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Abstract: Background: The effect of the apoptosis related gene Bcl-2 in the pathogenesis in NSCLC remains poorly investigated. Hence the aim of this study was to explore the potential role of Bcl-2 mRNA expression as a prognostic biomarker in patients with curatively resected NSCLC. Methods: 91 tumor and matching normal tissue samples from patients with NSCLC were analyzed using a quantitative real-time RT-PCR method. The relative Bcl-2 mRNA expression was measured using β-actin as a reference gene. 45 of the 91 patients had stage I tumors (49%), 19 had stage II (21%) and 27 had stage IIIa (30%). Squamous cell carcinoma was found in 43 patients (47%), adenocarcinoma in 33 (36%) and in large cell carcinoma in 15 (17%) of the patients. Results: Bcl-2 mRNA expression was detected in 83 (91%) of the investigated tumor samples and in 74 (81%) of the normal lung tissue. The median gene expression was 0.147 in tumor tissue and 0.144 in matching normal lung tissue (p =n.s., Wilcoxon Test). No associations were seen between the tumorous Bcl-2 mRNA expression levels and clinical or histopathologic parameters such as gender, tumor size, TNM stadium and grading, but with tumor histology and smoking. With a follow-up of 85.9 months, the median survival time was 59.7 months. Bcl-2 mRNA expression was significantly associated with patients prognosis (p =0.013, log-rank test). Multivariate regression analysis revealed Bcl-2 expression status and tumor stage as independent prognostic factor. Conclusions: Bcl-2 expression in NSCLC is not associated with the pathogenesis of this disease. Our data suggests that Bcl-2 mRNA expression plays a crucial role in the biological behavior of NSCLCs. Quantitation of Bcl-2 expression improves estimation of prognosis and appears to identify patients who will benefit from intensive adjuvant therapy. [ABSTRACT FROM AUTHOR]

PURPOSE: The role of adjuvant chemotherapy in patients with locally advanced bladder cancer still remains to be defined. We hypothesized that assessing the gene expression of the chemotherapy response modifiers multidrug resistance gene 1 (MDR1) and excision repair cross-complementing 1 (ERCC1) may help identify the group of patients benefiting from cisplatin-based adjuvant chemotherapy. EXPERIMENTAL DESIGN: Formalin-fixed paraffin-embedded tumor samples from 108 patients with locally advanced bladder cancer, who had been enrolled in AUO-AB 05/95, a phase 3 trial randomizing a maximum of three courses of adjuvant cisplatin and methotrexate (CM) versus methotrexate, vinblastine, epirubicin, and cisplatin (M-VEC), were included in the study. Tumor cells were retrieved by laser-captured microdissection and analyzed for MDR1 and ERCC1 expression using a quantitative real-time reverse transcription-polymerase chain reaction assay. Gene expression levels were correlated with clinical outcomes by multivariate Cox proportional hazards regression analysis. RESULTS: Expressions of MDR1 and ERCC1 were independently associated with overall progression-free survival (P = .001, relative risk = 2.9 and P = .01, relative risk = 2.24, respectively). The correlation of high MDR1 expression with inferior outcome was stronger in patients receiving M-VEC, whereas ERCC1 analysis performed equally in the CM and M-VEC groups. CONCLUSIONS: High MDR1 and ERCC1 gene expressions are associated with inferior outcome after cisplatin-based adjuvant chemotherapy for locally advanced bladder cancer. Prospective studies are warranted to define a role for MDR1 and ERCC1 analysis in individualizing multimodality treatment in locally advanced bladder cancer. [ABSTRACT FROM AUTHOR]

