advanced gastric cancer, but it is lower in non-dysplastic tissue adjacent to the tumour in comparison with that of chronic H. priori gastritis 17989 Ahernative Roles of c-Jnn Pathway in Cell Proliferation and Gemcitabine- Induced Apoptosis of Colon Cancer Cell Reiko Suto, Kazunari Tominaga, Razuhide Higuchi, Eiji Sasaki, Toshio Watanabe, Yasuhim F@wa*a, Nobuhide Oshitani, Takaypki Matsumoto, Tetsuo Arakawa It is reported that the induction of apoptosis by gemcitabine (GEM) relates c-jun pathway m some cancer cell lines But the interaction between c-jun pathway and apoptosis induced by GEM is not tidly understood in colon cancer. We investigated the role of c-jun in cell kineucs incluchng both cell proliferation and apoptosis induced by GEM using adenovirus dominant negative nmtant of c-jun. METHOD: AP- 1 DNA binding activity of colon adenocar- cinoma cells (HT29) was determined by electrophoreac moNlity shift assay" (EMSA). Cell proliferation *wasevaluated by PHI thymidine incorporatton. Susceptibility to GEM on HT29 cells was evaluated by MTS assa F We used adenovirus domimant negative mutant of c-jun (Ad-DN-c-hin) to inhibit endogenous AP-1 Distribution of cell cycle and apoptosis were evaluated by flow cytometry, RESULFS: DNA binding activity of AP-1 was increased by serum stimulation (1%) in a time dependent manner. Infection of HT-29 with Ad-DN-c- Jun decreased serum-induced protitm~tinn in a dose dependent manner. GEM significantly" inhibited these serumdnduced cell proliferation (IC50:IOnM). Flow cytometry analysis showed that GBM significantly induced apoptosis by 72hr (488% of the cell population being apoptotic) compared with control (6%) (p < 0.01) interestingly, however, inhibiting endogenous AP-1 with AD-DN-c-Jun decreased GEM-induced apoptosis by 30.5% (p < 001) CONCIYUSION:Our results demonstrated that in HT-29 ceils, c-jun pathway played au important role in the apoptosis induced by GEM, Whereas c-jun pathway was also critical for cell prolit)ration. Tbese findings suggest that the role of C-lUn pathway can be alternative in accordance with the external stimulations T990 Polypbenolic Compounds fl'om the Fruit Gareinia Strongly Inhibit the Growth of Human Colon Cancer Cells by G1/S Arrest and Apoptosis Petr l~otiva, Ed Kennelb/Scott Baggett, jnn Ma, Peter R Holt, 1 B. Weinstein, i. B. Weinstein Background: The role of dietary ~?uits and vegetables in colorectal cancer prevention is controversial in part this may stem from the fact that potentially active compounds derived from plant foods trove not been identified and their content may vary in dietary sources. Aim: The purpose ot this study is to identify novel plant polyphenolic compounds from the kuit Garcima xanthochymus that might be effective for colorectaI cancer prevention and/or treatment Methods: Nine fractioI~ of the ethyl-acetate extract f~:om Gascinia which is nch m polyqpbenols were tested for growth inhlhition on the human colon cancer cell lines HCT-11B and SW-480 by a modified MTT assay. The most active fraction denoted as A-C was assessed tot ability to induce apoptosis and cell cycle arrest by flow cytometry. Fraction AC was also analyzed by high perfbrmance liquid chromatography (HPLC) to identify"pure compounds responsible for colon cancer cell growth inhibition. Results: Fraction A-C of Gascinia xanthochymns displayed IC50's of approximately 6-8 mcg/ml with both ceIllines. In both ttCT- 11B and SW-480 cells fraction A~C induced dose-dependent apoptosis and G1/$ growth arrest. In HCT-116 ceils, apoptosis increased from 1% for controls to 10% for AC concentration of 10 mcg/ml and to 18% tor A-C concentration of 20 mcg/ml. The respective values in 5W-480 ceils veere 1.3% for controls, 6% {br A~C concentration of 10 mcg/mI and 11% lhr A-C concentration of 20 mcg./mL HPLC armlysis revealed that fraction AC contains two major compounds, xanthochymol and guuiferone E, which will be analyzed in k~ture studies Conclusion: tin