Search

Background: Postoperative survival of esophageal cancer patients has improved with advances in technology and treatment modalities. However, squamous cell carcinoma (SCC) often affects the esophagus together with the head and neck regions, with second primary head and neck cancer (SPHNC) adversely influencing the patients' quality of life. Therefore, patients with esophageal squamous cell carcinoma (ESCC) should be carefully followed up postoperatively. This study evaluated the risk of developing SPHNCs after an esophagectomy for ESCC., Methods: Patients with ESCC who underwent curative esophagectomy from January 2008 to December 2017 from two tertiary medical centers, Taipei Veteran General Hospital and Kaohsiung Veteran General Hospital, were retrospectively reviewed. SPHNC development was documented according to the anatomic region and the cumulative incidence rate and risk factors were analyzed., Results: The median follow-up period of the 435 patients included in our study was 48.5 months [interquartile range (IQR) 16.7-92.4 months]. Among the ESCC patients after curative esophagectomy, younger age <50 [hazard ratio 4.13, 95% CI: 2.53-6.75, p < 0.001) was the only independent factor for developing SPHNCs in the multivariable analysis. The cumulative incidence rate of SPHNCs of patients aged <50 years old was 14.3%, 22.1%, and 34.2% after 3, 5, and 10 years, respectively., Conclusions: There was a high risk of secondary cancer after ESCC in the upper aerodigestive tract, especially the head and neck regions; therefore, active surveillance is strongly recommended, especially in younger patients., (© 2024 International Society of Surgery/Société Internationale de Chirurgie (ISS/SIC).)

The 5‑year survival rate of patients with esophageal squamous cell carcinoma (ESCC) is <20%, highlighting the need for the development of novel therapeutic targets. Neuregulin‑1 (NRG1), a transmembrane protein involved in cell proliferation and survival signaling, has unclear biological functions and clinical value in ESCC. The present study investigated the association between NRG1 expression and ESCC by analyzing data from both patients with ESCC and The Cancer Genome Atlas database. Reverse transcription‑quantitative PCR and immunohistochemistry staining were used to determine the levels of gene and protein in the tissue. The findings revealed that NRG1 gene and protein levels were significantly higher in tumor tissues compared with the normal tissues. Elevated expression of NRG1 was associated with poor outcomes, particularly in patients with advanced ESCC. Silencing NRG1 decreased both its mRNA and protein levels, disrupting key signaling pathways, such as phosphorylated (p‑)AKT and cellular rapidly accelerated fibrosarcoma (p‑cRAF), which led to decreased cancer cell proliferation, migration and tumor sphere formation, along with increased cell death. High expression levels of NRG1 and cRAF were significantly associated with poor prognosis. Additionally, silencing NRG1 promoted autophagosome and autolysosome formation, decreasing LC3B levels. The use of the autophagy inhibitor chloroquine significantly enhanced cell death induced by NRG1 silencing, suggesting that autophagy functions as a survival mechanism in ESCC cells in which NRG1 is silenced. Furthermore, high co‑expression of NRG1 and LC3B was associated with a worse prognosis. On the whole, the present study demonstrated that targeting NRG1 with autophagy inhibitors may serve as a potential therapeutic strategy for ESCC.

Background: This retrospective study aimed to compare the efficacy and safety of one-stage computed tomography (OSCT)- to that of two-stage computed tomography (TSCT)-guided localization for the surgical removal of small lung nodules., Methods: We collected data from patients with ipsilateral pulmonary nodules who underwent localization before surgical removal at Veteran General Hospital Kaohsiung between October 2017 and January 2022. The patients were divided into the OSCT and TSCT groups., Results: We found that OSCT significantly reduced the localization time and risky time compared to TSCT, and the success rate of localization and incidence of pneumothorax were similar in both groups. However, the time spent under general anesthesia was longer in the OSCT group than in the TSCT group., Conclusions: The OSCT-guided approach to localize pulmonary nodules in hybrid operation room is a safe and effective technique for the surgical removal of small lung nodules., (© 2024. The Author(s).)

The primary reason for cancer-related fatalities is metastasis. The compound 4-carbomethoxyl-10-epigyrosanoldie E, derived from the Sinularia sandensis soft coral species grown in cultures, exhibits properties that counteract inflammation. Moreover, it has been observed to trigger both apoptosis and autophagy within cancerous cells. This research focuses on examining the inhibitory impact of 4-carbomethoxyl-10-epigyrosanoldie E on the migration and invasion processes in Cal-27 and Ca9-22 oral cancer cell lines. To assess how this compound affects cell migration and invasion, the Boyden chamber assay was employed. Furthermore, Western blot analysis was utilized to explore the underlying molecular mechanisms. In a dose-dependent manner, 4-carbomethoxyl-10-epigyrosanoldie E notably decreased the levels of matrix metalloproteinase-2 (MMP-2) and MMP-9, along with urokinase-type plasminogen activator (uPA), in both Cal-27 and Ca9-22 cell lines. Conversely, it elevated the concentrations of tissue inhibitors of metalloproteinases-1 (TIMP-1) and TIMP-2. In addition, the treatment with this compound led to the inhibition of phosphorylation in extracellular signal-regulated kinase (ERK), p38, and c-Jun N-terminal kinase (JNK). It also curtailed the expression of several key proteins including focal adhesion kinase (FAK), protein kinase C (PKC), growth factor receptor-bound protein 2 (GRB2), Rac, Ras, Rho A, mitogen-activated protein kinase kinase kinase 3 (MEKK3), and mitogen-activated protein kinase kinase 7 (MKK7). Furthermore, the expression levels of IQ-domain GTPase-activating protein 1 (IQGAP1) and zonula occludens-1 (ZO-1) were significantly reduced by the compound. The ability of 4-carbomethoxyl-10-epigyrosanoldie E to inhibit the migration and invasion of Cal-27 and Ca9-22 oral cancer cells was observed to be dose dependent. This inhibitory effect is primarily attributed to the suppression of MMP-2 and MMP-9 expression, as well as the downregulation of the mitogen-activated protein kinase (MAPK) signaling pathway., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2024 Rou-Yi Fang et al.)

Background: This study compares the surgical and long-term outcomes, including disease-free survival (DFS), overall survival (OS), and cancer-specific survival (CSS), between lobe-specific lymph node dissection (L-SND) and systematic lymph node dissection (SND) among patients with stage I non-small cell lung cancer (NSCLC)., Methods: In this retrospective study, 107 patients diagnosed with clinical stage I NSCLC undergoing video-assisted thoracic surgery lobectomy (exclusion of the right middle lobe) from January 2011 to December 2018 were enrolled. The patients were assigned to the L-SND ( n = 28) and SND ( n = 79) groups according to the procedure performed on them. Demographics, perioperative data, and surgical and long-term oncological outcomes were collected and compared between the L-SND and SND groups., Results: The mean follow-duration was 60.6 months. The demographic data and surgical outcomes and long-term oncological outcomes were not significantly different between the two groups. The 5-year OS of the L-SND and SND groups was 82% and 84%, respectively. The 5-year DFS of the L-SND and SND groups was 70% and 65%, respectively. The 5-year CSS of the L-SND and SND groups was 80% and 86%, respectively. All the surgical and long-term outcomes were not statistically different between the two groups., Conclusion: L-SND showed comparable surgical and oncologic outcomes with SND for clinical stage I NSCLC. L-SND could be a treatment choice for stage I NSCLC.

Esophageal cancer has a poor prognosis due to its aggressiveness and low survival rate. In Ease Asia, esophageal squamous cell carcinoma (ESCC) outnumbers esophageal adenocarcinoma (EAC). The ESCC patients still have high mortality despite modern surgical resection and neoadjuvant treatment. Determining patient and outcome prognostic factors is critical in ESCC treatment. In esophageal cancer, early growth response-1 (Egr-1) is a tumor suppressor gene, but the mechanism and associated genes are unknown. The study utilizes RNA interference method, the platform of Next Generation Sequencing (NGS) and bioinformatics analysis to investigate the influences after the Egr-1 gene slicing on the ESCC cells. The heat maps of differentially expressed mRNA and microRNAs were analyzed using the algorithm, Burrows-Wheller Aligner. The study showed that the expression of 51 mRNA and 26 microRNAs have significant changes in ESCC cells after Egr-1 knockdown. The KEGG enrichment analysis linked Egr-1-regulated genes and microRNAs. Egr-1 interactions with these genes and microRNAs may be important in tumor progression. In conclusions, this study provided the transcriptome patterns and relating pathway analysis for Egr-1 knockdown in ESCC cells. The mRNA and microRNAs altered by Egr-1 gene silencing might provide key information in the treatment of ESCC., (© 2022. The Author(s).)