Background: Pancreatic cancer still has one of the worst prognoses of all cancers with a 5-year survival rate of 5%, making it necessary to find markers or gene sets that would further classify patients into different risk categories and thus allow more individually adapted multimodality treatment regimens. Especially heparanase (HPSE) has recently been discussed as a key factor in pancreatic cancer.Materials and Methods: Paraffin-embedded tissue samples were obtained from 41 patients with pancreatic adenocarcinoma who were scheduled for primary surgical resection. Direct quantitative real-time reverse transcriptase polymerase chain reaction (TaqMan) assays were performed in triplicates to determine HPSE, hypoxia inducible factor-1 alpha (HIF1a), platelet-derived growth factor alpha (PDGFA), heparin-binding EGF-like growth factor (HB-EGF), and basic fibroblast growth factor (bFGF) gene expression levels.Results: HPSE was significantly correlated to PDGFA (p = 0.04) and HIF1a (p = 0.04). The correlation of HIF1a to bFGF and HB-EGF was significant (p = 0.04, p = 0.02). Stepwise multiple linear regression models showed a significant independent association of HPSE with lymph node metastasis (p = 0.025) and with dedifferentiation (p = 0.042).Conclusions: Heparanase seems to be significantly associated with lymph node metastasis (p = 0.025) as well as dedifferentiation (p = 0.042). We assume that HPSE plays a crucial role for the aggressiveness of pancreatic cancer. Larger studies including more patients seem to be warranted. [ABSTRACT FROM AUTHOR]

PURPOSE: Pancreatic cancer still has one of the worst prognoses in gastrointestinal cancers with a 5-year survival rate of 5%, making it necessary to find markers or gene sets that would further classify patients into different risk categories and thus allow more individually adapted multimodality treatment regimens. In this study, we investigated the prognostic values of HIF1a, bFGF, VEGF, and PDGFA gene expressions as well as their interrelationships. EXPERIMENTAL DESIGN: Formalin-fixed paraffin-embedded tissue samples were obtained from 41 patients with pancreatic adenocarcinoma (age, 65; range, 34-85 years). After laser capture microdissection, direct quantitative real-time reverse transcription-polymerase chain reaction assays were performed in triplicates to determine HIF1a, PDGFA, VEGF, and bFGF gene expression levels. Multivariate Cox proportional hazards regression analysis was used to assess the impact of HIF1a gene expression on prognosis. RESULTS: HIF1a was significantly correlated to every gene we tested: bFGF (P = .04), VEGF (P = .02), and PDGFA (P = .03). Tumor size, P = .04, and high HIF1a mRNA expression (cutoff, 75th percentile) had a significant impact on survival, P = .009 (overall model fit, P = .02). High HIF1a expression had a sensitivity of 87.1% and a specificity of 55.6% for the diagnosis short (<6 months) versus long (6-60 months) survival. CONCLUSIONS: Measuring PDGFA, bFGF, and HIF1a expression may contribute to a better understanding of the prognosis of patients with pancreatic cancer and may even play a crucial role for the distribution of patients to multimodal therapeutic regimens. Larger studies including patients treated with actual chemotherapeutics seem to be warranted. [ABSTRACT FROM AUTHOR]

Reliable quantification of gene expression offers the possibility of more accurate and prognostically relevant characterization of tissues than potentially subjective interpretations of histopathologists. We measured the expression of 18 selected genes and compared them to histological features in a spectrum of esophageal disease to evaluate the feasibility of molecular characterization of normal and pathologic esophageal epithelia. Esophageal tissue biopsies from 82 patients with foregut symptoms were laser capture microdissected, and the expression levels of 18 selected genes were measured by quantitative real-time polymerase chain reaction. Linear discriminant analysis, which uses combinations of genes to distinguish between histological groups, was performed to compare gene expression and the following five histological groups: (1) normal squamous epithelium (n = 35), (2) reflux esophagitis (n = 13), (3) non-dysplastic Barrett's (n = 33), (4) dysplastic Barrett's (n = 16), (5) adenocarcinoma (n = 31). A panel of seven genes had 90-94% predictive power to distinguish non-dysplastic and dysplastic Barrett's esophagus. Clustering analysis revealed structure in gene expression values even in the absence of histology. Expression levels in 17 genes differed significantly across histological groups. Classification based on gene expression agreed with histopathological assessment in the following percentage of cases: normal squamous epithelium = 53%, reflux esophagitis = 31%, non-dysplastic Barrett's = 76%, dysplastic Barrett's = 40%, and adenocarcinoma = 59%. Interestingly, predictive power improved markedly when inflammatory and dysplastic tissues were removed (77-94%). Gene expression classification agrees well with histopathological examination. When differences occur, it is unclear whether this effect is due to intraobserver variability in pathological diagnosis or to a genuine difference between gene expression and histopathology. [ABSTRACT FROM AUTHOR]