extract from the fruit Garcinia rich in xanthochymol and guttiterone E strongly inhibits the growth of human colon cancer cells by inducing G1/5 arrest and apoptosis Plant poIyphenols may include several highly active compounds that could be usetial in both colon cancer cherooprevention and therapy, Studies are in progress to determine the erecise molecular targets of these Garcinia compounds in colon cancer cells. T991 Gastric Cancer Is Associated With Reduced Apoptosis In Non-Lesional Gastric Mucosa Bather Than Increased Mucosal Proliferation Khay-Guan Yeoh Chun-l-ao Wai, Ming Teh, Kong-Bing Tan, Bee-l.eng Seer, Andrea Ralmlkova Background and Aim The pathogenesis of gastric cancer is incompletely understood. The aim of this study was to compare differences in mucosal apoptosis, proliferation and inflam- mation in non-lesional gastric mucosa in patients with and without gastric cancer, to identify ask tactors tbr further investigation. Patients and Methods Consecutive patients referred to a hospital gastroenterology" clinic for dyspepsia and scheduled for gastroscopy were enrolled n the study after reformed consent was obtained. Biopsies taken from non-lesional gastric antral mucosa were sent for histology and evaluation of apoptosis (by TUNNEL technique), prohteratior~ (Ki67) and inflammation (iNOS). Serum was tested for H pylori (HP) IgG, cag A and vac A antibodies. Results Over a 24-month period, 182 patients were recruited. Mean+ SD age was 53- + 13 years, 109 (6094) were male, and 153 (84%) were Chinese. HP infection was present in 115 (63%) patients, of which 110 (96%) ,,,,'ere cag A positive and 104 (91%) were vac A positive. Fifteen patients (8%) had gastric cancer. Apoptosis index n non-lesional nmcosa was significantly lower in gastric cancer than non-cancer (13.8 -+ 4.7 vs 24.5-+ 1.3, p=0 02), while there were no sigmficant differences in Ki67 (13,1 _+5.0 vs. 22.0• 1.5,p=0 ll) and iNOS (4.5 -+O.3 vs. 4.4+_0A, p=0.69). HP+ patients had higher K~67 indices (236- + 19 vs 17.3• p=0.04) and higher iNOS (4.6-+ 1.2 vs 4.0- + 1.5, p=O,002) but no differences in apoptotic indices (24.5 • 1.5vs 22.2 • 2.3, p = 0.39). At univariate analysis, patients with gastric cancer were older and had a lower level of apoptosis (Table below) At multivariate analysis, both older age and lower level of apoptosis were independently associated with gastric cancer. Conclusion Gastric cancer was associated with older patient age and reduced apoptosis m gastric mucosa, while there were no sigmficant differences m indices of inflammation or proliferation, We hypothesize that impaimrent of apoptosis by genetic or environmental factor(s) may play a critical role in the pathogenesis of gastric cancer. Factors associated with gastric cancer in univariate analysis Factors Patient1 with ga=dr can. Patients without g=~dr can. r n:lS cer, n:167 P Age, y~rsl" 62 4 52 1 0,003 ~.4de 12 (80%) 97 (58%) 0,11 Index of apopto. 13.8 4,7 24,5' 1,3 0.02 si,t Index of prolifeta- 13.0 ~5.0 21,9 15 0.117 lion Index of INOS 4.5,:0,3 4A0,1 0,69 l'sign~nt on multivariate analysis T992 Phenylacetate Induces G1/S Arrest and Apoptosis in Colon Cancer Cells but Antagonizes S-phase Specific Chemotherapy Petr Protiva, Banke Agarwal, Masahito Shimizu, i. B. Weinstein, Peter R. Holt Background: We previously" showed that phenylacetate (PA) inhibits grmvth and induces apoptosis in several colon cancer ceil lines. Whether PA can potentiate the eft)ct of conven- tional chemotherapeutic agents is controversial Aim: The purpose of this study is to test whether PA can potentiate effects of the S-phase specific chemotherapeutic agent 5 fluoroura- cyl (FU), and the non-specific agent cis platin (PT) on colon cancer ceils We hypothesized that PA induces G1/S arrest and apoptosis in colon cancer cells and that this GI/S arrest antagonizes the eft?ct on apoptosis of FU but not of PT Methods: HCT-116, SW-480 and HT-29 colon cancer cells were incubated fbr 36 h with PT I0 