Background: Esophageal squamous cell carcinoma (ESCC) is associated with poor survival despite surgical resection, and its pathogenesis has been broadly investigated in the past decade. Early growth response 1 (EGR-1) could involve regulating tumor development in ESCC cells. Methods: An attempt was made to examine the molecular and cellular influence of EGR-1 in esophageal cancer cells by RNA extraction, real-time PCR (qRT-PCR), cell culture, small interfering RNA (siRNA) knockdown, western blot, migration assay, and cell viability assay. One hundred and forty-four samples of ESCC were collected from our hospital and analyzed. Significantly higher EGR-1 expression was noted in tumor-adjacent normal tissue compared with tumor lesions. Results: The univariate analysis showed no significant impacts of EGR-1 expression on patients’ survival. However, after adjusting for the pathological stage, patients with EGR-1 expression > 68th percentile had lower risks of cancer-related death. Moreover, knockdown of EGR-1 significantly enhanced cell migration, invasion, and resistance to chemotherapeutic agents in two ESCC cell lines. Conclusions: EGR-1 plays a key role in tumor suppression involving tumor viability suppression and reflects the treatment effect of current chemotherapy for ESCC.

Background/aim: Chlorogenic acid (CGA) is a polyphenol compound found in a variety of foods, including coffee, tea, cherries, and apples. It has been found by a number of studies to affect the viability of human cancer cells. No study has investigated its effect on esophageal squamous cell carcinoma (ESCC) metastasis or the molecular mechanism underlying its effect on this disease., Materials and Methods: We first used the Taiwanese ESCC cell line CE81T/VGH to create CE81T-M4 cells. Treatment of higher motility cells with chlorogenic acid for 24 h led to inhibition of cell migration and invasion as shown by scratch migration and transwell assays., Results: Western blotting showed that chlorogenic acid halted the activation of EGFR/p-Akt/Snail pathway and suppressed the expression of MMP-2 and MMP-9. Knockdown of either EGFR or Akt inhibited Snail, MMP2, and MMP9 activity as well as cell migration and invasion., Conclusion: Chlorogenic acid inhibited cancer cell motility via the EGFR/p-Akt/Snail pathway and could potentially be used to develop an antimetastatic agent for ESCC in the future., (Copyright © 2022 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Background: Thymoma is a type of rare mediastinal tumor whose clinical characteristics and indicators of prognosis are poorly understood. This single-institution retrospective study aimed to assess the predictive value of tumor, node, metastasis (TNM) staging incorporating tumor size in predicting the risk of thymoma recurrence after resection., Methods: Four binary logistic regression models were developed. Models I and II included median tumor size and TNM stage, respectively. Model III included the above two variables. Model IV was model III containing these two variables and their interaction terms. All models were adjusted for WHO histological type, operational time, and adjuvant therapy., Results: A total of 276 patients with a median age of 51.0, including 21 patients with thymoma recurrence, were included in this study. Models II or III showed a lower -2LogL and higher AUC (0.735 and 0.738 vs. 0.576) with significantly better discrimination than model I, and model III and model II shared similar discrimination. In model III, TNM stage was positively correlated with thymoma recurrence. The recurrence risk of patients with TNM stage IV was significantly higher than those with TNM stage I (OR of 11.03, p = 0.022). No significant correlation between the tumor size and recurrence risk (p = 0.779) and no interaction was found between medium tumor size and TNM stage in model IV., Conclusions: This study suggests that the prediction contribution of the TNM stage combined with tumor size is similar to the TNM stage alone for tumor recurrence in patients with thymoma after surgical resection., (© 2021 The Authors. Thoracic Cancer published by China Lung Oncology Group and John Wiley & Sons Australia, Ltd.)

Background/aim: Urothelial carcinoma (UC) is the most common type of genitourinary cancer with high incidence and mortality rates in men. In this study, we used the BFTC-905 and T24 bladder cancer cell lines as in vitro models to investigate the pathways involved in flaccidoxide-induced apoptosis., Materials and Methods: We utilized MTT assays, colony assays, wound-healing assays and fluorescence with TUNEL to confirm the cytotoxicity of flaccidoxide in bladder cancer cell lines. Potential proliferative and apoptotic molecular mechanisms were evaluated by western blotting., Results: The expression of anti-apoptotic proteins Bcl-2 and phosphorylated Bad (p-Bad) was attenuated with an increasing flaccidoxide concentration, while the expression of proapoptotic proteins Bax, Bad, cleaved caspase-3, cleaved caspase-9 and cleaved PARP-1 was found increased. Additionally, phosphorylation of phosphoinositide 3-kinases (PI3K), protein kinase B (AKT) and mammalian target of rapamycin (mTOR) in the PI3K/AKT/mTOR pathway was reduced, leading to a reduction in the phosphorylation of downstream 70-kDa ribosomal protein S6 kinase 1 (p70S6K), S6 ribosomal protein (S6) and eukaryotic translation initiation factor 4B (eIF4B). However, eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1) protein phosphorylation was increased due to attenuation of the upstream phosphorylation of mTOR protein., Conclusion: Flaccidoxide-induced apoptosis in BFTC-905 and T24 cells is mediated by mitochondrial dysfunction and down-regulation the PI3K/AKT/mTOR/p70S6K signaling pathway., (Copyright © 2021 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Nobiletin (NOB) is a polymethoxylated flavonoid isolated from citrus fruit peel that has been shown to possess anti-tumor, antithrombotic, antifungal, anti-inflammatory and anti-atherosclerotic activities. The main purpose of this study was to explore the potential of using NOB to induce apoptosis in human bladder cancer cells and study the underlying mechanism. Using an MTT assay, agarose gel electrophoresis, a wound-healing assay, flow cytometry, and western blot analysis, this study investigated the signaling pathways involved in NOB-induced apoptosis in BFTC human bladder cancer cells. Our results showed that NOB at concentrations of 60, 80, and 100 μM inhibited cell growth by 42%, 62%, and 80%, respectively. Cells treated with 60 μM NOB demonstrated increased DNA fragmentation, and flow cytometry analysis confirmed that the treatment caused late apoptotic cell death. Western blot analysis showed that mitochondrial dysfunction occurred in NOB-treated BFTC cells, leading to cytochrome C release into cytosol, activation of pro-apoptotic proteins (caspase-3, caspase-9, Bad, and Bax), and inhibition of anti-apoptotic proteins (Mcl-1, Bcl-xl, and Bcl-2). NOB-induced apoptosis was also mediated by regulating endoplasmic reticulum stress via the PERK/elF2α/ATF4/CHOP pathway, and downregulating the PI3K/AKT/mTOR pathway. Our results suggested that the cytotoxic and apoptotic effects of NOB on bladder cancer cells are associated with endoplasmic reticulum stress and mitochondrial dysfunction.

Oral squamous cell carcinoma (OSCC) is one of the major leading causes of cancer-related death worldwide, with limited effective markers for diagnosis and therapy, which has caused a low overall survival rate in the past decades. Kinases play important roles in tumor development and malignancy in various types of cancer. However, little is known about the role of kinases in OSCC cells. In this study, an arrayed kinome small interfering RNA (siRNA) library was used to screen oral cancer cell lines and counter assayed with normal fibroblast cells to identify the genes required for cancer cell proliferation. We found that polo-like kinase 1 (PLK1) was one of the most potent genes required for OSCC cell proliferation. The knockdown of PLK1 with a siRNA or antisense oligonucleotide (ASO) consistently diminished cyclin-B1 (CCNB1) expression/phosphorylation and the G 2 -M phase transition. Similar effects were observed in cells treated with the PLK1 kinase inhibitor BI6727. Besides, The Cancer Genome Atlas (TCGA) analysis revealed that PLK1 was elevated in tumor tissues and associated with short survival in patients with OSCC. We also found that PLK1 expression was highly correlated with the expression of its downstream effector, CCNB1, in patients with OSCC. Coexpression of the two genes resulted in a poor prognosis of OSCC patients, particularly those in the advanced stages of OSCC. Taken together, our results suggest that PLK1 might be a diagnostic or therapeutic marker for OSCC., Competing Interests: The authors declare no conflict of interest.

Targeted therapy is currently limited for patients with hepatocellular carcinoma (HCC) due to the lack of suitable targets. Kinases play pivotal roles in many cellular biological processes, whereas dysregulation of kinases may lead to various diseases, particularly cancer. However, the role of kinases in HCC malignancy remains unclear. In this study, we employed a kinome small interfering RNA (siRNA) library, comprising 710 kinase-related genes, to screen whether any kinases were essential for cell proliferation in various HCC cell lines. Through a kinome siRNA library screening, we found that MAP3K7 was a crucial gene for HCC cell proliferation. Pharmacological or genetic ablation of MAP3K7 diminished the growth, migration, and invasion of HCC cells, including primary HCC cells. Stable knockdown of MAP3K7 attenuated tumor formation in a spheroid cell culture model and tumor xenograft mouse model. In addition, silencing MAP3K7 reduced the phosphorylation and expression of mammalian target of rapamycin (mTOR) in HCC cells. MAP3K7 expression was positively correlated with mTOR expression in tumors of patients with HCC. Higher co-expression of MAP3K7 and mTOR was significantly associated with poor prognosis of HCC. Taken together, our results revealed that the MAP3K7-mTOR axis might promote tumorigenesis and malignancy, which provides a potential marker or therapeutic target for HCC patients.

