Orlistat Reduces Proliferation and Enhances Apoptosis in Human Pancreatic Cancer Cells (PANC-1).

BACKGROUND/AIM: Pancreatic cancer is a disease with very poor prognosis, and none of currently available pharmacotherapies have proven to be efficient in this indication. The aim of this study was to analyze the expression of fatty acid synthase (FASN) gene as a potential therapeutic target in proliferating human pancreatic cancer cells (PANC-1), and verify if orlistat, originally developed as an anti-obesity drug, inhibits PANC-1 proliferation. MATERIALS AND METHODS: The effects of orlistat on gene expression, lipogenesis, proliferation and apoptosis was studied in PANC-1 cell culture. RESULTS: Expression of FASN increased during proliferation of PANC-1. Inhibition of FASN by orlistat resulted in a significant reduction of PANC-1 proliferation and enhanced apoptosis of these cells. CONCLUSION: This study showed, to our knowledge for the first time, that orlistat exhibits significant antitumor activity against PANC-1 cells. This implies that orlistat analogs with good oral bioavailability may find application in pharmacotherapy of pancreatic cancer.

Gene Expression, DNA Methylation and Prognostic Significance of DNA Repair Genes in Human Bladder Cancer.

BACKGROUND/AIMS: This study investigated the gene expression and DNA methylation of selected DNA repair genes (MBD4, TDG, MLH1, MLH3) and DNMT1 in human bladder cancer in the context of pathophysiological and prognostic significance. METHODS: To determine the relationship between the gene expression pattern, global methylation and promoter methylation status, we performed real-time PCR to quantify the mRNA of selected genes in 50 samples of bladder cancer and adjacent non-cancerous tissue. The methylation status was analyzed by methylation-specific polymerase chain reaction (MSP) or digestion of genomic DNA with a methylation-sensitive restriction enzyme and PCR with gene-specific primers (MSRE-PCR). The global DNA methylation level was measured using the antibody-based 5-mC detection method. RESULTS: The relative levels of mRNA for MBD4, MLH3, and MLH1 were decreased in 28% (14/50), 34% (17/50) and 36% (18/50) of tumor samples, respectively. The MBD4 mRNA expression was decreased in 46% of non-muscle invasive tumors (Ta/T1) compared with 11% found in muscle invasive tumors (T2-T4) (P<0.003). Analysis of mRNA expression for TDG did not show any significant differences between Ta/T1 and T2-T4 tumors. The frequency of increased DNMT1 mRNA expression was higher in T2-T4 (52%) comparing to Ta/T1 (16%). The overall methylation rates in tumor tissue were 18% for MBD4, 25% for MLH1 and there was no evidence of MLH3 promoter methylation. High grade tumors had significantly lower levels of global DNA methylation (P=0.04). There was a significant association between shorter survival and increased expression of DNMT1 mRNA (P=0.002), decreased expression of MLH1 mRNA (P=0.032) and the presence of MLH1 promoter methylation (P=0.006). CONCLUSION: This study highlights the importance of DNA repair pathways and provides the first evidence of the role of MBD4 and MLH3 in bladder cancer. In addition, our findings suggest that DNMT1 mRNA and MLH1 mRNA expression, as well as the status of MLH1 promoter methylation, are attractive prognostic markers in this pathology.

Chemerin gene expression is regulated by food restriction and food restriction-refeeding in rat adipose tissue but not in liver.

Chemerin is an adipokine that regulates adipocyte development and metabolism as well as inflammatory and immune function of some cells. Although chemerin may be linked to obesity and related diseases, little is known about the nutritional regulation of chemerin gene expression. We investigated the effect of prolonged food restriction, a common approach in treating obesity and related diseases, and prolonged food restriction-refeeding on chemerin gene expression in rat white adipose tissue and liver. The prolonged food restriction was accompanied by an approximately 2-fold decrease in chemerin mRNA level in rat white adipose tissue. Upon refeeding, an increase (approximately 8-fold as compared to rats maintained on restricted diet and 4-fold as compared to control) in chemerin mRNA level in white adipose tissue was found. Surprisingly, no effect of food restriction and food restriction-refeeding on chemerin mRNA level in the liver was found. Chemerin mRNA level in adipose tissue was positively correlated with serum insulin concentration. Moreover insulin increased significantly chemerin gene expression in primary rat adipocytes. The changes in chemerin mRNA level in adipose tissue and serum chemerin concentrations were associated with changes in serum leptin and free fatty acid concentrations. Collectively, the data presented here indicate that chemerin gene expression is regulated by nutritional status in rat adipose tissue but not in liver. It seems that insulin plays important role in stimulation of chemerin gene expression in adipose tissue. However, changes in serum leptin and free fatty acids concentrations after food restriction-refeeding suggest that the role of these factors in the regulation of chemerin gene expression in adipose tissue cannot be excluded. Lack of the effect of food restriction and food restriction-refeeding on liver chemerin gene expression suggests that adipose tissue is the dietary modifiable source of serum chemerin concentration.

Up-regulation of rat adipose tissue adiponectin gene expression by long-term but not by short-term food restriction.

Beneficial effects of food restriction as well as elevated circulating adiponectin concentration on the cardiovascular system, lipid metabolism and non-insulin dependent diabetes have been reported. The present article indicates that the reduction in rat body and adipose tissue weight after long-term (1 month) food restriction (either 75% or 50% of ad libitum food intake) was accompanied by the increase in serum adiponectin concentration. The increase in serum adiponectin concentration is the result of the significant increase in white adipose tissue adiponectin gene expression. In contrast to long-term food restriction, short-term (3 days) food restriction (either 75% or 50% of ad libitum food intake) did not influence body and fat mass as well as serum adiponectin concentration and white adipose tissue adiponectin mRNA level. The obtained results suggest that long-term food restriction is necessary to increase in serum adiponectin concentration that could be beneficial, to prevent some disorders associated with low serum adiponectin concentration like obesity and obesity-related diseases.

Clinical implications of p53 mutation analysis in bladder cancer tissue and urine sediment by functional assay in yeast.

In the present study we correlate the p53 gene mutations in tumour tissue with urine sediment using a functional assay in yeast, and relate the p53 status to the outcome in a group of patients with transitional cell carcinoma of the bladder. The p53 mutations were found in three of 30 (10%) Ta/T1 tumour tissue samples and in two of 20 (10%) corresponding urine sediments. In the stage T2-T4 tumour p53 mutations were found in tumour tissues and urine sediments in 13 of 31 (42%) and in seven of 18 (39%) samples, respectively. In 80% (8/10) of cases, the p53 mutations found in tumour tissue were re-detected in urine sediment. Median follow-up was at 20 months. Disease recurred in 18 of the 61 patients (30%) with a median time of 5 months. In Ta/T1 tumours the frequency of recurrence was 37% (11/30) compared with 23% (7/31) of T2-T4 tumours. The 3-year overall survival (OS) was 82% (50/61). The p53 status was significantly associated with stage (P = 0.0077, two-sided Fisher's exact test), grade (P < 0.001) and lymph node involvement (P = 0.027). There was an association between the p53 mutations and shorter OS (P = 0.033; log-rank test); however in a multivariate analysis adjusted for stage, grade, lymph node status and age the p53 mutation was not an independent predictor of survival. There was no correlation of the p53 status with decreased disease-free survival (P = 0.8; log-rank test). The data presented indicate that the yeast functional assay is a useful method for p53 gene mutation analysis in tumour tissue and p53 mutation can be re-detected in urine sediment, but further validation of the assay in non-invasive screening for p53 mutations is needed.