mcg/ml or FU 20 mcg/mI with or without PA 25 mM. Cell viability was assessed by MTT assay', and apoptosis and cell cycle analyses were evaluated by flow cytometry. Results: PA induced GI/S arrest and apoptosis in all 3 cell lines. PA slightly enhanced the df~:cts of PT on cell viability" and apoptosis in all cells. In contrast, the addition of PA to FU treated cells resulted in a dramatic amagonistic effect on cell viability in HT-29 but had no effect in HCT-I16 or SW-480 cells Furthermore, addition of PA decreased FU-induced apoptosis in alI three cell fines and this eft~:ct was most prominent in HT-29 cells. Conclusion: PA induces GI/S arrest and apoptosis in human colon cancer cells. PA enhances the effect of PT but antagonizes tbe effect of FU on apoptosis Because PA prevents cells trom entering the S phase PA probably should not be combined with chemotherapy agents that act only m the S phase, such as FU T993 15-Lox-1 Inhibits P21 (CiptNVAF1) Expression by Enhancing Mek-Erkl/2 Signaling in Colon Carcinoma Cells Masahiro Yoshinaga, Rayn'tond N Dubois Background & Aims: Previous studies have shown that 15dipoxygenase-1 (15-LOX-1) is expressed in the normal colon tissues and Colonic tumors. There is some controversy concerning its precise role in colorectal carcinogenesis, which needs fiarther clarification Others have shown that sodium butyrate (NaBT) induces 15-LOX-1 expression and G1 celI cycle arrest in colon cancer cells. The aim of this study was to evaluate the mvolvement of 15-LOX-I in cell cycle control and proliferation of colon cancer ceils in order to further understand its role in carcinogenesis. Methods: The expression of cell cycle control molecules was evaluated in Caco~2 cells treated with NaBT and/or Nordihydroguaiamtic acid (NDGA), a LOX inhiblton The dfect of 13-S-hydroxyoctadecadienoic acid (HODE), a product of 15- LOX-1, on cell cycle progression was evaluated in Caco-2 ceils treated with NaBT The effect of transiently transfecting HCT 116 cells with 15-LOX-1 was also evaluated. Results: 13-S-HODE levels (measured by EL1SA) were elevated in Caco-2 cells treated with NaBT and 15-kOX-transtected HCT 116 cells. NDGA treatment enhanced p21 (Cip/WAF i) expression in Caco-2 cells pre-treated with NaBT Low concentrations of 13-S-HODE decreased p21 expression and partially reversed the G1 arrest. 15-LOX-1 transtected HCT 116 cells had decreased p21 expression and dramatically increased cell growth. 13-HODE treatment restored NaBT-induced inhibition of ERK 1/2 phosphorylation Treatment with PD98059 enhanced p21 expression levels in NaBT-treated Caco-2 ceils Conclusion: Our results indicate that 15-LOX-1 inhibits p21 expression by enhancing ERK 1/2 signaling in colorectal cammoma cells Our in vitro experimental results support the hypothesis that 15- LOX-1 may have a pro-neoplastic effect. T994 5-Fluoruracil and Cisplatin Indnce Nf-Kb, Ap-1 and COX-2 Transcription in Oesophageal Epithelial Cells: Role for PKC and MAPK Signalling Pathways Involvement Mohamed M. M. Abdel-Latif, Dermot Kelleher, John V, Reynolds Background The regulation of gene expression by certain transcription factors, such as NF- KB, AP-1 and COX-2, is fundamental to the phenotype of all cells engaged in intlammatory and immune responses, Since antineoplastic agents have used in different settings for treating of oesophageal cancer, we have examined the effect of 5-fluoruracd and cisplatin on NF- KB, AP-1 and COX-2 activity in the oesophageaI epitheliaI cell lines SKGT-4 and OE33. Methods: NF-KB and AP-1 activ W were assessed by electroplmretic mobility gel shift assay (EMSA) and Western blotting COX-2 acti'dty was assessed by Western blotting. Reki. immunofluorescence staining was also performed, Results: Treatment of OE33 and SKGT- 4 ceils with 5-Fu (1-1000 p.M) and cisplatin (1-10 p.M) reduced NF-KB and AP-1 activity m a dose- and time-dependent manner. 5-Fu and cisplatin also induced RelA tmndocation, A-463 AGA Abstracts