Rationale: Bronchoscopic cryotherapy has been considered as one of the optional interventions for unresectable malignant central airway obstruction (CAO). And it provides high safety and effectiveness in airway patency re-establishment. This report describes the interventional bronchoscopic cryotherapy for a patient with CAO caused by squamous cell carcinoma of the esophagus. We display a series of dramatic change of chest radiographs before and after the intervention., Patient Concerns: A 70-year-old man with squamous cell carcinoma of the middle third of the esophagus (initial staging, pT2N0M0; stage IIB; in January 2017) underwent Video-assisted esophagectomy and reconstruction with a gastric conduit via a substernal route. Following Chest computed tomography and positron emission tomography revealed disease progression with paratracheal metastases. Progressive dyspnea and chest pain lasted for a month, and he was admitted to the ER., Diagnoses: Blood gas analysis revealed type I respiratory failure (pH, 7.445; PaO2, 69.4 mmHg; PaCO2, 40.6 mmHg). Other laboratory data were grossly normal. Chest radiography revealed a total left lung collapse. Chest CT identified a tumor blocking the left mainstem bronchus with the consolidation of the left lung., Interventions: Dexamethasone and epinephrine inhalation were administered for initial symptom relief. Bronchoscopy performed 4 days after admission revealed a huge tumor completely occluding the left mainstem bronchus orifice. The occlusion was completely resolved following cryotherapy. Then, the first course of palliative chemotherapy with cisplatin plus fluorouracil, followed by the second course a month later, was administered., Outcomes: The latest chest radiograph showed a patent airway. The patient's condition remained stable for at least the following 2 months., Lessons: Malignant CAO is a rare but potentially life-threatening condition. Several acceptable bronchoscopy techniques exist for treatment. Cryotherapy has high safety and effectiveness in airway patency re-establishment.

Pleomorphic carcinoma (PC) of lung, a rare malignant lung tumor, is predominated in male smokers with over 65 years of age. The clinical presentations of PC are various including chest pain, cough, and dyspnea and so on. Asymptomatic patients have been also reported. In our case, a female non-smoker with middle age, who initially developed symptoms like empyema was diagnosed advanced PC. Poor progression occurred in this patient within one month from diagnosis to expiration. The lesson from this case is that malignancy of lung such as PC could not be excluded if a patient develops unmanageable empyema., Competing Interests: Conflicts of Interest: The authors have no conflicts of interest to declare.

Thymomas are enigmatic tumors for which surgical resection is the mainstay of treatment. However, there are still many debates about resected thymomas with pleural recurrence. Repetitive operations for thymomas involving pleural recurrence are still the treatment of choice. Herein, we present a case with a suitable performance status for re-operation with a new application for argon beam coagulation. Both our experience and the currently available evidence suggest that surgical resection could be considered for patients with advanced thymomas, even for patients with locally advanced or Masaoka-Koga stage IV thymomas. Multimodality or multimodal, treatments resulted in better oncological outcomes for these patients. In this case, we proved that the new application of argon beam coagulation for a thymoma patient with pleural recurrence is safe and feasible. Additional evidence should be collected, and patients should be followed to assess long-term benefits., Competing Interests: Conflicts of Interest: The authors have no conflicts of interest to declare.

Background: Tumor cells require proficient autophagy to meet high metabolic demands and resist chemotherapy, which suggests that reducing autophagic flux might be an attractive route for cancer therapy. However, this theory in clinical cancer research remains controversial due to the limited number of drugs that specifically inhibit autophagy-related (ATG) proteins. Methods: We screened FDA-approved drugs using a novel platform that integrates computational docking and simulations as well as biochemical and cellular reporter assays to identify potential drugs that inhibit autophagy-required cysteine proteases of the ATG4 family. The effects of ATG4 inhibitors on autophagy and tumor suppression were examined using cell culture and a tumor xenograft mouse model. Results: Tioconazole was found to inhibit activities of ATG4A and ATG4B with an IC 50 of 1.3 µM and 1.8 µM, respectively. Further studies based on docking and molecular dynamics (MD) simulations supported that tioconazole can stably occupy the active site of ATG4 in its open form and transiently interact with the allosteric regulation site in LC3, which explained the experimentally observed obstruction of substrate binding and reduced autophagic flux in cells in the presence of tioconazole. Moreover, tioconazole diminished tumor cell viability and sensitized cancer cells to autophagy-inducing conditions, including starvation and treatment with chemotherapeutic agents. Conclusion: Tioconazole inhibited ATG4 and autophagy to enhance chemotherapeutic drug-induced cytotoxicity in cancer cell culture and tumor xenografts. These results suggest that the antifungal drug tioconazole might be repositioned as an anticancer drug or chemosensitizer., Competing Interests: Competing Interests: The authors have declared that no competing interest exists.

Background and Objectives: Thymoma has a variable long-term oncological outcome after surgical resection. Survival and tumor recurrence were analyzed to determine the predisposing factors for tumor recurrence., Methods: A total of 235 patients who underwent surgery for thymoma or thymic carcinoma from December 1997 to March 2013 were analyzed using Masaoka staging system and World Health Organization (WHO) histological classification. Surgical intervention included extended thymothymectomy via median sternotomy and thymomectomy via thoracotomy/ video-assisted thoracoscopic surgery (VATS)., Results: The median duration of follow-up was 105 months (12-198 months). Among these 235 patients, recurrence was observed in 25 patients (10.7%). according to Masaoka stage I, IIA, IIB, III, IVA, IVB, recurrence rates were 1/65(1.5%), 8/106(7.5%), 1/32(3.1%), 6/20(30.0%), 8/10(80.0%), 1/1(100.0%), respectively. Disease or treatment-related mortality was observed in 13 patients. Overall survival rate was 94.4%. After univariate analysis, predisposing factors for tumor recurrence included Masaoka stage, WHO histologic type, tumor size, adjuvant therapy and margin status., Conclusions: Due to the indolent behavior of thymoma, tumor recurrence appears to be a better assessment of oncological outcome rather than survival. Factors associated with tumor recurrence include Masaoka stage, WHO histologic type, tumor size, adjuvant therapy and margin status.

Angiomatosis is a nonneoplastic proliferative vascular lesion, which occurs mostly in diverse soft tissues. We observed a rare case of mediastinal angiomatosis with intraspinal invasion that mimicked a dumbbell tumor in a 63-year-old man with a history of prostate adenocarcinoma. A roentgenogram of the chest showed that the patient had left pleural effusion and a left paraspinal mass, computed tomography disclosed a low-density fusiform lesion over the left paraspinal region, and magnetic resonance imaging confirmed a large posterior mediastinal tumor with T4 intraspinal invasion. The tumor was completely excised through a laminectomy of the T3-5 spine, followed by thoracoscopic removal of the mediastinal part. The definitive diagnosis was angiomatosis. Surgical removal of such a dumbbell-mimicked tumor is mandatory because it may progress to spinal cord compression., (Copyright © 2016 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.)

Lung cancer is the leading cause of cancer mortality worldwide. Although many biomarkers have been identified for lung cancer, their low specificity and sensitivity present an urgent need for the identification of more candidate biomarkers. In this study, we conducted MRM-based targeted analysis to evaluate the potential utility of a list of candidate proteins for lung cancer diagnosis. A total of 1249 transitions of 420 peptides representing 102 candidate proteins from our previous study and the literature were first screened by MRM analysis in pooled plasma samples, resulting in 78 proteins remaining in the list. Relative quantification of these 78 proteins was further performed in 60 individual plasma samples from lung adenocarcinoma patients in stages I-III and matched healthy control subjects. Ultimately, nine proteins were found to be able to distinguish patients from controls. Further combinations of five, three, and two candidate marker proteins improved the sensitivity to discriminate patients from controls and resulted in a merged AUC value of nearly 1.00 in stages I-III patients versus controls. Our results highlighted several possible markers for lung adenocarcinoma, and the proposed protein panels require further validation in a larger cohort to evaluate their potential use in clinical applications or development of therapeutics.

Background: Differential expression and secretion of alpha-actinin 4 (ACTN4) in the lung cancer cell lines CL1-0 and CL1-5 have been reported in previous proteomic studies. The aim of this study is to investigate the functional properties of the ACTN4 protein in non-small-cell lung cancer (NSCLC) cells and evaluate its clinical importance., Methods: We used RNA interference to knock down and overexpress ACTN4 protein to evaluate the effects of this intervention on cancer cell invasion and migration, as well as on microscopic cellular morphology. Furthermore, we examined by immunohistochemistry the expression of ACTN4 protein in tissue samples at different stages of lung cancer and compared the protein levels of ACTN4 in blood plasma samples from patients with histologically confirmed lung cancer and healthy controls., Results: CL1-5 cell motility was significantly suppressed by the knockdown of ACTN4 protein. The morphology of CL1-5 cells changed from a predominantly mesenchymal-like shape into a globular shape in response to ACTN4 protein knockdown. A quantitative immunohistochemical assessment of lung cancer tissues revealed that ACTN4 protein level was considerably higher in cancerous tissues than in the adjacent normal ones, and the area under the receiver operating characteristic curve was 0.736 (p < 0.001). According to an enzyme-linked immunosorbent assay, the plasma levels of ACTN4 protein were significantly different between cancer patients and healthy controls, and the areas under the receiver operating characteristic curves were 0.828 and 0.909, respectively, for two independent cohorts (p < 0.001)., Conclusions: We demonstrate that the knockdown of ACTN4 protein inhibited cell invasion and migration. These results suggest that ACTN4 is associated with lung cancer cell motility. Thus, the level of ACTN4 in cancerous tissue and plasma is related to the presence of lung cancer.

