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BACKGROUND: Non-small cell lung cancer (NSCLC) is one of the most lethal malignancies worldwide. Deepening understanding of the pathogenesis of NSCLC is quite important for its treatment. Circular (circ) RNA_0015278 has been found to be downregulated in NSCLC, but its role in NSCLC and the underlying regulatory mechanism is unknown. METHODS: Circ_0015278, microRNA (miR)-1278 and SOCS6 were analyzed with real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) or western blot. Cell Counting Kit-8 (CCK-8) and 5-ethynyl-2'-deoxyuridine (EdU) staining were used to evaluate cell proliferation. The colony forming capacity and invasion of NSCLC cells were assessed with colony formation and transwell assays, respectively. The interaction among circ_0015278, miR-1278, and SOCS6 was evaluated using luciferase, receptor interacting protein (RIP), and RNA-pull down assays. Cell apoptosis was analyzed using flow cytometry. A subcutaneous NSCLC xenograft mouse model was established for evaluating circ_0015278-mediated effects on the growth of NSCLC in vivo. RESULTS: Circ_0015278 was downregulated in NSCLC tissues and cells, and its reduced expression indicated poor prognosis. Overexpression of circ_0015278 restrained the proliferation, colony formation, invasion, and epithelial-mesenchymal transition (EMT) of NSCLC cells and induced NSCLC cell apoptosis. Moreover, overexpression of circ_0015278 inhibited the growth of NSCLC in vivo. Mechanically, circ_0015278 acted as an miR-1278 sponge to reduce its quantity, and miR-1278 targeted SOCS6 to inhibit its expression in NSCLC cells. Circ_0015278 promoted SOCS6 expression by sponging miR-1,278 in NSCLC cells. Overexpression of circ_0015278 attenuated the malignant phenotypes of NSCLC through sponging miR-1278 and consequently promoting SOCS6 expression. CONCLUSIONS: We demonstrated for the first time that circ_0015278 attenuated the progression of NSCLC via targeting the miR-1278/SOCS6 axis, which provides potential diagnostic biomarkers and therapeutic targets.

Considerable differences in molecular characteristics have been defined between non-smoker and smokers in patients with lung adenocarcinoma (LUAD), yet study of open chromatin patterns associated with LUAD progression caused by smoking is still lacking. Here, we constructed a novel network based on correlations between each ATAC-seq peak from TCGA data using our previously developed algorithm. Subsequently, principal component analysis was performed on LUAD samples with retained peaks filtered by the correlation network and pathway analysis was conducted for potential pathways identification. Results were verified in an independent dataset from primary LUAD samples. We identified a set of peaks that clearly differentiated long-term from short-term smokers in LUAD patients and also significantly associated with overall survival of these patients. We then investigated the gene set related to those peaks and found that the comprising genes are strongly associated with LUAD development, such as B3GNT3, ACTN4 and CLDN3. They are consistent with the important roles for the associated pathways in LUAD oncogenesis induced by smoking, including glycosphingolipid biosynthesis and tight junction pathways.In summary, our study may provide valuable insights on exploration of ATAC-seq peaks and on smoking-related LUAD carcinogenesis from a perspective of open chromatin changes.

Objective: assess the feasibility of intraoperative intrathoracic chemotherapy and debulking surgery for pulmonary adenocarcinoma patients with pleural dissemination and no lymph node metastasis. Methods: We retrospectively reviewed medical records of 23 pulmonary adenocarcinoma patients with pleural dissemination and no lymph node metastasis underwent debulking surgery. They were divided into intraoperative intrathoracic chemotherapy (IC) group comprising patients who not intraoperative intrathoracic chemotherapy (NIC) group. Result: There was no significant difference in the adverse reactions of chemotherapy between the two groups. The median progression-free survival (PFS) was 38.0 months (95% CI: 25.6-50.4) in IC group and 24.0months (95% CI: 6.3-41.7) in NIC group, respectively. There was statistical significance between two group (IC vs NIC, p=0.016).The median overall survival (OS) was42.0 months (95% CI: 37.4-46.6) in IC group and was 36.0months (95% CI: 28.4-43.6) in NIC group, respectively. But there were no statistical significance between two groups (p=0.082). Conclusions: Intraoperative intrathoracic chemotherapy might improve PFS for unexpected pulmonary adenocarcinoma with pleural dissemination and no lymph node metastasis in primary tumor resected patients and with less adverse reactions.

