Glucose transporters 1, 3, 6, and 10 are expressed in gastric cancer and glucose transporter 3 is associated with UICC stage and survival.

BACKGROUND: Due to proliferation and increased metabolism, cancer cells have high glucose requirements. The glucose uptake of cells is influenced by a group of membrane proteins denoted the glucose transporter family (Glut-1 to -12). Whereas increased expression and a negative correlation with survival have been described for Glut-1 in several types of cancer, the impact of other glucose transporters on tumor biology is widely unknown. METHODS: In this retrospective study, gastric cancer specimens of 150 patients who underwent total gastrectomy between 2005 and 2010 were stained for Glut-1, -3, -6, and -10 by immunohistochemistry. Expression of Glut-1, -3, -6, and 10 was correlated to prognosis as well as clinical and pathological parameters. RESULTS: Glut-1, Glut-3, Glut-6, and Glut-10 were expressed in 22.0, 66.0, 38.0, and 43.3 % of the analyzed samples. Whereas Glut-1, -6, and -10 did not show a correlation with prognosis, positive staining for Glut-3 was associated with higher UICC stage and inferior prognosis. The mean overall survival was 38.6 months for Glut-3 positive patients, as compared to 51.2 months for Glut-3 negative patients (p < 0.05). Coexpression of two or more of the analyzed glucose transporters was correlated to inferior prognosis. Glut-3 and UICC stage were significant prognostic factors in multivariate analysis. CONCLUSIONS: All of the analyzed glucose transporters were expressed in a significant proportion of the gastric cancer samples. Glut-3 was associated with higher UICC stage and inferior prognosis. These findings are relevant to therapeutic approaches that target glucose metabolism as well as to imaging using radioactively labeled glucose.

Death-associated protein kinase (DAPK) promoter methylation and response to neoadjuvant radiochemotherapy in esophageal cancer.

BACKGROUND: Multiple studies have shown that promoter methylation of tumor suppressor genes underlies esophageal carcinogenesis. Hypothetically, methylation resulting in tumor suppressor gene inactivation might result in tumors that are unresponsive to chemotherapy and radiation. Accordingly, our aim was to investigate if aberrant methylation of the apoptosis-related gene Death-Associated Protein Kinase (DAPK) could be used as a predictor of response to neoadjuvant therapy in locally advanced cancer of the esophagus. METHODS: Tumor and normal esophageal tissues were obtained from 50 patients with locally advanced cancer of the esophagus prior to neoadjuvant radiochemotherapy. DAPK methylation analysis was performed on all samples by methylation-specific real-time polymerase chain reaction (PCR). RESULTS: Seventeen (34%) patients showed a major and 33 (66%) a minor histomorphological response to neoadjuvant therapy. DAPK methylation was detectable in normal esophageal tissues with a frequency of 10% and in tumor tissue with a frequency of 78%. The median methylation level for DAPK was 2.7 x 10(-3) in tumor compared with 0.1 x 10(-3) in normal tissues (p < 0.001). DAPK methylation was not associated with response to neoadjuvant therapy or prognosis after esophagectomy. CONCLUSION: Aberrant DAPK methylation in tumor tissues is significantly higher compared with matching normal esophageal tissues, suggesting a fundamental role of this epigenetic alteration in the pathogenesis of this disease. The level of DAPK methylation in pretreatment biopsies of patients with locally advanced cancer of the esophagus is no marker for the prediction of histomorphological regression or prognosis following neoadjuvant chemoradiation in this disease.

The role of the homeobox genes BFT and CDX2 in the pathogenesis of non-small cell lung cancer.

