Antiproliferative Effect of Phenylbutyrate in AsPC-1 Pancreatic Cancer Cell Line

PURPOSE: Phenylbutyrate is an effective redifferentiating agent in several human cancers. Recently phenylbutyrate has been reported to inhibit histone deacetylase activity. We investigated the effects of sodium 4-henylbutyrate (Na-4-PB) on cell proliferation in a human pancreatic cancer cell line. METHODS: A human pancreatic cancer cell line, Aspc-1 was purchased from Korean Cell Line Bank. Antiproliferative effects of sodium 4-phenylbutyrate were measured by MTT assay and their mechanisms were evaluated by apoptosis assay and cell cycle analysis. RESULTS: After 3 days of treatment with Na-4-PB at the concentration of 2.5, 5, 7.5, and 10 mM, relative growth inhibition compared to control was 21.3+/-8.3% (mean+/-SD), 37.8+/-2.3%, 46.7+/-0.5%, and 56.7+/-1.7% respectively (p < 0.05). Antiproliferative effect of Na-4-PB was also time-dependent. Combination treatment with Na-4-PB and troglitazone, a PPARg agonist, increased antiproliferative effects but was not synergistic. After 48 hour treatment with Na-4-PB, early apoptotic cell population in control, 2.5, and 5 mM of Na-4-PB was 29.6%, 44.2%, and 65.9%, respectively. After 24 hour treatment with Na-4- PB, G0/G1 phase population in control, 2.5, and 5 mM of Na-4-PB was 55.0%, 67.4%, and 65.8%, respectively. CONCLUSION: Na-4-PB inhibited pancreatic cancer cell proliferation by inducing apoptosis and cell cycle arrest at G0/G1 phase in time- and dose-dependent manner. Combination treatment with Na-4-PB and other chemotherapeutic agents such as troglitazone, a PPARg agonist, can enhance antiproliferative effects. Na-4-PB might be a promising potential therapeutic agent for patients with pancreatic cancer.

Antiproliferative Effect of NS-398, a Cyclooxygenase-2 Inhibitor, in Pancreatic Cancer Cell Lines

PURPOSE: Selective cyclooxygenase (COX)-2 inhibitors have been reported to inhibit cancer cell proliferation. We investigated the effects of NS-398, a selective COX-2 inhibitor, on cell proliferation in human pancreatic cancer cell lines. METHODS: Human pancreatic cancer cell lines, Aspc-1, Capan-1, and Capan-2 were used. We used western blot and/or RT-PCR to evaluate COX-2 and vascular endothelial growth factor expression. Antiproliferative effects were measured by MTT assay, apoptosis assay and cell cycle analysis. Epidermal growth factor (EGF) and troglitazone were used for combined treatment. RESULTS: COX-2 was relatively overexpressed in Capan-1 and Capan- 2, but minimal in Aspc-1 cell line. COX-2 mRNA expression was upregulated by 50 microM of NS-398 in Aspc-1 cell line but was downregulated at 100 microM in all cell lines. Treatment with NS-398 increased cell population of G0/G1 phase and also induced early apoptotic changes in a dose-dependent manner in all three cell lines. Combined treatment with EGF or troglitazone did not seem to affect antiproliferative effects of NS-398. All three cell lines expressed vascular endothelial growth factor constitutively and its expression was downregulated by treatment with NS-398. Pretreatment with NS-398 prior to radiation exposure increased radiosensitivity in Capan-2 cells. CONCLUSION: COX-2 expression was variable in pancreatic cancer cell lines. NS-398 inhibited pancreatic cancer cell proliferation by inducing apoptosis and cell cycle arrest in a dose-dependent manner. Treatment with NS-398 also inhibited expression of VEGF and enhanced radiosensitivity in pancreatic cancer cell lines. COX-2 inhibitors might be promising potential therapeutic agents for patients with pancreatic cancer.

Cyclooxygenase-2 Expression in Periampullary Cancer

PURPOSE: Cyclooxygenase-2 (COX-2) expression is up-regulated in several types of human cancers and it has been suggested that COX-2 is closely inked to carcinogenesis. The objectives of this study were to investigate COX-2 expression in periampullary cancer and to evaluate the association of the clinicopathological factors with its expression. METHODS: Thirty specimens which were resected from patients with periampullary cancers (13 pancreatic adenocarcinomas, 8 common bile duct cancers, 9 ampulla of vater cancers) were investigated by immunohistochemical staining using Anti COX-2 monoclonal Ab. The 30 specimens were divided into stain-positive and stain-negative groups. The correlation between COX-2 expression and the various clinicopathological factors including the tumor size, nodal metastasis, differentiation, perineural and vascular invasion, were studied. RESULTS: COX-2 was expressed in 69% of pancreatic adenocarcinomas, 100% of common bile duct cancers and 78% of ampulla of vater cancers. However there was no significant correlation between COX-2 expression and the clinicopathological factors. CONCLUSION: COX-2 is highly expressed in periampullary cancer. Even though there was no correlation with the clinicopathological factors, the utility of the COX-2 inhibitors in preventing or treating periampullary cancer remains undetermined but warronts further investigation.