B cell leukemia-2 (Bcl-2) plays important roles in the development of tumor and drug resistance. The growth of tumor cells can be inhibited by downregulating the abnormal expression of Bcl-2 protein. TW37, an effective inhibitor of Bcl-2 protein, has now been widely studied in many tumors. In our study, it was found that TW37 exerted a significant effect on the proliferation, apoptosis and migration of Non-Small Cell Lung Cancer cells. Bcl-2 is also a key downstream factor of many signaling pathways such as Epidermal Growth Factor Receptor (EGFR). TW37 enhanced the inhibition of tumorigenesis by gefitinib, an EGFR-Tyrosine Kinase Inhibitor drug. Moreover, TW37 can promote apoptosis ability by inhibiting the phosphorylation level of EGFR protein in H1975 cells. Overall, TW37 enhances the pro-apoptosis and anti-migration ability of gefitinib in Non-Small Cell Lung Cancer.

Considerable differences in molecular characteristics have been defined between non-smoker and smokers in patients with lung adenocarcinoma (LUAD), yet study of open chromatin patterns associated with LUAD progression caused by smoking is still lacking. Here, we constructed a novel network based on correlations between each ATAC-seq peak from TCGA data using our previously developed algorithm. Subsequently, principal component analysis was performed on LUAD samples with retained peaks filtered by the correlation network and pathway analysis was conducted for potential pathways identification. Results were verified in an independent dataset from primary LUAD samples. We identified a set of peaks that clearly differentiated long-term from short-term smokers in LUAD patients and also significantly associated with overall survival of these patients. We then investigated the gene set related to those peaks and found that the comprising genes are strongly associated with LUAD development, such as B3GNT3, ACTN4 and CLDN3. They are consistent with the important roles for the associated pathways in LUAD oncogenesis induced by smoking, including glycosphingolipid biosynthesis and tight junction pathways. In summary, our study may provide valuable insights on exploration of ATAC-seq peaks and on smoking-related LUAD carcinogenesis from a perspective of open chromatin changes.

Exosomes, released from various cell types, serve as vehicles of intercellular communication. Rearranged anaplastic lymphoma kinase (ALK) has been detected in exosomes released from cancer cells in ALK-positive nonsmall cell lung cancer (NSCLC), however, the functional consequence of ALK in exosomes has not been studied. This study aims to address whether exosomal ALK release is affected by stress, and whether exosomal ALK can modulate survival of recipient cells in vitro and in vivo. Exosomes, isolated from ALK-containing H3122 cells with (Exo-Apo) or without (Exo-Ctrl) irradiation treatment, were transferred to recipient H3122 cells in vitro or mouse xenograft in vivo. Western blot, flow cytometry, MTT, and xenograft were employed to respectively assess activation of the ALK pathway, apoptosis, cell viability, and tumor growth. Exo-Apo contained much higher levels of phosphorylated ALK (p-ALK) than that of Exo-Ctrl, and it activated AKT, STAT3, and the ERK pathway in recipient H3122 cells. ALK-specific inhibitors, including Crizotinib, Ceritinib, and TAE684, exhibited less effects on H3122 cells preincubated with Exo-Apo than on those treated with Exo-Ctrl in either inhibition of cell viability or promotion of apoptosis. Moreover, in an H3122 xenograft model, the Exo-Apo treatment resulted in a greater tumor growth and less sensitivity to Ceritinib than the Exo-Ctrl treatment. The ALK protein cargo in exosomes could be a key element to drive tumor growth and compromise therapeutic efficacy of ALK inhibitors for ALK-positive NSCLC.