BACKGROUND: The role of the homeobox genes Backfoot (BFT) and caudal-related Homeobox 2 (CDX2) in the pathogenesis of non-small cell lung cancer (NSCLC) is unclear. The goal of this study was to investigate the mRNA expression of BFT and CDX2 in NSCLC and to determine the association with the pathogenesis and the potential as a biomarker of this disease. MATERIALS AND METHODS: The mRNA expression of BFT and CDX2 was analyzed by quantitative real-time RT-PCR in the tumor and matching normal tissue from 23 patients with NSCLC. RESULTS: The mRNA expression was detectable with the following frequencies in the tumor (t) and normal (n) tissues: BFT=100% (n), 100% (t); CDX2=100% (n), 100% (t). The median CDX2 mRNA expression was 0.85 (range: 0.01-15.47) in the tumor tissue and 0.045 (range: 0-1.36) in the matching normal lung tissue (p=0.001). The median BFT mRNA expression was 0.0034 (range: 0-0.35) in the tumor tissue and 0.0001 (range: 0-0.10) in the matching normal lung tissue (p=n.s.). There were no associations between the mRNA expression levels of BFT and CDX2 and clinicopathological variables. CONCLUSION: The mRNA expression of the homeobox genes is detectable at a high frequency in the tumor and normal tissue of patients with non-small cell lung cancer. Up-regulation of CDX2 mRNA expression appears to be associated with the pathogenesis of this malignant disease. The quantification of CDX2 and BFT mRNA expression in lung tissue is a potential biomarker for the identification of patients at risk of the development of NSCLC.

The upregulation of hypoxia-related miRNA 210 in primary tumor of lymphogenic metastatic prostate cancer.

AIM: To show the association between the expression level of hsa-miR-210 (miR-210) and tumor progression in prostate cancer (PCa). METHODS: Quantitative PCR was performed to measure miR-210 on 55 subjects with different tumor stages; our results were then validated using three external datasets. ANOVA and Tukey's post hoc analysis were performed for comparative analyses between different tumor stages. Using the transcriptome data from The Cancer Genome Atlas for CaP, the gene expression analyses were performed on experimentally validated target genes of miR-210 identified in Tarbase and miRWalk datasets. RESULTS & CONCLUSION: miR-210 was significantly higher in N1 PCa compared with nonmetastatic PCa, whereas the metastatic tumor revealed a lower expression level of miR-210 than the primary tumor.

COX-2 mRNA expression is significantly increased in acid-exposed compared to nonexposed squamous epithelium in gastroesophageal reflux disease.

BACKGROUND: Little is known about the role of cyclooxygenase (COX)-2 in gastroesophageal reflux disease (GERD) and the development of Barrett's metaplasia. The objectives of this study were to further analyze COX-2 mRNA expression in patients with GERD compared to Barrett's esophagus (BE) and Barrett's cancer (BC). METHODS: Tissue samples from 110 patients with GERD (n = 43), BE (n = 20), and BC (n = 47) were obtained in routine upper GI endoscopy. Expression levels of COX-2 were measured by quantitative real-time reverse trancriptase polymerase chain reaction (RT-PCR). Also, 24-h pH monitoring was performed in all patients of the GERD study group and the DeMeester composite score was used to match COX-2 mRNA expression with the severity of acid exposure in the lower esophagus. RESULTS: COX-2 mRNA is progressively upregulated within the metaplasia-dysplasia-adenocarcinoma (MDA) sequence (p = 0.001). COX-2 levels of the squamous epithelium in the distal esophagus from patients with GERD and a pathologic mean DeMeester score (>14.72) were significantly higher than in patients with normal DeMeester scores (p = 0.01). CONCLUSION: In summary our findings suggest that alterations in COX-2 mRNA expression occur independently of endoscopic or histologic signs of GERD in the acid-exposed squamous epithelium of the distal esophagus. However, this early COX-2 increase in GERD is further upregulated within the MDA sequence for yet unknown reasons.

HIF-1alpha mRNA is not associated with histopathological regression following neoadjuvant chemoradiation in esophageal cancer.