Background: Pin1 promotes oncogenesis by regulating multiple oncogenic signaling. In this study, we investigated the involvement of Pin1 in tumor progression and in the prognosis of human esophageal squamous cell carcinoma (ESCC)., Results: We observed that proliferation, clonogenicity and tumorigenesis of CE81T cells were inhibited by Pin1 knockdown. We next analyzed Pin1 expression in clinical ESCC specimens. When compared to the corresponding non-tumor part, Pin1 protein and mRNA levels in tumor part were higher in 84% and 62% patients, respectively. By immunohistochemistry, we identified that high Pin1 expression was associated with higher primary tumor stage (p = 0.035), higher overall cancer stage (p = 0.047) and poor overall survival (p < 0.001). Furthermore, the association between expression of Pin1 and levels of β-catenin and cyclin D in cell line and clinical specimens was evaluated. β-catenin and cyclin D1 were decreased in CE81T cells with Pin1 knockdown. Cyclin D1 level correlated with Pin1 expression in clinical ESCC specimens., Conclusions: Pin1 upregulation was associated with advanced stage and poor prognosis of ESCC. Pin1 knockdown inhibited aggressiveness of ESCC cells. β-catenin and cyclin D1 were positively regulated by Pin1. These results indicated that targeting Pin1 pathway could represent a potential modality for treating ESCC.

Tumor susceptibility gene 101 (TSG101), a mammalian homologue of yeast vps23, is involved in protein sorting, vesicular trafficking and maintenance of genomic integrity. Upregulation of the TSG101 gene was found in human thyroid papillary and breast tumors. Here, we define the proximal promoter of human TSG101 at -1 to -436 by reporter assay. Intact Sp1 and MAZ binding sequences within this region are essential, and mutation of both sites eliminates proximal promoter activity implying cooperation between these two cis-elements. Chromatin immunoprecipitation and DNA affinity precipitation assay confirmed in vivo Sp1 binding on the GGGGCGGGTT sequence. MAZ protein was essential for TSG101 promoter activity because its knockdown using siRNA decreased reporter activity. An upstream regulatory element (URE) at the -1280 to -1757 region was identified to confer the orientation-independent enhancement of the promoter activity in transformed COS-1, ARO and WRO cell lines but not in a normal thyroid FRTL cell line. The sequence of this URE region contains putative binding sites for thyroid transcription factor 2 (TTF-2) and thyroid hormone receptor (T3R), which might be relevant to differential regulation of TSG101 promoter activity in transformed and primary cells.

Background: Tumor size is an important prognostic factor in many cancers, but its role in esophageal cancer remained undetermined. The aim of this study is to investigate the impact of tumor length on survival for patients with resected esophageal squamous cell carcinoma., Methods: A total 582 esophageal squamous cell carcinoma patients underwent surgical resection as the primary treatment was enrolled into this retrospective review. The longitudinal tumor length was defined as a uniformly measurement from the surgeons in the operating room immediately after completion of the esophagectomy. The impact of tumor length on patient's overall survival was assessed and compared with the factors among the current tumor-nodes-metastasis (TNM) staging system published in 2009., Results: The overall 1-, 3-, and 5-year survival rates were 70.4%, 37.8%, and 30.0%, respectively, with a median for 22 months. The length adversely affected the overall survival, and the 5-year survival rate was 77.3%, 48.1%, 38.5%, and 23.3 % for tumor lengths of 1 cm, 2 cm, 3 cm, and more than 3 cm, respectively (p < 0.001). In multivariate survival analysis, tumor length (more or less than 3 cm) remained an independent prognostic factor (p = 0.020) as did the other current TNM factors. For subgroup analysis, the predictive value of tumor length was significant in patients with T1 or T2 disease (p < 0.001), T3 or T4 disease (p = 0.029), and patients with N0 disease (p < 0.001), but not for patients with N1, N2, or N3 disease., Conclusions: Tumor length, which represents longitudinal spreading of the cancerous cells, could impact the overall survival of patients with resected esophageal squamous cell carcinoma, especially among those with nodal-negative disease. It may provide additional prognostic information to the current TNM staging system., (Copyright © 2011 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.)

The present study was designed to exploit optimum lipophilicity and high water-exchange rate (k(ex)) on low molecular weight Gd(III) complexes to generate high bound relaxivity (r(1)(b)), upon binding to the lipophilic site of human serum albumin (HSA). Two new carbon backbone modified TTDA (3,6,10-tri(carboxymethyl)-3,6,10-triazadodecanedioic acid) derivatives, CB-TTDA and Bz-CB-TTDA, were synthesized. The complexes [Gd(CB-TTDA)(H(2)O)](2-) and [Gd(Bz-CB-TTDA)(H(2)O)](2-) both display high stability constant (log K(GdL) = 20.28 and 20.09, respectively). Furthermore, CB-TTDA (log K(Gd/Zn) = 4.22) and Bz-CB-TTDA (log K(Gd/Zn) = 4.12) exhibit superior selectivity of Gd(III) against Zn(II) than those of TTDA (log K(Gd/Zn) = 2.93), EPTPA-bz-NO(2) (log K(Gd/Zn) = 3.19), and DTPA (log K(Gd/Zn) = 3.76). However, the stability constant values of [Gd(CB-TTDA)(H(2)O)](2-) and [Gd(Bz-CB-TTDA)(H(2)O)](2-) are lower than that of MS-325. The parameters that affect proton relaxivity have been determined in a combined variable temperature (17)O NMR and NMRD study. The water exchange rates are comparable for the two complexes, 232 × 10(6) s(-1) for [Gd(CB-TTDA)(H(2)O)](2-) and 271 × 10(6) s(-1) for [Gd(Bz-CB-TTDA)(H(2)O)](2-). They are higher than those of [Gd(TTDA)(H(2)O)](2-) (146 × 10(6) s(-1)), [Gd(DTPA)(H(2)O)](2-) (4.1 × 10(6) s(-1)), and MS-325 (6.1 × 10(6) s(-1)). Elevated stability and water exchange rate indicate that the presence of cyclobutyl on the carbon backbone imparts rigidity and steric constraint to [Gd(CB-TTDA)(H(2)O)](2-)and [Gd(Bz-CB-TTDA)(H(2)O)](2-). In addition, the major objective for selecting the cyclobutyl is to tune the lipophilicity of [Gd(Bz-CB-TTDA)(H(2)O)](2-). The binding affinity of [Gd(Bz-CB-TTDA)(H(2)O)](2-) to HSA was evaluated by ultrafiltration study across a membrane with a 30 kDa MW cutoff, and the first three stepwise binding constants were determined by fitting the data to a stoichiometric model. The binding association constants (K(A)) for [Gd(CB-TTDA)(H(2)O)](2-) and [Gd(Bz-CB-TTDA)(H(2)O)](2-) are 1.1 × 10(2) and 1.5 × 10(3), respectively. Although the K(A) value for [Gd(Bz-CB-TTDA)(H(2)O)](2-) is lower than that of MS-325 (K(A) = 3.0 × 10(4)), the r(1)(b) value, r(1)(b) = 66.7 mM(-1) s(-1) for [Gd(Bz-CB-TTDA)(H(2)O)](2-), is significantly higher than that of MS-325 (r(1)(b) = 47.0 mM(-1) s(-1)). As measured by the Zn(II) transmetalation process, the kinetic stabilities of [Gd(CB-TTDA)(H(2)O)](2-), [Gd(Bz-CB-TTDA)(H(2)O)](2-), and [Gd(DTPA)(H(2)O)](2-) are similar and are significantly higher than that of [Gd(DTPA-BMA)(H(2)O)](2-). High thermodynamic and kinetic stability and optimized lipophilicity of [Gd(CB-TTDA)(H(2)O)](2-) make it a favorable blood pool contrast agent for MRI.