Expression of Inducible Nitric Oxide Synthase (iNOS) in the Epithelial Cells of the Distended Bile Duct by the Bile Ductal Ligation

PURPOSE: Recent studies in an obstructive jaundice rat model showed that the bile duct epithelium is also very important in the bile duct dilatation besides the increased luminal pressure. This study evaluated the role of iNOS in the bile duct epithelium in a rat obstructive jaundice model. METHODS: Bile duct ligations were performed in male Sprague-Dawley rats. The bile ducts were harvested on seven consecutive days. Immunohistochemical staining in the bile duct was performed using anti-iNOS polyclonal antibodies. Aminoguanidine (an iNOS antagonist) was injected intraperitoneally after bile duct ligation (0, 100, and 200 mg/kg/day, n=6 in each group). One week after surgery, the diameter of bile duct was measured and bile was collected for NO analysis by 280NOA (Silvers). RESULTS: The iNOS expression level was increased in the dilated ductal epithelium after the bile duct ligation but not in the normal epithelium. Aminoguanidine decreased the mean diameter of the bile duct after the bile duct ligation: 11/-2.3 mm in the duct ligation only group; 7.5+/-0.75 mm in the 100 mg/kg/day aminoguanidine; 6+/-0.82 mm in the 200 mg/kg/day of aminoguanidine group (mean+/-SE, P<0.05). The NO concentration in the bile was decreased by aminoguanidine: 16+/-4.2 mM in the sham operation group; 40+/-4.5 mM in duct ligation only group; 34+/-6.4 mM in the 100 mg/kg/day of aminoguanidine group; 11+/-1.2 mM in the 200 mg/kg/day of aminoguanidine group (mean+/-SE). CONCLUSION: Bile duct ligation induced iNOS expression in the dilated bile duct epithelium and the iNOS antagonist partially inhibited bile duct dilatation. iNOS induction in the epithelium is partly responsible for the dilatation of the bile duct after duct ligation.

Antiproliferative Effect of NS-398, a Cyclooxygenase- 2 Inhibitor in TPC-1 Thyroid Cancer Cell Line / 대한내분비외과학회지

PURPOSE: Cyclooxygenase (COX) enzymes catalyze the ratelimiting step in arachidonate metabolism. COX-1 is expressed constitutively in many cell types. However COX-2 is an inducible enzyme responsible for prostaglandin production at site of inflammation. Recently, there has been increasing evidence that COX-2 involves in development and progression of human tumors. The aim of the present investigation is to evaluate the antiproliferative effect of NS-398, a selective COX-2 inhibitor, and its mechanism in a papillary thyroid cancer cell line, TPC-1. METHODS: We used TPC-1 cell line, NS-398 and EGF. COX-2 expression was detected by RT-PCR and western blot. We used MTT assay to evaluate antiproliferative effect of NS- 398. The mechanisms of growth inhibition were evaluated by apoptosis assay and cell cycle analysis using flow cytometry. RESULTS: COX-2 expression was identified by both RT-PCR and western blot in TPC-1 cells and it was upregulated by serum, EGF (10 ng/ml), and NS-398 (50 mM). NS-398 induced a dose-dependent inhibition of cell proliferation but did not increases apoptotic cell population significantly in the TPC-1 cell line. EGF treatment (10 ng/ml) for 72 hours did not seem to change the antiproliferative effect of NS-398. The proportion of G0/G1 cell cycle was increased by 10% compared with control after 36 hours of treatment with NS-398. CONCLUSION: TPC-1 cells expressed COX-2 constitutively and its expression was upregulated by serum, EGF, and NS-398. The selective COX-2 inhibitor, NS-398 inhibited cell proliferation in TPC-1 cell line rather by cell cycle arrest at G₀/G₁ phase than by inducing apoptosis.

The Effects of Paclitaxel-Coated Nylon Thread on the Proliferative Cholangitis in a Rat / 한국간담췌외과학회지

BACKGROUND/AIMS: Local drug delivery to the bile duct may be effective to prevent proliferative cholangitis (PC) through capability of high dose administeration with minimal systemic side effects. Paclitaxel is an anticancer drug whose side effect on the stabilization of microtubule leads to cell death. The aims of this study were to establish the proliferative cholangitis model in rat, mimicking biliary stricture in human, and to test whether paclitaxel-coated nylon thread prevents biliary stricture in a PC model of rat. METHODS: PC was induced by introducing a fine nylon thread into the bile duct in a rat from 1 week to 4 week. To evaluate the effects of paclitaxel as a locally-delivered anti-proliferative drug, dog gallbladder epithelial cells were exposed to sequential concentrations of paclitaxel (0.1microM, 1microM, 10microM, and 100microM) for 20 min. in vitro, and inhibition of proliferation was measured by (3)H-thymidine uptake assay. Paclitaxel- coated 5-0 nylon threads (1.8+/-0.5 ug/3 cm thread, measured by HPLC) were made by immersion of ethanolic paclitaxel (50 mg/ml) and evaporation of the solvent. Nylon threads were inserted into the bile duct of male Sprague-Dawley rats weighing 200~50 g. Paclitaxel (n=15) and control (n=15) groups were divided with or without paclitaxel-coating procedure. The paclitaxel effects were assessed by histomorphological examination one week after thread implantation. RESULTS: The decrease of (3)H-thymidine uptake was observed at 100microM of paclitaxel exposured for 20 minutes in the presence of epidermal growth factor (50 nM/ml) than control. PC model characterized by epithelial-glandular proliferation and fibrous thickening of the bile duct wall through 1~ week. This model was established at 1 week. The effect of paclitaxel-coated nylon thread into the bile duct were evaluated after 1 week. In paclitaxel treated the luminal area, luminal length and the ratio of lumen to bile duct cross sectional area increased by 276% (p=0.044), 87% (p=0.012) and by 330% (p=0.000), respectively, versus control. The total wall area, epithelial-glandular area, and stromal area were similar between paclitaxel treated group and control (p>0.05). The bile duct wall thickness of paclitaxel group decreased by 33% (p=0.011, 273 (90)microM vs 410 (95)microM, paclitaxel vs control). CONCLUSION: Paclitaxel-coated nylon thread into bile duct was effective for the suppression of luminal stenosis, and may offer a therapeutic option for biliary stricture and biliary stricture associated disease.