BACKGROUND: Hypoxia-inducible factor-1alpha (HIF-1alpha) expression was reported to be associated with tumor growth, progression and resistance to radio-/chemotherapy. Whether HIF-1alpha mRNA or protein expression is associated with histomorphological response or prognosis following neoadjuvant chemoradiation and surgery in resectable, locally-advanced esophageal cancer was analyzed. PATIENTS AND METHODS: Fifty-three patients received neoadjuvant chemoradiation (cisplatin, 5-fluorouracil, 36 Gy) followed by transthoracic en bloc esophagectomy. HIF-1alpha mRNA and protein expressions were analyzed by quantitative RT-PCR and immunostaining. RESULTS: In squamous cell carcinoma, HIF-1alpha mRNA expression was significantly higher than in paired normal epithelium (p < 0.001). Normal squamous epithelium showed significant elevated expression in adenocarcinomas, suggesting a field effect (p < 0.04). HIF-1alpha protein expression showed a significant regulation following chemoradiation. Neither HIF-1alpha mRNA nor protein expression was associated with histomorphological regression or prognosis. CONCLUSION: HIF-1alpha mRNA expression is differentially upregulated in esophageal squamous cell carcinoma compared to adenocarcinomas, but does not predict tumor regression or prognosis.

High cyclooxygenase-2 expression following neoadjuvant radiochemotherapy is associated with minor histopathologic response and poor prognosis in esophageal cancer.

PURPOSE: High expression of cyclooxygenase-2 (COX-2) was shown to inhibit chemotherapy- and radiotherapy-induced apoptosis. We analyzed the association of COX-2 mRNA and protein expression with histomorphologic response to neoadjuvant radiochemotherapy in esophageal cancer. EXPERIMENTAL DESIGN: Fifty-two patients with resectable esophageal cancers (cT2-4, Nx, and M0) received neoadjuvant radiochemotherapy (cisplatin, 5-5-fluorouracil, 36 Gy) followed by transthoracic en bloc esophagectomy. Histomorphologic regression was defined as major response when resected specimens contained less than 10% of residual vital tumor cells. RNA was isolated from endoscopic biopsies (paired tumor and normal tissue) before neoadjuvant treatment and quantitative real-time reverse transcriptase-PCR (Taqman) assays were done to determine COX-2 mRNA expression levels standardized for beta-actin. COX-2 protein expression in pretreatment biopsies and post-therapeutic resection specimens was analyzed by immunostaining of tumor cells. RESULTS: Median COX-2 mRNA expression levels were significantly (P < 0.0001) different between paired tumor (median, 2.2) and normal tissues (median, 0.159). Comparison of pre-therapeutic and posttherapeutic specimens showed a significant difference (P < 0.006) in COX-2 protein expression. Twelve of 52 tumors showed down-regulation and 3 of 52 showed up-regulation of COX-2 protein expression during neoadjuvant radiochemotherapy. High COX-2 protein expression in post-therapeutic resection specimens was significantly associated with minor histopathologic response (P < 0.04) and poor prognosis (5-year survival probabilities: 26.3 +/- 8.2% for minor and 58.6% +/- 12.9% for major histopathologic response; P < 0.01). CONCLUSION: High COX-2 protein expression following neoadjuvant radiochemotherapy in resection specimens is significantly associated with minor histopathologic response to neoadjuvant therapy and very poor prognosis.

High specificity of quantitative excision repair cross-complementing 1 messenger RNA expression for prediction of minor histopathological response to neoadjuvant radiochemotherapy in esophageal cancer.

PURPOSE: The excision repair cross-complementing 1 (ERCC1) gene is coding for a nucleotide excision repair protein involved in the repair of radiation- and chemotherapy-induced DNA damage. We examined the potential of quantitative ERCC1 mRNA expression to predict minor or major histopathological response to neoadjuvant radiochemotherapy (cisplatin, 5-fluorouracil, and 36 Gy of radiation) followed by transthoracic en bloc esophagectomy in patients with locally advanced esophageal cancer (cT(2-4), N(x), M(0)). EXPERIMENTAL DESIGN: Tissue samples were collected by endoscopic biopsy before treatment. RNA was isolated from biopsies, and quantitative real-time reverse transcriptase PCR assays were performed to determine ERCC1 mRNA expression. Relative mRNA levels (tumor/normal ratios) were calculated as (ERCC1/beta-actin in tumor)/(ERCC1/beta-actin in paired normal tissue). ERCC1 expression levels were correlated with the objective histopathological response in resected specimens. Histomorphological regression was defined as major response when resected specimens contained <10% of residual vital tumor cells or in case a pathologically complete response was achieved. RESULTS: Twelve of 36 tumors showed a major histopathological response, and 24 of 36 showed a minor histopathological response. Relative expression levels of ERCC1 of >1.09 were not associated with a major histopathological response (sensitivity, 62.5%; specificity, 100%) and 15 of 24 patients with minor histopathological response to the delivered neoadjuvant radiochemotherapy could be unequivocally identified. This association of dichotomized relative ERCC1 mRNA expression and histopathological response was statistically significant (P < 0.001). CONCLUSIONS: Relative expression levels of ERCC1 mRNA determined by quantitative real-time reverse transcriptase-PCR appear highly specific to predict minor response to our neoadjuvant radiochemotherapy protocol in patients with locally advanced esophageal cancer and could be applied to prevent expensive, noneffective, and potentially harmful therapies in a substantial number (42%) of patients.