Purpose: Inhibitors of antiapoptosis protein (IAP) have been implicated in the resistance to cisplatin. Therefore, verifying which pathway is involved in cIAP2 upregulation may be helpful in finding a feasible pathway inhibitor to increase the chemotherapeutic efficacy in human papillomavirus (HPV)-infected lung cancer., Experimental Design: Specific inhibitors of different pathways were used to verify which pathway is involved in cIAP2 transcription. cIAP2 promoter fragments with various deletions and/or mutations were constructed by site-directed mutagenesis. cIAP2, epidermal growth factor receptor (EGFR), and phospho-AKT (p-AKT) expressions in 136 lung tumors were evaluated by immunohistochemistry., Results: Our data show that two NF-κB (-209 to -200 and -146 to -137) and one CREB (cyclic AMP-responsive element binding protein; -52 to -42) binding sites in cIAP2 promoter region were responsible for cIAP2 upregulated by E6 in TL-1 cells. Moreover, CREB was phosphorylated by EGFR/phosphatidylinositol 3-kinase (PI3K) pathway. To test the involvement of cIAP2 on cisplatin resistance, IC(50) was lowered to 8.6 μmol/L in TL-1 cells with cIAP2 short hairpin RNA (shRNA) transfection and compared with 39.7 μmol/L in TL-1 cells with nonspecific shRNA. Pretreatment with EGFR or PI3K inhibitor in TL-1 cells diminished the resistance to cisplatin. Among the tumor groups, cIAP2 expression correlated significantly with HPV16/18 E6, EGFR, and p-AKT. We followed up 46 of 136 patients who had tumor recurrence and/or metastasis and underwent chemotherapy. Tumors with cIAP2-positive immunostaining were associated with a poorer tumor response to chemotherapy compared with those with negative immunostaining., Conclusions: cIAP2 upregulated by E6 via EGFR/PI3K/AKT cascades may contribute to cisplatin resistance, revealing that the EGFR or PI3K inhibitor combined with cisplatin may improve the chemotherapeutic efficacy in HPV-infected lung cancer., (©2010 AACR.)

Background: Once diagnosed, esophageal cancer has a very low overall 5-year survival rate. This study investigates the mechanisms behind the invasiveness and severity of esophageal squamous cell carcinoma (ESCC)., Materials and Methods: Transwell invasion chamber was used to subdivide one Taiwanese ESCC cell line, CE81T/VGH, into sublines (CE81T-0, CE81T-1, CE81T-2, CE81T-3, and CE81T-4) in four rounds of assays; the most invasive were identified, and various factors related to their invasiveness measured., Results: CE81T-1, CE81T-2, CE81T-3 and CE81T-4 sublines were significantly more invasive than the parental cells (CE81T/VGH) and CE81T-0 subline. CE81T-1 and CE81T-4, the sublines we chose to study further, had significantly greater colony-forming ability (3.5- to 2.7-fold) and wound migrating activity (1.95- to 2.6-fold) than the parental cells in vitro (p<0.01). They also displayed greater tumorigenesis in immunodeficient BALB/c Foxlnn mice than the parental cells. We found an inverse correlation between expression of tissue inhibitor of metalloproteinase-2 and invasive ability, and a significant positive correlation between expressions of matrix metalloproteinase-1, vimentin, and p-Src (pY416) in these cell lines and their invasiveness (all p<0.05)., Conclusion: The subline model may be used to study the molecular and genetic mechanisms underlying the invasion and metastasis of ESCC.

In a subset of lung adenocarcinomas, the epidermal growth factor receptor (EGFR) is activated by kinase domain mutations and/or gene amplification, but the interaction between the two types of abnormalities is complex and unclear. For this study, we selected 99 consecutive never-smoking women of East Asian origin with lung adenocarcinomas that were characterized by histologic subtype. We analyzed EGFR mutations by PCR-capillary sequencing, EGFR copy number abnormalities by fluorescence and chromogenic in situ hybridization and quantitative PCR, and EGFR expression by immunohistochemistry with both specific antibodies against exon 19 deletion-mutated EGFR and total EGFR. We compared molecular and clinicopathologic features with disease-free survival. Lung adenocarcinomas with EGFR amplification had significantly more EGFR exon 19 deletion mutations than adenocarcinomas with disomy, and low and high polysomy (100% versus 54%, P = 0.009). EGFR amplification occurred invariably on the mutated and not the wild-type allele (median mutated/wild-type ratios 14.0 versus 0.33, P = 0.003), was associated with solid histology (P = 0.008), and advanced clinical stage (P = 0.009). EGFR amplification was focally distributed in lung cancer specimens, mostly in regions with solid histology. Patients with EGFR amplification had a significantly worse outcome in univariate analysis (median disease-free survival, 16 versus 31 months, P = 0.01) and when adjusted for stage (P = 0.027). Lung adenocarcinomas with EGFR amplification have a unique association with exon 19 deletion mutations and show distinct clinicopathologic features associated with a significantly worsened prognosis. In these cases, EGFR amplification is heterogeneously distributed, mostly in areas with a solid histology.

MicroRNAs (miRNAs) make up a novel class of gene regulators; they function as oncogenes or tumor suppressors by targeting tumor-suppressor genes or oncogenes. A recent study that analysed a large number of human cancer cell lines showed that miR-330 is a potential tumor-suppressor gene. However, the function and molecular mechanism of miR-330 in determining the aggressiveness of human prostate cancer has not been studied. Here, we show that miR-330 is significantly lower expressed in human prostate cancer cell lines than in nontumorigenic prostate epithelial cells. Bioinformatics analyses reveal a conserved target site for miR-330 in the 3'-untranslated region (UTR) of E2F1 at nucleotides 1018-1024. MiR-330 significantly suppressed the activity of a luciferase reporter containing the E2F1-3'-UTR in the cells. This activity could be abolished with the transfection of anti-miR-330 or mutated E2F1-3'-UTR. In addition, the expression level of miR-330 and E2F1 was inversely correlated in cell lines and prostate cancer specimens. After overexpressing of miR-330 in PC-3 cells, cell growth was suppressed by reducing E2F1-mediated Akt phosphorylation and thereby inducing apoptosis. Collectively, this is the first study to show that E2F1 is negatively regulated by miR-330 and also show that miR-330 induces apoptosis in prostate cancer cells through E2F1-mediated suppression of Akt phosphorylation.

LATS2 is a member of the LATS tumor suppressor family. It has been implicated in regulation of the cell cycle and apoptosis. Frequent loss of heterozygosity (LOH) of LATS2 has been reported in human esophageal cancer. But, the LATS2 gene expression and its regulatory mechanism in esophageal cancer remain unclear. The present study has shown that LATS2 protein expression was mediated by miR-373 at the post-transcriptional level and inversely correlated with miR-373 amounts in esophageal cancer cell lines. Furthermore, we demonstrated that the direct inhibition of LATS2 protein was mediated by miR-373 and manipulated the expression of miR-373 to affect esophageal cancer cells growth. Moreover, this correlation was supported by data collected ex vivo, in which esophageal cancer tissues from esophageal squamous cell carcinoma (ESCC) patients were analyzed. Finally, by miRNA microarray analysis, four miRNAs including miR-373 were over-expressed in ESCC samples. Our findings reveal that miR-373 would be a potential oncogene and it participates in the carcinogenesis of human esophageal cancer by suppressing LATS2 expression.

Genetic variants in alcohol dehydrogenase-1B (ADH1B) and aldehyde dehydrogenase-2 (ALDH2) genes modulate acetaldehyde removal upon alcohol ingestion. Although these genetic vulnerabilities have been linked to higher esophageal squamous cell carcinoma (ESCC) risks, it is unclear whether they also determine the time of malignancy presentation. The purpose of this investigation was to unravel genotoxic effects of the two alcohol-metabolizing genes with regard to alcohol and tobacco consumption on the age at ESCC diagnosis and tumor dissemination. ADH1B/ALDH2 genotyping was performed on lymphocyte DNA specimens taken from 406 consecutively registered incident patients with pathology-proven ESCC. To fully utilize individual genetic and survival information, survival analyses and gene-longevity applied approaches were introduced. Among heavy drinkers, the ADH1B Arg/Arg (55 years) and ALDH2 Glu/Lys genotypes (54 years) were found to confer a 15 and 16 years earlier carcinoma diagnosed age than His/His and Glu/Glu nondrinkers (both 70 years), respectively. For drinkers, 1-year age advancement was, separately, associated with a 0.977 and 0.953-fold stepwise reduced likelihood of being ADH1B Arg homozygote and ALDH2 Lys variant. Noticeably elevated hazard-ratio (HR) for drinkers of ADH1B slow-form genotype and ALDH2 inactive-form allele were identified in smokers (HR = 2.3-2.6), but no in nonsmokers. In smokers, appreciably higher cumulative cancer onset risks were correspondingly recognized from the age of 45 and 49 upward among any + Lys allele and Arg/Arg + Glu/Glu combined-ADH1B/ALDH2-genotype drinkers than nondrinkers. In conclusion, consumption of tobacco and alcohol, coupled with genetic susceptibilities associated with acetaldehyde elimination, as modulated by ADH1B and ALDH2 genotypes, determines a substantial magnitude of tumorigenetic effect on earlier age ESCC diagnosis., (2009 UICC.)