Background: To find out if neoadjuvant therapy could alter tumor response determinants that might affect tumor sensitivity to the treatment, we investigated intratumoral expressions of genes associated with chemosensitivity, radiosensitivity, or both before and after radiochemotherapy. Study design: Twenty-four patients with locally advanced, resectable esophageal cancer (cT2–4,Nx,M0) received neoadjuvant 5-FU/cisplatin/36 Gy treatment followed by transthoracic en bloc esophagectomy. Expression levels of thymidylate synthase, dihydropyrimidine dehydrogenase, excision repair cross-complementing gene 1 , glutathione S-transferase Pi, epidermal growth factor receptor, and HER2 were measured in matched preradiochemotherapy endoscopic tumor biopsies and in postradiochemotherapy resection specimens. mRNA was isolated from formalin-fixed, paraffin-embedded, laser microdissected tumor tissues, and a quantitative fluorescent dye real-time reverse transcription polymerase chain reaction system was used for gene expression measurement. Results: There was a significant reduction in the expression levels of thymidylate synthase (p &#60; 0.01), dihydropyrimidine dehydrogenase (p = 0.03), excision repair cross-complementing gene 1 (p &#60; 0.01), glutathione S-transferase Pi (p = 0.03), HER2 (p &#60; 0.01), and epidermal growth factor receptor (p = 0.04). The change in the levels of epidermal growth factor receptor mRNA in post- compared with pretreatment specimens was significantly associated with the histopathologic grade of regression (p = 0.01). Conclusions: The expression levels of a set of genes that are possible determinants of 5-FU/cisplatin/radiation therapy antitumor activity are significantly downregulated by neoadjuvant radiochemotherapy in esophageal cancer. [Copyright &y& Elsevier]

Cyclooxygenase (Cox-2) is implicated in the pathogenesis of many cancers including esophageal adenocarcinoma (EAC), whereas the role of the isoform Cox-1 in carcinogenesis is not well understood. To further elucidate the role of these factors in the development of EAC, we measured the gene expressions (mRNA levels) of Cox-2 and Cox-1 by real-time quantitative polymerase chain reaction (QRT-PCR) in tissues from normal esophagus with and without erosive gastroesophageal reflux disease (GERD), Barrett''s esophagus (BE), dysplasia, adenocarcinoma, and in healthy gastric antrum. All tissues were purified by laser capture microdissection from endoscopic or surgical resection specimens. Median Cox-2 gene expression did not differ significantly among the esophageal control groups but was elevated 5-fold in BE, 8-fold in dysplasia and 16-fold in EAC compared to normal esophageal controls with no erosive GERD. Erosive GERD tissue had slightly higher median Cox-2 expression but Cox-2 expression in normal antrum was much higher than that in a normal esophagus, close to that of dysplasia. In contrast to that of Cox-2, Cox-1 expression was significantly decreased in all neoplastic tissues compared to normal controls. Cox-1 and Cox-2 expression varied over a wide range in the neoplastic tissues but over a relatively narrow range in the esophageal normal tissues. The occurrence of substantial alterations in Cox-1 and Cox-2 expression at the BE stage indicates that these are early events in the development of EAC. These results confirm the important role of Cox-2 amplification in the pathogenesis of esophageal adenocarcinoma, but the unexpected down-regulation of Cox-1 raises questions about its role in carcinogenesis. [Copyright &y& Elsevier]

