BackgroundSquamous cell carcinomas (SCC) account for approximately 30% of non-small cell lung cancer. Investigation of the mechanism of invasion and metastasis of lung SCC will be of great help for the development of meaningful targeted therapeutics. This study is intended to understand whether the activation of Hedgehog (Hh) pathway is involved in lung SCC, and whether activated Hh signaling regulates metastasis through epithelial-mesenchymal transition (EMT) in lung SCC.MethodsTwo cohorts of patients with lung SCC were studied. Protein expression was examined by immunohistochemistry, Western blot, or immunofluorescence. Protein expression levels in tissue specimens were scored and correlations were analyzed. Vismodegib and a Gli inhibitor were used to inhibit Shh/Gli activity, and recombinant Shh proteins were used to stimulate the Hh pathway in lung SCC cell lines. Cell migration assay was performed in vitro.ResultsShh/Gli pathway components were aberrantly expressed in lung SCC tissue samples. Gli1 expression was reversely associated with the expression of EMT markers E-Cadherin and β-Catenin in lung SCC specimens. Inhibition of the Shh/Gli pathway suppressed migration and up-regulated E-Cadherin expression in lung SCC cells. Stimulation of the pathway increased migration and down-regulated E-Cadherin expression in lung SCC cells.ConclusionsOur results suggested that the Shh/Gli pathway may be critical for lung SCC recurrence, metastasis and resistance to chemotherapy. Inhibition of the Shh/Gli pathway activity/function is a potential therapeutic strategy for the treatment of lung SCC patients.

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BackgroundThis retrospective study investigated the association between the Charlson comorbidity index (CCI) score and the survival of patients with stage IIIB-IV (advanced, non-resectable) non-small cell lung cancer (NSCLC) who also did not have gene mutations in epidermal growth factor receptor (EGFR) or anaplastic lymphoma kinase (ALK).MethodsThe records of 165 patients (28–80 y, median 61 y) who met the above criteria and were admitted to Beijing Friendship Hospital Capital Medical University from 1 May 2010 to 1 October 2014were reviewed. Associations between baseline variables and the CCI score were assessed via univariate and multivariate logistic regression analysis. Overall survival was defined as the time from the first clinic visit to death from any cause, or to the end of follow-up. Survival curves were estimated via the Kaplan-Meier method and compared with the log-rank test.ResultsLogistic regression analyses indicated that smoking and performance status were independently associated with the CCI score. Smoking was associated with an increased risk of mortality (odds ratio (OR) 4.12 (95% confidence interval [CI] 1.92–8.84) compared to non-smokers), as was performance status 2 (ambulatory, capable of self-care, unable to perform any work activities; active for >50% of waking hours) (OR 2.22 (95% CI, 1.14–4.33) compared to performance status 1). Univariate Cox’s regression analyses showed that the hazard ratios were significantly associated with the CCI score (P = 0.009), smoking (P = 0.042), and male gender (P = 0.021).ConclusionThe CCI score is an important prognostic factor for the prediction of overall survival in patients with stage IIIB-IV NSCLC who are negative for EGFR and ALK gene mutations.

Objective: To isolate exosomes from the serum of colon cancer patients and identify RNAs in the small vesicles.Methods: ExoQuick-TC™ Exosome Precipitation Solution was used for exosome isolation and the shapes of exosomes were observed under a transmission electron microscope. Mass spectrometry was used to identify the classification of miRNAs encapsulated in exosomes, and the expression levels of miRNA-21,-133a, and -181b in exosomes were detected by RT-PCR.Results: Exosomes isolated from serum of colon cancer patients were circular-or oval-shaped and vary in size with a diameter of 40-100 nm. Mass spectrometry shows that the main RNAs in exosomes are small RNAs; the levels of these small RNAs in exosomes are significantly higher compared with fresh serum. There is still a tiny amount of small RNAs in exosome-free serum, but the amounts are significantly lower than that in exosomes. No more RNAs were detected in the repeated freezing and thawing serum, but there were still some RNAs detectable in the exosomes extracted from these serums. MiRNA-21, -133a and -181b can be detected in exosomes, and the level of miRNA-21 is associated with early diagnosis of colon cancer.Conclusion: This study proves that commercial kits for exosome separation are more convenient and time-saving and that mass spectrometry is capable of identifying the miRNAs in exosomes. Compared with direct extraction of miRNAs from the serum, the method of isolating exosomes from the serum firstly and then extracting miRNAs from the exosomes can enhance the stability and integrity of their inner miRNAs. Also, we demonstrate that the exosomal miRNA-21 expression is associated with the early diagnosis of colon cancer.

BackgroundEsophageal cancer is a common and aggressive malignant tumor. This study aimed to investigate the association between human papillomavirus (HPV) Types 16 and 18 and esophageal carcinoma (EC) in the world population by conducting a meta-analysis.Materials and MethodsComputerized bibliographic and manual searches were performed to identify all eligible literatures between 1982 and 2014. PUBMED (http://www.ncbi.nlm.nih.gov/pubmed/) and CNKI (http://www.cnki.net/) were the primary sources of case-control studies, and key words used include human papillomavirus, HPV, esophageal, esophagus, cancer, carcinoma, and tumor. All searches were performed by reviewing articles and abstracts cited in the published systematic reviews and case-control studies. Prospective studies that reported relative risk (RR) estimates with 95% CIs for the association between HPV and EC were included.ResultsThirty-three randomized studies were identified, and the main features of these trials were included in this systematic review. HPV infection rate in the EC group was 46.5%, while HPV infection rate in the control group was 26.2% (OR = 1.62; 95% CI, 1.33–1.98). In China, the merger OR value was 1.62 (95% CI: 1.26–2.07); while in the Asian region, the merger OR value was 1.63 (95% CI: 1.29–2.04). There were statistical differences in HPV testing due to different detection methods such as PCR, IHC and ISH. In the PCR detection group, the merger OR value was 1.61 (95% CI: 1.33–1.95).ConclusionsThese results indicate that HPV infection and the incidence of EC are closely associated.