Objective: The main objective of this study was to further elucidate the effect of consuming various foods on the development of squamous cell carcinoma (SCC) in three different sections of the esophagus., Methods: A total of 343 patients with SCC of the esophagus and 755 cancer-free control subjects were recruited for this study from 1996 to 2005., Results: We found that intake of vegetables, raw onions/garlic, and fruits are significantly protective against esophageal SSC risk, whereas intake of hot foods can significantly increase its risk. There was a significant inverse relation between the frequency of tea consumption and esophageal SCC risk (P for trend = 0.005), with a 0.5-fold lower risk associated with the intake of unfermented tea (green tea, oolong tea, or jasmine tea). The effects of dietary factors on esophageal SCC were similar in all subsites, with the exception of consumption of coffee. Coffee consumption was more pronounced in having a protective effect in the middle third section compared with the lower third section of the esophagus (adjusted odds ratio 0.4, 95% confidence interval 0.2-0.9), although this protective effect was marginally significant (adjusted odds ratio 0.6, 95% confidence interval 0.4-1.0) against esophageal SCC in all subsites. Our data also suggest that discomfort when eating hot foods may exert a carcinogenic effect by direct contact with the esophageal mucosa and tend to have more harmful effects in the upper than in the lower esophagus. In contrast, vegetables, fruits, and tea with components that are thought to inhibit carcinogenesis by absorbed components affected all subsites similarly., Conclusion: Our results add additional information that certain dietary components may affect carcinogenesis locally and systemically.

Background: Overexpression of Cyclooxygenase-2 (COX-2) was observed in many types of cancers, including esophageal squamous cell carcinoma (ESCC). One functional SNP, COX-2 -1195G/A, has been reported to mediate susceptibility of ESCC in Chinese populations. In our previous study, the presence of Helicobacter pylori (H. pylori) was found to play a protective role in development of ESCC. The interaction of COX-2 and H. pylori in gastric cancer was well investigated. However, literature on their interaction in ESCC risk is scarce. The purpose of this study was to evaluate the association and interaction between COX-2 single nucleotide polymorphism (SNP), H. pylori infection and the risk of developing ESCC., Methods: One hundred and eighty patients with ESCC and 194 controls were enrolled in this study. Personal data regarding related risk factors, including alcohol consumption, smoking habits and betel quid chewing, were collected via questionnaire. Genotypes of the COX-2 -1195 polymorphism were determined by PCR-based restriction fragment length polymorphism. H. pylori seropositivity was defined by immunochromatographic screening test. Data was analyzed by chi-squared tests and polytomous logistics regression., Results: In analysis adjusting for the covariates and confounders, H. pylori seropositivity was found to be inversely association with the ESCC development (adjusted OR: 0.5, 95% CI: 0.3 - 0.9). COX-2 -1195 AA homozygous was associated with an increased risk of contracting ESCC in comparison with the non-AA group, especially among patients with H. pylori seronegative (adjusted OR ratio: 2.9, 95% CI: 1.2 - 7.3). The effect was strengthened among patients with lower third ESCC (adjusted OR ratio: 6.9, 95% CI 2.1 - 22.5). Besides, H. pylori seropositivity conveyed a notably inverse effect among patients with COX-2 AA polymorphism (AOR ratio: 0.3, 95% CI: 0.1 - 0.9), and the effect was observed to be enhanced for the lower third ESCC patients (AOR ratio: 0.09, 95% CI: 0.02 - 0.47, p for multiplicative interaction 0.008), Conclusion: H. pylori seropositivity is inversely associated with the risk of ESCC in Taiwan, and COX-2 -1195 polymorphism plays a role in modifying the influence between H. pylori and ESCC, especially in lower third esophagus.

Background: The purpose of this study was to better understand the role of osteopontin (OPN) in esophageal squamous cell carcinoma (ESCC) by comparing the OPN mRNA level in ESCC tumor tissue and matched normal tissue and by determining the prognostic significance of the gene expression., Methods: Initially, by an oligo-nucleotide microarray hybridization technique, OPN expressions were found to consistently elevate at least twofold in three ESCC tissues compared with their adjacent normal ones. Subsequently, the expression of OPN mRNA was detected by real-time QRT-PCR among the 58 fresh surgical ESCC specimens. The clinical information was obtained by chart review., Results: OPN mRNA expression was detectable in 58 of 58 (100%) tumor specimens and 57 of 58 (98.3%) nonmalignant esophageal specimens. OPN expression was higher in tumor tissue than in the matched normal tissue in 54 of 58 (93.1%) individual cases. The overall median mRNA expression level of OPN was approximately 8.8-fold higher in tumor tissues (4.1; range: 0.02-247) compared with matched normal esophageal tissues (0.5; range, 0.0-21.4; p < 0.001). Overexpression of OPN mRNA was significantly associated with clinical stage (p = 0.01). The more severe the clinical stage (from I-II, III, to IV) was the higher frequency of overexpression of OPN mRNA. No significant associations were found between overexpression of OPN mRNA and the patients' survival (p = 0.27)., Discussion: Our findings suggest OPN is associated with esophageal tumorigenesis and progression, but not patients' survival.

Methylation of the O(6)-methylguanine-DNA-methyltransferase (MGMT) promoter is associated with G:C to A:T transitions in the p53 gene in various human cancers, including lung cancer. In tumors with p53 mutation, MGMT promoter methylation is more common in advanced tumors than in early tumors. However, in tumors with wild-type p53, MGMT promoter methylation is independent of tumor stage. To elucidate whether p53 participates in MGMT promoter methylation, we engineered three cell models: A549 cells with RNA interference (RNAi)-mediated knockdown of p53, and p53 null H1299 cells transfected with either wild-type p53 (WT-p53) or mutant-p53 (L194R, and R249S-p53). Knockdown of endogenous p53 increased MGMT promoter methylation in A549 cells, and transient expression of WT-p53 in p53 null H1299 cells diminished MGMT promoter methylation, whereas the MGMT promoter methylation status were unchanged by expression of mutant-p53. Previous work showed that p53 modulates DNA-methyltransferase 1 (DNMT1) expression; we additionally examined chromatin remodeling proteins expression levels of histone deacetylase 1 (HDAC1). We found that p53 knockdown elevated expression of both DNMT1 and HDAC1 in A549 cells. Conversely, expressing WT-p53 in p53 null H1299 cells reduced DNMT1 and HDAC1 expression, but the reduction of both proteins was not observed in expressing mutant-p53 H1299 cells. CHIP analysis further showed that DNMT1 and HDAC1 binding to the MGMT promoter was increased by MGMT promoter methylation and decreased by MGMT promoter demethylation. In conclusion, MGMT promoter methylation modulated by p53 status could partially promote p53 mutation occurrence in advanced lung tumors.

Pedunculated lipoma of the esophagus is rare and easily misdiagnosed in clinical practice. The presenting symptoms of esophageal lipoma are dysphagia, regurgitated mass and persistent sensation of a lump in the throat. The most frequent location of the tumor pedicle is the upper esophageal sphincter. Although the lipoma is pathologically benign, if it is large enough, it may cause airway obstruction secondary to the mechanical pressure to the larynx when the tumor is regurgitated. We present the case of a 67-year-old man who had the symptoms of dysphagia, nausea and vomiting. Esophagography and chest computed tomography revealed that he might have an esophageal submucosal or intraluminal tumor mass. Panendoscopy showed a pedunculated tumor mass within the esophageal lumen with its peduncle arising from the cervical esophagus. The tumor mass measured 9.0 x 4.7 x 2.5 cm in size. Thoracic approach via the right chest wall was performed for confirmation. After removal of the intraluminal mass, the patients symptoms dramatically improved. Pathology showed a lipoma arising from the submucosa of the esophagus.

The consumption of alcohol, tobacco and betel quid has been found to be an important contributor to esophageal squamous cell carcinoma (ESCC) in Taiwan. The genotoxic effect of the ADH1B and ALDH2 genes modulating an individual's alcohol-metabolizing capacity on ESCC may be linked to drinking behavior, intake pattern and other exogenous factors. To investigate the interplay of these genetic and environmental factors in determining the risk of ESCC, a multicenter case-control study was conducted. Here, 406 patients with pathology-proven ESCC, as well as 656 gender, age and study hospital matched controls were recruited. Genetic polymorphisms of ADH1B and ALDH2 appeared to correlate with the abstinence of alcohol, though not with tobacco and betel quid. Within the same levels of alcohol consumption, carcinoma risks increased along with an increase in the numbers of ADH1B*1 and ALDH2*2 alleles. The inactive ALDH2*1/*2 genotype was found to multiplicatively interact with a low-to-moderate (0.1-30 g/day) and a heavy (>30 g/day) ethanol intake to increase the ESCC risk (the joint aOR = 14.5 and 102.6, respectively). Among low-to-moderate drinkers, a smoking-dependent carcinogenetic effect for the ADH1B*1/*1 and ALDH2*1/*2+*2/*2 genotypes was recognized, with significant risks found in smokers, but not in nonsmokers. Further, a supra-multiplicative combined risk of ESCC for alcohol and tobacco use was identified among carriers of the ADH1B*1/*1 genotype (p for interaction = 0.042). In conclusion, the interplay of the ADH1B and ALDH2 genotypes, in conjunction with a behaved drinking habit and a practiced drinking pattern, along with continued tobacco consumption, plays an important pathogenic role in modulating ESCC risk., ((c) 2007 Wiley-Liss, Inc.)