Lung cancer is a leading cause of cancer mortality worldwide. Promoter methylation of transcription factor 21 (TCF21) was frequently observed in the early stage of non-small cell lung cancer (NSCLC). However, clinical relevance and molecular functions of TCF21 in NSCLC progression remain unclear. In this study, we analyzed the associations between TCF21 expression and clinicopathological features in 100 patients with NSCLC and revealed the underlying molecular mechanisms of TCF21 methylation on cell viability, apoptosis and invasion of H1299 cells. We found that the expression of TCF21 was significantly regulated by its methylation level in patients with NSCLC and was associated with tumor stage, metastasis and invasion. Demethylation of H1299 cells by 5-aza-2'-deoxycytine (5-Aza) demonstrated that a higher level of TCF21 expression led to remarkable decreases of cell viability and invasion ability but an increase of cell apoptosis. Accordingly, TCF21 knockdown showed converse results to high expression of TCF21. TCF21 knockdown cells exhibited significantly upregulated ATG-9, BECLIN-1, and LC3-I/II expressions but decreased p62 expression compared to wildtype cells. Inhibition of autophagy by 3-methyladenine (3-MA) elevated TCF21 expression and increased cell apoptosis. TCF21 expression is clinically related to the progress of lung cancer and may inhibit autophagy by suppressing ATG-9 and BECLIN-1. In turn, autophagy may also play an important role in regulation TCF21 expression.

// Hao Wu 1, * , Jingcheng Zhou 1, * , Chao Zeng 1 , Da Wu 1 , Zhimin Mu 1 , Baokun Chen 1 , Yuancai Xie 1 , Yiwang Ye 1 , Jixian Liu 1 1 Department of Thoracic Surgery, Peking University Shenzhen Hospital, Shenzhen 518036, Guangdong Province, China * These authors have contributed equally to this work Correspondence to: Jixian Liu, email: jixianliudr@hotmail.com Hao Wu, email: haowu-dr@hotmail.com Keywords: curcumin, TCF21, DNMT1, exosome, lung cancer Received: August 31, 2016 Accepted: October 28, 2016 Published: November 22, 2016 ABSTRACT Curcumin is a novel drug for lung cancer treatment. However, the mechanism underlying the anti-tumor effect of curcumin remains elusive. Previous evidences indicated that, the methylating transferase DNMT1 is downregulated by curcumin, and the transcription factor 21 (TCF21) is suppressed by DNMT1. We hereby attempt to elucidate the correlation between curcumin treatment and TCF21 expression. Exosomes derived from curcumin-pretreated H1299 cells were used to treat BEAS-2B cells, which induced proliferation, colony formation and migration of BEAS-2B cells. An increase in TCF21 expression in response to curcumin was also seen, as revealed by real-time PCR (RT-PCR) and western blot. Analysis using the GEO database (access #GSE21210) indicated that a positive correlation existed between TCF21 levels and lung cancer patient survival. TCF21 overexpression and knockdown was introduced to H1299 cells through lentiviral system, which led to suppression and promotion of tumor growth, respectively. We also demonstrated that DNMT1 expression was downregulated by curcumin. Therefore, curcumin exerts its anti-cancer function by downregulating DNMT1, thereby upregulating TCF21.

