, SEPT2 overexpression increased cisplatin resistance in LSCC cells. Next, by conducting the dual-luciferase reporter gene system assay, we identified that the LncRNA FGD5-AS1/miR-497-5p axis regulated SEPT2 in LSCC. Specifically, LncRNA FGD5-AS1 sponged miR-497-5p to upregulate SEPT2 in LSCC cells in a competing endogenous RNA (ceRNA) mechanisms-dependent manner. Interestingly, upregulated LncRNA FGD5-AS1 and downregulated miR-497-5p were observed in LSCC tissues and cells, and LncRNA FGD5-AS1 ablation inhibited cancer progression. Also, LncRNA FGD5-AS1 overexpression increased cisplatin-resistance in LSCC by modulating the miR-497-5p/SEPT2 axis. Collectively, we conclude that targeting the LncRNA FGD5-AS1/miR-497-5p/SEPT2 signaling cascade may be an alternative strategy to treat LSCC in the clinic.

Prognostic value of platelet distribution width and mean platelet volume in patients with laryngeal cancer.

Aims: To investigate the prognostic relevance of platelet volume indices for survival in laryngeal cancer. Patients & methods: The study included 640 patients with laryngeal cancer. We analyzed the optimal cutoff values through receiver operating characteristic analysis, then analyzed the univariate factor and multivariate variables. Kaplan-Meier curves and log-rank tests were conducted to compare the overall survival (OS) and recurrence-free survival rates between the groups. Results: In multivariate analysis, elevated platelet distribution width (PDW) and PDW/platelet count ratio were significantly correlated with poor prognosis for OS; however, elevated mean platelet volume (MPV) and MPV/platelet count ratio suggested a notable correlation with favorable prognosis for OS. Meanwhile, elevated PDW and decreased MPV were significantly correlated with poor prognosis for recurrence-free survival. Conclusions: Our findings indicate that elevated PDW and decreased MPV could serve as independent biomarkers for worse survival in laryngeal cancer.

Gene expression profiling for the diagnosis of multiple primary malignant tumors.

BACKGROUND: The incidence of multiple primary malignant tumors (MPMTs) is rising due to the development of screening technologies, significant treatment advances and increased aging of the population. For patients with a prior cancer history, identifying the tumor origin of the second malignant lesion has important prognostic and therapeutic implications and still represents a difficult problem in clinical practice. METHODS: In this study, we evaluated the performance of a 90-gene expression assay and explored its potential diagnostic utility for MPMTs across a broad spectrum of tumor types. Thirty-five MPMT patients from Sir Run Run Shaw Hospital, College of Medicine, Zhejiang University and Fudan University Shanghai Cancer Center were enrolled; 73 MPMT specimens met all quality control criteria and were analyzed by the 90-gene expression assay. RESULTS: For each clinical specimen, the tumor type predicted by the 90-gene expression assay was compared with its pathological diagnosis, with an overall accuracy of 93.2% (68 of 73, 95% confidence interval 0.84-0.97). For histopathological subgroup analysis, the 90-gene expression assay achieved an overall accuracy of 95.0% (38 of 40; 95% CI 0.82-0.99) for well-moderately differentiated tumors and 92.0% (23 of 25; 95% CI 0.82-0.99) for poorly or undifferentiated tumors, with no statistically significant difference (p-value > 0.5). For squamous cell carcinoma specimens, the overall accuracy of gene expression assay also reached 87.5% (7 of 8; 95% CI 0.47-0.99) for identifying the tumor origins. CONCLUSIONS: The 90-gene expression assay provides flexibility and accuracy in identifying the tumor origin of MPMTs. Future incorporation of the 90-gene expression assay in pathological diagnosis will assist oncologists in applying precise treatments, leading to improved care and outcomes for MPMT patients.

Prognostic Significance of Lactate Dehydrogenase in Patients Undergoing Surgical Resection for Laryngeal Squamous Cell Carcinoma.

The aim is to estimate the prognostic value of lactate dehydrogenase (LDH) in patients undergoing surgical resection for laryngeal squamous cell carcinoma (LSCC). A total of 640 resected LSCC patients were included. Preoperative lactate dehydrogenase (LDH) was assessed. Kaplan-Meier survival analysis and Cox regression analysis were conducted for overall survival (OS) and recurrence-free survival (RFS). Kaplan-Meier analysis, univariate analysis and multivariate analysis demonstrated significant prognostic value for preoperative LDH. Although LDH was predictor of OS, it failed to be a predictor of RFS. The univariate HR and 95% CI of LDH were 0.484 and 0.357-0.658 (P < 0.0001). The multivariate analysis showed that LDH (HR = 0.518, 95% CI: 0.380-0.705, p < 0.0001) was related to OS. Elevated preoperative LDH >132 IU/L was significantly associated with better survival. Preoperative LDH might be an independent prognostic marker of OS in LSCC patients undergoing surgical resection.