Inactivation of p53 by human papillomavirus 16/18 E6 plays a crucial role in cervical tumorigenesis. To investigate the involvement of HPV16/18 in lung tumorigenesis, the association between HPV16 or HPV18 E6 and p53 protein expression in 122 lung tumors was evaluated by immunohistochemistry, and data showed that HPV16/18 E6 expression correlated inversely with p53 expression, which was further confirmed by tissue in situ immunostaining. Real-time reverse transcription-PCR analysis indicated that E6-positive tumors had lower p21(WAF1/CIP1) and mdm2 mRNA levels than E6-negative tumors. To elucidate the role of E6 in p53 inactivation, we successfully established lung adenocarcinoma cell lines with or without HPV16 infection from patients' pleural effusions. Western blotting showed that E6 protein was indeed expressed in HPV16-infected cells and a lower level of p53 protein was observed in E6-positive cells compared with E6-negative cells. Moreover, the levels of p21(WAF1/CIP1) and mdm2 mRNA in E6-positive cells were lower than in E6-negative cells. The interaction of E6 with p53 protein was revealed by immunoprecipitation assay showing that p53 could be inactivated by E6 protein. Conversely, p53 proteins and p21(WAF1/CIP1) and mdm2 mRNA expressions were restored in E6-knockdown cells by RNA interference compared with control cells. These results reveal that HPV16/18 E6 may be partially involved in p53 inactivation to down-regulate p21(WAF1/CIP1) and mdm2 transcription. In conclusion, HPV16/18 E6 is indeed expressed in HPV DNA-positive lung tumors and is involved in p53 inactivation to contributing to HPV-mediated lung tumorigenesis.

Epidermal growth factor receptor (EGFR) gene copy number correlates with response to tyrosine kinase inhibitors in patients with nonsmall cell lung carcinoma. Fluorescence in situ hybridization (FISH), a standard methodology to detect EGFR copy number abnormalities in nonsmall cell lung carcinoma, is limited by instrumentation and cost. Chromogenic in situ hybridization (CISH) is an emerging alternative detection technique using light microscopy, but its utility in assessing EGFR copy number in lung cancer is not established. To address the utility of CISH, we studied paraffin-embedded nonsmall cell lung carcinoma specimens from 77 Taiwanese nonsmoking women treated by surgery alone. We recorded the number of signals per tumor cell nucleus, correlated EGFR copy number by CISH with FISH results, and used receiver operating characteristics to identify cut-off points for the CISH results. Tumors were classified as adenocarcinoma (n=28), mixed adenocarcinoma with bronchioloalveolar features (n=25), bronchioloalveolar carcinoma (n=2), squamous cell carcinoma (n=15), and adenosquamous carcinoma (n=7). By FISH, 29% of cases had no amplification, 18% had low polysomy, 35% had high polysomy, and 12% had gene amplification. EGFR copy number detected by CISH highly correlated with FISH (Spearman r=0.81, P<0.0001). We determined the optimal EGFR CISH cut-off points that discriminate between no amplification and low polysomy (2.8 signals, P=0.09); no amplification plus low polysomy and high polysomy plus gene amplification (4.5 signals, P<0.0001); and high polysomy and gene amplification (7.1 signals, P=0.0003). CISH is an alternative assay to FISH in determining EGFR copy number status that may contribute to stratification of patients with nonsmall cell lung carcinoma for clinical trials and identify a subset of patients that should be treated with tyrosine kinase inhibitors.

Consumption of alcohol and tobacco, separately or jointly, can increase the risk of oesophageal squamous cell carcinoma (OSCC). It is unclear whether the amount of alcohol consumption by individual drinkers affects the joint carcinogenetic action of both agents. To demonstrate how the intensity of alcohol intake determines the risk of OSCC in relation to tobacco smoking, we conducted a multicentre case-control study. A total of 652 patients with pathology-proven OSCC, as well as 1127 gender, age, and study hospital matched controls were recruited. To identify a possible curvature in the continuous relationship between exposure and risk, we applied the generalised additive models to the collected data. Both non-drinkers who smoked tobacco and non-smokers who drank heavy alcohol (>30 g/day) were observed to have elevated cancer risks. A smoking habit-specific, non-linear increase in oesophageal cancer risk was recognised. Tobacco was found to interact with light-to-moderate alcohol (0.1-30 g/day) to increase the risk of oesophageal cancer in a supra-multiplicative way (Odds ratio (OR) ratio=5.5-5.7, p<0.05), whereas with heavy alcohol consumption in a simple multiplicative model (OR ratio=1.7-2.3, p>0.05). Weekly intake frequency had the strongest influence on the risk of neoplasm development. Alcohol consumption was responsible, respectively, for 18% and 77% of nonsmoking and smoking OSCC cases in this population. In conclusion, both light-to-moderate and heavy alcohol intake interact separately with tobacco in differently synergistic processes that can determine the development of this type of cancer.

The carcinogenetic impact of risk factors on esophageal cancer (EC) may differ according to the portion of the esophagus where the tumor occurs. It is unclear why more esophageal squamous cell carcinomas (SCC) developed in the middle location. We carried out a multicenter case-control study in Taiwan to assess anatomical subsite risk discrepancy for this neoplasm in regard to the consumption of alcohol, tobacco and betel quid. Four hundred forty seven incident patients with pathology-proven SCC of the esophagus (107 were upper-third [U/3-EC], 199 middle-third [M/3-EC] and 141 lower-third [L/3-EC] cases), as well as 1,022 gender, age and study hospital matched controls were analyzed by unordered polytomous logistic regression. All consumption of the three substances was related to the development of each subsite of EC, with a heterogeneously higher risk for current smokers (adjusted odds ratio (AOR) = 6.2) found in M/3-EC and for current chewers, in U/3-EC (AOR = 4.9). The joint risk of contracting lower two-third EC for drinking and smoking appeared to significantly surpass those estimated by a multiplicative interaction model. Concomitant exposure to these two agents brought the risks of EC at all three subsites up to 10- to 23.9-fold and additional tobacco-free betel quid to a 30.3- to 75.0-fold. In conclusion, tumor subsite discrepancy risk is related to prolonged exposure to tobacco and betel quid with inflorescence. Alcohol interacts with tobacco in a stronger supra-multiplicative way in the middle portion of the esophagus, probably explaining why esophageal SCC occurs more commonly at this anatomical location., ((c) 2007 Wiley-Liss, Inc.)

Objective: Esophageal squamous cell carcinoma and adenocarcinoma were increasingly recognized as two entities with different biologic behaviors and prognosis. Surgical risks and oncologic benefits of transthoracic and transhiatal esophagectomy for esophageal squamous cell carcinoma patients are not confessed., Methods: Between 1994 and 2005, 216 esophageal squamous cell carcinoma patients underwent esophagectomy were enrolled and analyzed retrospectively., Results: One hundred sixty-six patients underwent transthoracic esophagectomy and 50 patients underwent transhiatal esophagectomy. The overall hospital mortality and postoperative complication rates were 9.7 and 49%, respectively. The amount of intra-operative blood loss or transfusion, postoperative complication rate, lengths of hospital stay and hospital mortality rate were not significantly different between both groups. However, shorter operative time was noticed in transhiatal group (p<0.001). The overall 5-year survival rate was 16.8%. ESCC patients underwent either transthoracic or transhiatal esophagectomy had comparable long-term survival. The pTNM stage was independent prognostic factor for patients underwent transthoracic esophagectomy. However, location of tumor (p=0.009) and pathologic tumor length (p=0.012) were predictors of prognosis for patients underwent transhiatal esophagectomy., Conclusions: For esophageal squamous cell carcinoma patients, no significant differences in postoperative mortality or morbidity rates were observed between transthoracic and transhiatal esophagectomy. However, traditional pTNM staging system might underestimate the severity of esophageal squamous cell carcinoma patients who underwent transhiatal esophagectomy. The information of dissimilar prognostic factors for transhiatal or transthoracic esophagectomies will be helpful in tailoring more individualized adjuvant therapy to optimize esophageal squamous cell carcinoma patient's outcome.