Previous studies have suggested that the B-cell lymphoma 2 (Bcl-2) inhibitor, TW37, may induce apoptosis of the non-small cell lung cancer cell line, H1975/epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI), which exhibits secondary resistance to EGFR-TKI. However, the effects of TW37 on H1975/EGFR-TKI cells remain unclear. The aim of the present study was to investigate the effects of TW37 on the growth of H1975/EGFR-TKI cells and explore the underlying mechanisms. An in vitro study was performed, whereby H1975/EGFR-TKI cells were treated with serially increasing concentrations of TW37. MTT, flow cytometry, migration and transwell invasion assays were preformed to investigate the proliferation, apoptosis, migration and invasion of H1975/EGFR-TKI cells, respectively. In addition, reverse transcription-polymerase chain reaction and western blot analyses were performed to detect the mRNA and protein expression levels of apoptosis-associated factors, respectively. An enzyme-linked immunosorbent assay was performed to detect phosphatidylinositol [3,4,5] tris-phosphate (PIP3) expression. The results suggested that the mRNA and protein expression levels of Bcl-2 were significantly decreased in TW37-treated cells when compared with the untreated control group. Following treatment with TW37, the proliferation, migration and invasion ability of H1975/EGFR-TKI cells decreased in a dose-dependent manner, while the percentage of apoptotic cells increased. In addition, the results demonstrated that TW37 reduced the expression of PIP3 and the phosphorylation of AKT serine/threonine kinase 1 (AKT) in H1975/EGFR-TKI cells in a dose-dependent manner. In conclusion, TW37 inhibited H1975/EGFR-TKI cell growth and induced cell apoptosis potentially via suppression of AKT signaling pathway activation.

The molecular mechanisms involved in the progression of clear cell renal cell carcinomas (ccRCCs) are still unclear. The aim of this study was to analyse the relationships between expression of RALYL and clinical characteristics. In 41 paired samples of ccRCCs and adjacent normal tissues, we used real-time qPCR to evaluate the expression of RALYL mRNA. RALYL protein levels were determined in 146 samples of ccRCC and 37 adjacent normal tissues by immunohistochemistry. Statistical analysis was used to explore the relationships between expression of RALYL and the clinical characteristics (gender, age, tumor size, T stage, N stage, M stage, survival times and survival outcome) in ccRCC. In addition, these patients were follow-up period 64 months (range: 4~116 months) to investigate the influence on prognosis. We found significantly differences between ccRCC tissues and normal tissues (p

Pulmonary sequestration (PS) is an uncommon lung disease. Carcinoid tumorlets in pulmonary sequestration are extremely rare. This case report presents a rare clinical case of carcinoid tumorlet in pulmonary sequestration with bronchiectasis after breast cancer. A 64-year-old female was diagnosed with infiltrating ductal carcinoma of the left breast in February 2009. Chest computer tomography (CT) revealed a cystic low-density mass of ∼2.5×4.7 cm in the right lower lung field, as well as cystic bronchiectasis in the right lower lobe. A right lower lobectomy was performed. In the surgery, abnormal vessel growth from the mass was found. Therefore, intralobar PS was diagnosed and pathological examination supported the diagnosis. Subsequently, pathological examination identified a carcinoid tumorlet in the PS. This report presents a rare clinical case of PS and bronchiectasis as well as carcinoid tumorlet in PS following diagnosis of breast cancer three years earlier. When a mass is found in the lung of patents with bronchiectasis with a history of breast cancer, aggressive therapy should be considered, since the mass may be a tumor or precancerous lesion.

Transcription factor 21 (TCF21) has been identified as a candidate tumor suppressor at 6q23-q24 that is epigenetically inactivated in many types of human cancers. We recently found that TCF21 methylation level was significantly increased in clear cell renal cell carcinoma (ccRCC). The purpose of this study was to investigate the prognostic impact of TCF21 expression in ccRCC and analyze the relationship between TCF21 expression and methylation level. We used real-time PCR and immunohistochemical staining to detect the expression of TCF21, and used methylation specific-PCR (MS-PCR) to determine the methylation status of TCF21 in ccRCC samples and cell line 786-O. The results showed that TCF21 expression level in ccRCC samples was significantly lower than in normal adjacent tissue samples (NAT samples). The Kaplan-Meier survival analysis demonstrated that TCF21 was a significant prognosticator of cancer-specific survival (p=0.001). Furthermore, the DNA demethylating agent 5'-azacytidine restored part of TCF21 expression by suppressing TCF21 methylation in 786-O. The methylation level of TCF21 in ccRCC samples was much higher than in NAT samples. These results suggest that the expression of TCF21 was an independent prognostic factor for poor survival in patients with ccRCC. Aberrant methylation was an important reason for the down-regulation the expression of TCF21, and may be associated with tumorigenesis in ccRCC.