Long noncoding RNA LINC-PINT regulates laryngeal carcinoma cell stemness and chemoresistance through miR-425-5p/PTCH1/SHH axis.

Functional, noncoding RNA of about 200 nucleotides in length are known as long noncoding RNA (lncRNA). Advances in -omics have revolutionized the information with respect to the coding and noncoding regions of the genome. Several studies have illustrated the role of lncRNA in cell growth and cancer. Profiling and bioinformatic studies of laryngeal cancer has identified LINC-PINT as one of the lncRNA. However, the functional aspects of the deregulation have not been studied in laryngeal tumors. In this study, LINC-PINT expression in normal and tumor tissues were studied. Using a bioinformatic approach, microRNA (miRNA) targets of LINC-PINT and gene targets of the miRNA were determined. The impact of LINC-PINT on cell proliferation and chemoresistance was determined. Further through a set of silencing and re-expression studies phenotype rescue was studied. LINC-PINT expression was downregulated in laryngeal tumors. LINC-PINT targeted miR-425-5p by three sites. miR-425-5p also targeted PTCH1 a protein of the Hedgehog pathway. Downregulation of LINC-PINT was associated with increased cancer stemness and chemoresistance to cisplatin. Our results indicate a probable role of LINC-PINT in the pathology of laryngeal tumors. LINC-PINT re-expression in laryngeal tumors may be explored for reversion of cancer cell stemness and also for rescue of drug resistance phenotype.

Long non-coding RNA AFAP1-AS1/miR-320a/RBPJ axis regulates laryngeal carcinoma cell stemness and chemoresistance.

AFAP1-AS1 is a long non-coding RNA that is associated with tumorigenesis and poor prognosis in a variety of cancers. We have been suggested that AFAP1-AS1 increases tumorigenesis in laryngeal carcinoma specifically by enhancing stemness and chemoresistance. We assessed AFAP1-AS1 expression in human laryngeal specimens, paired adjacent normal tissues and human HEp-2 cells. Indeed, we found not only that AFAP1-AS1 was up-regulated in laryngeal carcinoma specimens and cells, but also that stemness-associated genes were overexpressed. Silencing of AFAP1-AS1 promoted HEp-2 cell chemoresistance under cisplatin treatment. Expression of AFAP1-AS1 was increased in drug-resistant Hep-2 cells. We then probed the mechanism of AFAP1-AS1 activity and determined that miR-320a was a potential molecular target of AFAP1-AS1. Luciferase reporter and qRT-PCR assays of AFAP1-AS1 and miR-320a levels in human specimens and cell cultures indicated that AFAP1-AS1 negatively regulates miR-320a. To discover the molecular mechanism of miR-320a, we again used the DIANA Tools algorithm to predict its genetic target, RBPJ. After cloning the 3'-untranslated regions (3'-UTR) of RBPJ into a luciferase reporter, we determined that miR-320a did in fact reduce RBPJ mRNA and protein levels. Ultimately, we determined that AFAP1-AS1 increases RBPJ expression by negatively regulating miR-320a and RBPJ overexpression rescues stemness and chemoresistance inhibited by AFAP1-AS1 silencing. Taken together, these results suggest that AFAP1-AS1 can serve as a prognostic biomarker in laryngeal carcinoma and that miR-320a has the potential to improve standard therapeutic approaches to the disease, especially for cases in which cancer cell stemness and drug resistance present significant barriers to effective treatment.

miR-217 inhibits laryngeal cancer metastasis by repressing AEG-1 and PD-L1 expression.

High incidences of laryngeal cancer have been reported recently. Increasing our understanding of the molecular mechanisms underlying this malignancy could reveal more effective approaches to treating laryngeal cancer patients and so improve their prognoses. In this study, we explored the biological effects of miR-217 on laryngeal cancer. miR-217 potently inhibited multiple metastatic traits, including cell migration, invasion, proliferation, apoptosis, and EMT, as well as angiogensis. These effects were achieved through downregulation of the miR-217 target gene, AEG-1 and PD-L1. Clinical expression and animal model studies further confirmed our results. These findings provide new insight into the physiological effects of miR-217 in laryngeal cancer and its potential therapeutic use.

Effect of EphA7 Silencing on Proliferation, Invasion and Apoptosis in Human Laryngeal Cancer Cell Lines Hep-2 and AMC-HN-8.