The Barrett''s multistage process is characterized histopathologically by progression from Barrett''s intestinal metaplasia to Barrett''s esophagus with dysplasia and ultimately adenocarcinoma. Understanding of the molecular alterations in this multistage process may contribute to improved diagnosis and treatment. Retinoid X receptors (RXR) play an important role in regulating the morphogenesis, development, growth, and differentiation of cells. Alterations in RXR expression have been observed in a variety of solid tumors; however, the role in Barrett''s esophagus disease has yet to be determined. The aim of this study was to assess the prevalence and timing of RXR messenger RNA expression in the Barrett''s metaplasia-dysplasia-adenocarcinoma sequence and to investigate its role in the development and progression of this disease. We analyzed the mRNA expression of all three RXR subtypes (RXR-alpha, RXR-beta, and RXR-gamma) by using a quantitative real-time reverse transcription–polymerase chain reaction method in 108 specimens from 19 patients with Barrett''s esophagus without carcinoma (BE group), 20 patients with Barrett''s-associated adenocarcinoma (EA group), and a control group of 10 patients without evidence of gastroesophageal reflux disease (CG). RXR-α mRNA expression was significantly decreased (P < 0.001; Kruskal-Wallis test), and RXR-γ was significantly increased (P < 0.001) at higher stages in Barrett''s esophagus. RXR-β expression was highest in Barrett''s tissues and was significantly increased compared to normal squamous tissues (P = 0.01; Wilcoxon test) and adenocarinoma tissues (P = 0.018, Mann-Whitney test). RXR-α and RXR-β mRNA expression was significantly associated in normal squamous esophagus tissues (r2 = 0.49; P < 0.001; Spearman test), Barrett''s tissues (r2 = 0.63; P < 0.001), and adenocarcinoma tissues (r2 = 0.68; P = 0.001). There were significant differences in RXR-α (P = 0.011) and RXR-β (P = 0.005) mRNA expression in histopathologically normal squamous esophagus tissues in patients with cancer and the control group without evidence of gastroesophageal reflux disease. These findings suggest that alterations in the mRNA expression of all three RXR subtypes are frequent events in the development and progression of Barrett''s esophagus and associated adenocarcinoma, that RXR mRNA expression levels may be useful biomarkers for this disease, and that a widespread “field-effect” is present in the normal esophagus of patients with esophageal adenocarcinoma. [Copyright &y& Elsevier]

The glutathione S–transferases (GSTs) are a family of enzymes that play an important role in the prevention of cancer by detoxifying numerous potentially carcinogenic compounds. GSTs conjugate reduced glutathione to a variety of electrophilic and hydrophobic compounds, converting them into more soluble, more easily excretable compounds. Decreased glutathione S–transferase-pi (GSTPI) enzyme activity has been reported in Barrett''s esophagus, and an inverse correlation was demonstrated between GST enzyme activity and tumor incidence in the gastrointestinal tract, but the role of GSTPI messengerRNA (mRNA) expression in Barrett''s esophagus and associated adenocarcinomas is uncertain. The purpose of this study was to investigate the role of GSTPI mRNA and protein expression in the development and progression of the Barrett''s metaplasia-dysplasia-adenocarcinoma sequence, and to investigate the potential of GSTPI quantitation as a biomarker in the clinical management of this disease. GSTPI mRNA expression levels, in relation to the housekeeping gene β-actin, were analyzed using a quantitative real-time reverse transcription–polymerase chain reaction method (TaqMan) in 111 specimens from 19 patients with Barrett''s esophagus without carcinoma (BE group), 21 patients with Barrett''s-associated adenocarcinoma (EA group), and a control group of 10 patients without evidence of Barrett''s esophagus or chronic gastroesophageal reflux disease. GSTPI mRNA expression was detectable in all 111 samples investigated. Analyzed according to histopathologic group, the median GSTPI mRNA expression was highest in normal squamous esophagus epithelium, intermediate in Barrett''s esophagus, and lowest in adenocarcinoma tissues (P < 0.001). The median GSTPI expression was significantly decreased in Barrett''s esophagus tissues compared to matching normal squamous esophagus from either the BE group (P = 0.001) or the EA group (P = 0.023). GSTPI expression levels in adenocarcinoma tissues were decreased compared to matching normal esophagus tissues from the patients with adenocarcinoma (P = 0.011). Furthermore, GSTPI mRNA expression values were significantly different between metaplastic, dysplastic, and adenocarcinoma tissues (P = 0.026). GSTPI expression levels were also significantly lower in histologically normal squamous esophagus tissues from patients with cancer (EA group) compared to both normal esophagus tissues from patients without cancer (BE group; P = 0.007) and normal esophagus tissues from the control group with no esophageal abnormality (P = 0.002). GSTPI protein expression was generally highest in the basal layer of normal squamous esophagus epithelium and lowest in adenocarcinoma cells, with Barrett''s cells showing intermediate staining intensity. Our results show that downregulation of GSTPI expression is an early event in the development of Barrett''s esophagus and esophageal adenocarcinoma. Loss of GSTPI expression may have an important role in the development and progression of this disease. ( J Gastrointest Surg 2002;6:359–367.) [ABSTRACT FROM AUTHOR]