Roles of thymidylate synthase and dihydropyrimidine dehydrogenase expression in blood as predictors of response to multimodal therapy in esophageal cancer.

BACKGROUND: Thymidylate synthetase (TS) and dihydropyrimidine dehydrogenase (DPD) RNA expression in peripheral blood was examined as a noninvasive molecular predictor of response to neoadjuvant radiochemotherapy in patients with locally advanced cancer of the esophagus. METHODS: Blood samples were drawn from 29 patients with esophageal cancer (10 squamous cell carcinomas and 19 adenocarciomas) before neoadjuvant radiochemotherapy. After extraction of cellular tumor RNA from blood samples, quantitative expression analysis of TS and DPD was performed with quantitative real-time reverse-transcription polymerase chain reaction. RESULTS: Twenty of 29 (68%) of patients had a minor histopathologic response, and 9 of 29 (32%) had a major response to neadjuvant radiochemotherapy. RNA expression in the blood of patients was detectable for TS in 86%, for DPD in 97%, and in 100% for ß-actin. No significant associations were detected between TS and DPD expression levels and clinical variables of the patients. A high expression level for TS was associated with a minor response to neoadjuvant treatment (P = .046), while there was no significant association between DPD and response to therapy. Combined analysis of TS and DPD expression increased the specificity for the prediction of response to 100%. No major responder to therapy had high expression levels for both genes in their peripheral blood. CONCLUSION: Quantitation of TS and DPD in peripheral blood may be a highly specific analysis to identify a subset of patients who do not respond to neoadjuvant radiochemotherapy and may therefore prevent expensive, noneffective, and potentially harmful therapies in a substantial number of patients with esophageal cancer.

Methylated DAPK and APC promoter DNA detection in peripheral blood is significantly associated with apparent residual tumor and outcome.

BACKGROUND: Death-associated protein kinase (DAPK) and adenomatous polyposis coli gene (APC) have been recently shown to be associated with outcome in patients with esophageal carcinoma, especially adenocarcinoma. We wanted to validate the correlation of these two markers with outcome by detecting methylated DNA sequences in peripheral blood. METHODS: Circulating cell-free DNA extracted from blood plasma of 59 patients with esophageal cancer was analyzed before and after surgical resection by quantitative real-time methylation-specific RT-PCR (TaqMan) assays. RESULTS: Thirty-six of 59 patients (61.0%) with esophageal cancer had detectable levels of methylated DAPK or APC promoter DNA and preoperative detection was significantly associated with an unfavorable prognosis as revealed by multivariate Cox proportional hazards regression analysis [Exp(b) = 4.578; P = 0.01]. The combination of both markers significantly increased sensitivity and specificity for discriminating between short (<2.5 years) and long survivors (P = 0.04, ROC curve analysis). Postoperative APC detection was significantly different if residual tumor was apparent (P = 0.03). CONCLUSIONS: Preoperative measurement of methylated DAPK and APC promoter DNA in peripheral blood may contribute to better estimate postoperative survival chances of patients with esophageal carcinoma, especially adenocarcinoma. The postoperative detection of methylated APC in peripheral blood might provide crucial information on apparent residual tumor and might be used as a "molecular" R0-Marker in addition to the pathologic examination.