Background Due to recent leaps forward in DNA synthesis and sequencing technology, DNA manipulation has been extended to the level of whole-genome synthesis. Bacteriophages occupy a special niche in the micro-organic ecosystem and have potential as a tool for therapeutic agent. The purpose of this study was to carry out chemical synthesis of the bacteriophage G4 and the study of its infectivity. Methodology/Principal Findings Full-sized genomes of bacteriophage G4 molecules were completed from short overlapping synthetic oligonucleotides by direct assembly polymerase chain reaction and ligase chain reaction followed by fusion polymerase chain reaction with flanking primers. Three novel restriction endonuclease sites were introduced to distinguish the synthetic G4 from the wild type. G4 particles were recovered after electroporation into Escherichia coli and were efficient enough to infect another strain. The phage was validated by electron microscope. Specific polymerase chain reaction assay and restriction analyses of the plaques verified the accuracy of the chemical synthetic genomes. Conclusions Our results showed that the bacteriophage G4 obtained is synthetic rather than a wild type. Our study demonstrated that a phage can be synthesized and manipulated genetically according to the sequences, and can be efficient enough to infect the Escherichia coli, showing the potential use of synthetic biology in medical application.

Gastrointestinal stromal tumors (GISTs) are rare mesenchymal neoplasms of the gastrointestinal tract. Less than 1% occurs in the esophagus. Surgery is the primary treatment for patients with GISTs. We report a 29-year-old male was admitted after the detection of a posterior mediastinal mass during work-up with routine examination. He did not have any disease-related symptoms. The physical examination was unremarkable. Chest computed tomographic scan, the barium esophagogram and endoscopic esophageal ultrasound showed benign neoplasm. The patient was performed an enucleation surgery through the right posterolateral thoracotomy. The pathology revealed a 13.0 cm × 12.0 cm × 5.0 cm mass. The tumor was CD117 (C-kit), PDGFRA and DOG1 positive. These findings were consistent with a GIST of the esophagus. So the diagnosis of GIST of esophagus was confirmed. The pathological diagnosis of low grade of GIST of esophagus was confirmed. The patient has no evidence of recurrence and is in good clinical conditions up-to date, five years after surgery.

14501 Background: As is known the polycomb group protein BMI1 has been shown to support normal stem cell proliferation via its putative stem cell factor function. However, the potential molecular mechanism of BMI1 act as a cancer stem cell factor may also promote cancer development is unclear. In this study we demonstrated for the first time that Notch signal transduction pathway regulating expression of Bmi1 in colorecta caner cells and investigate the Notch1 mRNA and Bmi1 mRNA level in human colorectal cancer samples and their correlation with clinicopathological parameters of colorectal cancer. Methods and Results: We assessed Notch1 and Bmi1 mRNA in 50 cases of colorectal cancer and adjacent normal tissue by RT-PCR. The immunohistochemistry was used to determine the expression of ICN and in the 50 tissues of colorectal cancer and normal mucosa. Bmi1 SW480 cells were treated with DAPT (g-secretase inhibitor) at different times. MTT assay and flow cytometry were used to measure the proliferation and apoptosis of SW480 cells. The expression of ICN, Bmi1 were measured by western blot. Notch1 and Bmi1 mRNA level were increased in colorectal cancer compared with adjacent normal mucosa (P No significant financial relationships to disclose.