AIMS: This study aimed to investigate the expression of EphA7 in human laryngeal squamous cell carcinoma (LSCC) tissues and disclose the potential roles and molecular mechanisms of EphA7 in LSCC. METHODS: In the present study, we examined EphA7 expression and its function and mechanism in LSCC. EphA7 expression levels were investigated by quantitative real-time PCR (qRT-PCR), western blotting, and immunohistochemistry in a panel of 35 LSCC patient cases. To investigate the potential mechanism of EphA7 in human laryngeal cancer, we employed EphA7 siRNA to knockdown EphA7 expression in LSCC cell line Hep-2 and AMC-HN-8. Subsequently, MTT, TUNEL, qRT-PCR, and western blotting were performed to disclose the roles of EphA7 on proliferation, invasion and migration, and apoptosis in LSCC cell line Hep-2 and AMC-HN-8. RESULTS: Depletion of EphA7 remarkably inhibited the proliferation and invasion of Hep-2 and AMC-HN-8 cells in comparison to control and EphA7 siRNA negative control (NC)-transfected cells. TUNEL staining assay demonstrated that, compared with the control group, the rate of apoptosis in the EphA7 siRNA group was significantly increased. In addition, knockdown of EphA7 in Hep-2 or AMC-HN-8 cells markedly decreased the expression of EphA7 and PTEN, which could contribute to apoptosis. However, the bpV(phen), a PTEN inhibitor, could attenuate anti-proliferation and pro-apoptotic effects of EphA7 siRNA in Hep-2 and AMC-HN-8 cells. CONCLUSION: Up-regulation of EphA7 was observed in human LSCC samples and down-regulation of EphA7 effectively suppressed laryngeal carcinoma cell growth and promoted its apoptosis. Thus, EphA7 has a critical role in modulating cell growth and apoptosis, which serves as a potential therapeutic target in human LSCC.

miR-219 inhibits the growth and metastasis of TSCC cells by targeting PRKCI.

Tongue squamous cells carcinoma (TSCC) is the most common type in oral cancers. Recently, accumulating evidence suggests that microRNAs (miRNAs) play critical roles in tumorigenesis. Here, we demonstrated that miR-219 was significantly downregulated in TSCC tissues and cell lines. miR-219 overexpression remarkably suppressed cell proliferation, colony formation, migration and invasion of TSCC cells. In addition, protein kinase CI (PRKCI) was identified as a target of miR-219, and overexpression of PRKCI could significantly attenuated the tumor suppressive effects of miR-219. Furthermore, PRKCI inversely correlates with miR-219 in TSCC tissues. Taken together, miR-219 inhibited growth and metastasis by targeting PRKCI and might be used as a potential target for the treatment of TSCC.

Krüppel-like factor 5: a novel biomarker for lymph node metastasis and recurrence in supraglottic squamous cell laryngeal carcinoma.

Krüppel-like factor 5 (KLF5), a zinc finger-containing transcription factor, is involved in important biological processes including cell transformation, proliferation, and carcinogenesis. However, its clinical significance has remained largely unknown in laryngeal cancer. Here, specimens from 144 patients with laryngeal tumors were investigated by immunohistochemical staining for KLF5, integrin-linked kinase (ILK), and E-cadherin expressions. A clinicopathological study revealed that the KLF5 expression level in tumor cells was significantly correlated with lymph node metastasis (P < 0.05) and local recurrence (P < 0.05). In addition, KLF5, ILK, and E-cadherin (epithelial-mesenchymal transition (EMT) biomarker) expressions were correlated with each other. These findings suggest that KLF5 may be an epithelial-mesenchymal transition-associated biomarker in human laryngeal carcinomas and play important roles in the progression of laryngeal carcinomas. KLF5 immunoreactivity is therefore considered a potential lymph node metastasis and recurrence factor in human laryngeal cancers. In addition, the KLF5-mediated pathway is a potential target for elimination of laryngeal cancer in the future.

Inhibition of 15-lipoxygenase (15-LOX) reverses hypoxia-induced down-regulation of potassium channels Kv1.5 and Kv2.1Inhibition of 15-lipoxygenase (15-LOX) reverses hypoxia-induced down-regulation of potassium channels Kv1.5 and Kv2.1.

The inhibition of voltage-gated potassium channels (Kv) plays an important role in the cerebral hypoxia-induced cell death. The activity of Kv can be inhibited by 15-hydroxyeicosatetrienoic acid (15-HETE). Therefore, as the key enzyme which catalyzed the formation of 15-HETE, 15-LOX may be involved in Kv inhibition induced by cerebral hypoxia. In our study, Wistar rats cerebral arterial smooth muscle cells (CASMCs) were placed under the condition of hypoxia and control, 15-LOX was proved involved in hypoxia-induced vasoconstriction. Furthermore, 15-LOX gene over expression under normoxic condition, as well as 15-LOX gene knockout or inhibition under hypoxic condition was performed to investigate the expression and activity of Kv1.5 and Kv2.1 in CASMCs. Results showed that both hypoxia and 15-LOX over expression could cause Kv1.5 and Kv2.1 suppression, but no suppression was observed under hypoxic condition when 15-LOX gene was knockout or inhibited, which made 15-LOX a new target for the treatment of cerebral hypoxia. In conclusion, AA/15-LOX/15-HETE induces vasoconstriction by down-regulating Kv channels, and Kv2.1/1.5 channels are the targets. Our study also suggests a therapeutic strategy to improve ischemic vascular occlusion by lowering 15-HETE level and preventing Kv channel down-regulation, which makes 15-LOX as a new target for the treatment of cerebral hypoxia.