In our previous study, we found that polymorphisms of alcohol and aldehyde dehydrogenase (ADH1B and ALDH2) are important risks for esophageal squamous cell carcinoma in a Taiwanese population. In this study, we increased the sample size to investigate the modifying effect of lifetime alcohol consumption on the association between ADH1B and ALDH2 genotypes and the risks of esophageal cancer. A multicenter hospital-based case-control study was conducted between August 2000 and June 2004. Three hundred and thirty newly-diagnosed esophageal squamous cell carcinoma patients and 592 controls were recruited from National Taiwan University Hospital in Taipei and Kaohsiung Veterans General Hospital and Kaohsiung Medical University Hospital in Kaohsiung, Taiwan. Controls were matched to the case patients by gender and age within 4 years (case:control = 1:1-4). Polymorphisms of ADH1B and ALDH2 were genotyped by the method of PCR-RFLP. Individuals with ADH1B*1/*1 genotype had a 3.99-fold risk (95% CI = 2.13-7.48) of developing esophageal cancer, compared with those with ADH1B*2/*2 genotype, after adjusted for appropriate covariates. Individuals with ALDH2*1/*2 and ALDH2*2/*2 had 4.99-fold risk (95% CI = 3.11-7.99) and 4.24-fold risk (95% CI = 1.52-11.84), respectively, of developing esophageal cancer, compared with those with ALDH2*1/*1, after adjusted for appropriate covariates. We also found a modifying effect of lifetime alcoholic consumption on the association between genotypes of ADH1B and ALDH2 on esophageal cancer risk. These results suggest that ADH1B and ALDH2 polymorphisms play a pivotal role on esophageal cancer and that the effect of these polymorphisms was modified by the amount of alcohol consumed., (Copyright 2006 Wiley-Liss, Inc.)

Background: Inconsistency in the prevalence of infection by human papillomavirus (HPV) in lung cancer patients was found between different countries with racial and geographic variations. Our previous reports have indicated that a high-risk HPV 16/18 DNA was frequently detected in Chinese lung cancer patients living in Taichung, Taiwan (Cheng et al. Cancer Res. 2001;61:2799-803). Thus, we conducted this study to verify whether there was a similar HPV 16/18 infection prevalence in lung cancer patients from Wuhan, China., Methods: To reduce the false positive HPV detection, the paraffin sections of 73 lung tumors and 34 non-cancer controls from Wuhan, China were collected for detection of the presence of HPV 16/18 DNA by in situ hybridization (ISH)., Results: Our results showed that the rates of HPV 16 and/or 18 infections in patients with lung tumors were significantly higher than in 34 non-cancer control subjects (26.0 versus 2.8% for HPV 16, P = 0.030; 23.3 versus 5.7% for HPV 18, P = 0.031; 27.7 versus 5.9% for HPV 16 or 18, P = 0.003) with a similar infection frequency of HPV 16 and 18 types in lung tumors. This result indicated that HPV 16/18 infection may be associated with lung cancer development in Chinese patients from Wuhan, China. Further statistical analyses revealed that HPV 16 or 18 infection was not correlated with any clinico-pathological parameter studied, including age, gender, smoking status, tumor type, tumor stage and tumor grades. Interestingly, smoking and male patients had a higher prevalence of HPV 16, although not reaching a statistical significance, compared with non-smoking and female patients, respectively (33.3% for smokers versus 20.0% non-smokers; 33.3% for male versus 17.6% for female). As compared with the HPV 16/18 infection in Taiwan, Chinese patients with lung cancer from Wuhan had a different HPV 16/18 infection prevalence., Conclusion: Difference in HPV 16/18 infection in lung cancer patients from Wuhan, China and Taichung, Taiwan suggests that HPV 16/18 might play a different role in lung cancer development among Chinese living in different areas.

A recent report suggested that BQ (BQ) chewing significantly correlated with the occurrence of esophageal squamous cell carcinoma (ESCC) in Taiwanese. BQ chewing was shown to be associated with p53 mutation in oral cancers. However, the relationship between BQ chewing and p53 mutation in ESCC is unclear. Seventy-five primary ESCC patients were enrolled for mutational analysis of the p53 gene using polymerase chain amplification and direct sequencing of amplified product. Thirty-seven mutations of the p53 gene were detected in 45.5% (34/75) of tumor specimens. These mutations significantly clustered in exon 5 (21/37) of the p53 gene. The incidence of p53 mutations did not associate with clinicopathological characteristics or the habits of cigarette smoking or alcohol consumption. However, BQ chewers exhibited significantly higher incidence of p53 gene mutations than non-chewers (67.6% vs 32.4%, P = 0.007). After controlling the confounding factors of cigarette smoking and alcohol intake, BQ chewing still showed significant association with the incidence of p53 mutation in ESCCs (RR = 4.23; 95% CI, 1.317-13.60). The A:T to G:C transition (8/37, 21.6%) and G:C to T:A transversion (5/23, 13.5%) were the prevalent spectrum of p53 gene mutations. All A:T to G:C transitional mutations occurred in patients with the habits of BQ chewing and cigarette smoking. Noticeably, alcohol consumption could enhance this peculiar spectrum of p53 mutation in ESCC. Accordingly, p53 might be an important molecular target of BQ carcinogens in the development of ESCC in Taiwanese.

Background: Prognostic effects of cell cycle regulation associated tumor suppressor genes for esophageal squamous cell carcinoma patients are controversial. Clarifying the effects of these genes is beneficial for optimizing patient's outcome., Methods: Forty esophageal squamous cell carcinoma patients were included in the study. Tissue samples were analyzed for cell proliferation, DNA content, mutation of p53 gene, and expression of p21 and p53 proteins. Prognostic effects of these parameters were assessed by multivariate analysis., Results: Seventy-five percent of tumors exhibited aneuploid DNA content. Significantly higher S-phase fractions were detected in tumor samples (p < 0.001). The p53 gene mutated in 52.5% (21 of 40) of tumor samples, p53 immunostaining was detected in 62.5% (25 of 40) of tumor tissues and 50% of tumors over expressed p21. Overexpression of p21 protein did not correlate with p53 gene status, but significantly correlated with abnormal DNA content (p = 0.028). Advanced pTNM stage, lymph node metastasis and p21 overexpression conferred survival disadvantages in univariate analysis (p = 0.013, 0.045 and 0.017, respectively). A Cox multivariable analysis revealed pTNM stage (IIB/III/IV vs. I/IIA; p = 0.024) associated with p21 overexpression (positive vs. negative; p = 0.035) as independent prognostic factors in esophageal squamous cell carcinomas. Surprisingly, p21 overexpression significantly compromised the survival of patients with mutated p53 gene (p = 0.035). However, no significant dismal effect of p21 overexpression can be seen in patients with wild-type p53 gene (p = 0.175)., Conclusions: Overexpression of p21 correlates with chromosomal instability and serves as an adverse prognostic predictor for patients with esophageal squamous cell carcinoma. Its adverse effect is more prominent when the p53 gene is mutated.

In this study, human oropharyngeal epidermoid carcinoma KB cells that were resistant to 2,2-difluorodeoxycytidine (dFdCyd) were selected and designated the KB-Gem clone. The KB parental cell line IC50 was 0.3 microM dFdCyd, as compared with the KB-Gem clone IC50 of 32 microM dFdCyd. The KB-Gem clone demonstrated overexpression of ribonucleotide reductase (RR) M2 subunit mRNA (9-fold) and overexpression of M2 protein (2-fold); RR activity was 2.3-fold higher than the KB parental cell line. Both the dATP and dCTP pools of the KB-Gem clone increased 2-fold over the parental cell line, with no change in the dGTP and dTTP pools. Reverse transcriptase-PCR was used to clone the cDNA of deoxycytidine kinase (DCK). Resulting sequences revealed two silent mutations in the KB-Gem clone. The amino acid sequence of the DCK protein and mRNA expression remained unchanged. The KB-Gem clone's DCK enzyme activity was 56% of that of the parental cell line. After the endogenous dNTPs were removed with a G-25 column, no difference was evident between the enzyme activities of the KB-Gem clone and parental cells. Thus, contrary to previous hypotheses, DCK deficiency does not play the primary role in the resistance mechanism of dFdCyd, accepting a secondary role to the overexpression of the target gene, RR, and pool expansion.

Background: Although several retrospective studies have been published concerning nonoperative management of minor liver and spleen injuries, few studies have prospectively analyzed the results of nonoperative management for higher-grade liver and spleen injuries. Is it possible to manage extensive hepatic or splenic injuries with hemoperitoneum nonoperatively? The current study was conducted to evaluate the safety of nonoperative management of blunt hepatic and splenic trauma with significant hemoperitoneum in hemodynamically stable patients regardless of injury severity., Methods: We used the nonoperative methods prospectively to treat consecutive patients with blunt spleen or liver injury during a 35-month period. Patients with unstable conditions underwent emergency laparotomies, and those who were stable underwent abdominal computed tomography for further evaluation. We analyzed the clinical characteristics and the success rate of this method thoroughly., Results: Twenty-four patients with severe hepatic or splenic injuries treated nonoperatively were included in this study. Among these 24 patients, 18 (75%) with hepatic or splenic injuries had grades of III or greater on the Organ Injury Scale. Twenty patients (83.3%) had moderate to large amounts of hemoperitoneum. Four patients (16.7%) failed at observation and underwent emergency celiotomy, two for liver-related and two for spleen-related causes. There were no differences between the nonoperative and operative management groups in terms of mean age, initial systolic blood pressure, initial heart rate, emergency room fluid requirement except emergency blood transfusion, abdominal complications, and hospital length of stay., Conclusion: We suggest that nonoperative management may be undertaken successfully in appropriately designed areas with close observation for the hemodynamic stable patient.