Altered expression of claudin-3 (CLDN3), a key cytoskeletal structural protein of the tight junctions in the epithelium, is associated with the development and metastasis of various human cancers. CLDN3 expression has been shown to be significantly associated with the prognosis of lung squamous cell carcinoma (SqCC). This study investigated the role of CLDN3 in inhibiting lung SqCC cell migration and invasion as well as the underlying molecular mechanisms. The CLDN3 levels were assessed between 20 paired lung SqCC tissues and adjacent normal tissues using quantitative real-time polymerase chain reaction (qRT-PCR) and western blot. The ectopic CLDN3 overexpression or knockdown was generated by using a plasmid carrying CLDN3 cDNA or shRNA, respectively. CLDN3 expression was significantly reduced in lung SqCC tissues vs. the adjacent normal tissues. The ectopic CLDN3 overexpression markedly inhibited the migration, invasion, and epithelial-mesenchymal transition (EMT) of lung cancer H520 cells, whereas CLDN3 knockdown had an inverse effect on SK-MES-1 cells. However, cell viability and plate colony formation assays showed that both CLDN3 knockdown and overexpression did not affect SqCC cell proliferation. Both tissue and cell data revealed that CLDN3 expression was significantly associated with the expression of the EMT biomarkers E-cadherin and Vimentin. Furthermore, CLDN3-modulated EMT and expression of the EMT markers were through regulation of the Wnt/β-catenin signaling pathway. In conclusion, this study identified reduced CLDN3 expression in lung SqCC tissues, which was associated with the progression and metastasis of lung SqCC and was attributed to EMT by activation of the Wnt pathway. Thus, CLDN3 could be further evaluated as a novel biomarker for predicting the prognosis of lung SqCC and as a target for the treatment of lung SqCC in the future.

BackgroundSquamous cell carcinomas (SCC) account for approximately 30% of non-small cell lung cancer (NSCLC). Current staging methods do not adequately predict outcome for this disease. EMX2 is a homeo-domain containing transcription factor known to regulate a key developmental pathway. This study assessed the significance of EMX2 as a prognostic and predictive marker for resectable lung SCC.MethodsTwo independent cohorts of patients with lung SCC undergoing surgical resection were studied. EMX2 protein expression was examined by immunohistochemistry, Western blot, or immunofluorescence. EMX2 expression levels in tissue specimens were scored and correlated with patient outcomes. Chemo-sensitivity of lung SCC cell lines stably transfected with EMX2 shRNAs to cisplatin, carboplatin, and docetaxel was examined in vitro.ResultsEMX2 expression was down-regulated in lung SCC tissue samples compared to their matched adjacent normal tissues. Positive EMX2 expression was significantly associated with improved overall survival in stage I lung SCC patients, and in stage II/IIIA lung SCC patients receiving adjuvant chemotherapy. EMX2 expression was also associated with expression of EMT markers in both lung SCC cell lines and tissue samples. Knock-down of EMX2 expression in lung SCC cells promoted chemo-resistance and cell migration.ConclusionsEMX2 expression is down-regulated in lung SCC and its down-regulation is associated with chemo-resistance in lung SCC cells, possibly through regulation of Epithelial-to-Mesenchymal Transition (EMT). EMX2 may serve as a novel prognostic marker for stage I lung SCC patients and a prediction marker for stage II/IIIA lung SCC patients receiving adjuvant chemotherapy.

The deregulation of claudin-3 has been reported to correlate with the invasion and metastasis of various cancers, but little is known about its expression level and the prognostic value in squamous cell lung carcinoma (SqCC). The purpose of this study is to determine the expression levels and the prognostic value of claudin-3 in completely resected SqCC tissues, and the potential underlying mechanism. The protein expression of claudin-3, E-cadherin, β-catenin, and vimentin in the tumor tissues from 103 patients with surgically resected SqCC was examined using immunohistochemistry, western blots, as well as semi-quantitative estimation. The claudin-3 protein level was significantly associated with E-cadherin, β-catenin, and vimentin protein expression. A decreased claudin-3 protein level was significantly correlated with TNM stage, lymph node metastasis, and disease recurrence. Similarly, downregulation of E-cadherin was significantly correlated with lymph node metastasis and disease recurrence. Decreased β-catenin expression also had a significant correlation with disease recurrence. Univariate analyses indicated that the T stage, lymph node metastasis, the TNM stage, and the expression of claudin-3, β-catenin, and vimentin were significant predictors for overall survival (OS). Moreover, multivariate analyses demonstrated that the TNM stage and protein levels of claudin-3, β-catenin, and vimentin were independent predictors for OS of SqCC patients. Claudin-3 plays an important role in the epithelial-mesenchymal transition of SqCC and might be used as a potential prognostic factor for SqCC.