- Email: [email protected]
Sa1967 Lipid Profile and Polyp Size in Subjects With Colon Adenomas
Sa1965 Garcinol Interferes With Oncogenic IL1b-STAT3 and Facilitates Tumor Suppressive KLF11-Sin3A to Down-Regulate AKT1 Expression and Cell Growth in Barrett's Epithelium Anamay N. Sharma, Sarah Kossak, Sonia Chowdhury, Raghav Chandra, Ishtpreet Singh, Anushka Baruah, Cathrine J. DeMars, Prasad G. Iyer, Raul A. Urrutia, Kenneth K. Wang, Kausilia K. Krishnadath, Navtej Buttar BACKGROUND: Chronic reflux injury promotes Barrett's metaplasia and adenocarcinoma, one of the most rapidly increasing, highly lethal cancers in Western countries. We have previously shown that transcriptional up-regulation of AKT1 expression plays a critical role during this neoplasia and is involved in increased cell growth, transformation, colony formation in-vitro and the growth of tumor implants in-vivo. This regulation is an important node on which pro-inflammatory oncogenic IL1b-STAT3 and tumor suppressive KLF11Sin3a pathways compete to regulate carcinogenesis. AIM: Our goal was to investigate the chemopreventive potential of the naturally occurring compound Garcinol, through interference with oncogenic IL1b-STAT3 and facilitation of tumor suppressive KLF11-Sin3a pathways. METHODS AND RESULTS: We treated Barrett's epithelial cells (FLO and SKGT) with Garcinol at baseline and under the conditions of activated IL1b-pSTAT3 or KLF11-Sin3A pathway. We used promoter-reporter luciferase assay, PCR, Western blot and BrDU incorporation. We noted that while IL1b-pSTAT3 increased AKT1 promoter activity and AKT1 expression, KLF11-Sin3A decreased them. A 24-hour treatment of Barrett's epithelial cells with 20uM Garcinol resulted in marked down-regulation of pSTAT3 protein expression and facilitated KLF11-mediated repression of AKT1 promoter, decreased AKT1 promoter activity, as well as decreased AKT1 mRNA expression. These molecular changes had functional consequences as similar treatment with Garcinol resulted in up to 50±1% reduction in cell growth compared to control treated cells (p<0.05). CONCLUSIONS: The interference with oncogenic IL1b-STAT3, facilitation of tumor suppressive KLF11-Sin3A, and down-regulation of AKT1 expression, as well as decreased cell growth in-vitro support the chemopreventive potential of Garcinol. However this agent needs to be tested further in-vivo in pre-clinical and clinical studies.
Sa1963 Low Molecular Weight Procyanidins From Grape Seeds Enhance the Impact of 5-Fluorouracil Chemotherapy on Colon Cancer Cells Ker Y. Cheah, Gordon S. Howarth, Keren A. Bindon, James A. Kennedy, Suzanne Mashtoub, Susan E. Bastian Introduction: Grape seed procyanidins (PC) are flavan-3-ol oligomers and polymers known for their biological activity in the gut. Grape seed PCs have been reported to reduce intestinal injury in rat models of intestinal mucositis [Cheah et al. Cancer Biol Ther (2009)] and ulcerative colitis [Cheah et al. Dig Dis Sci (2013)]. We sought to determine the differential efficacy of purified PC fractions differing in mean degree of polymerization (mDP) on the viability of colon cancer cells (Caco-2) when combined with 5-Fluorouracil (5-FU) chemotherapy. Methods: Six PC fractions (F1-F6) were isolated from Cabernet Sauvignon seeds at two ripeness stages: pre-veraison unripe (immature) and ripe (mature), utilizing step gradient, low-pressure chromatography on a Sephadex LH-20 resin. Fractions were characterized by phloroglucinolysis and gel permeation chromatography (GPC). The antioxidant capacity of the fractions was determined by ferric reducing antioxidant power (FRAP) assay. Fractions were tested on Caco-2 cells, alone and in combination with 5-FU. Cell viability was determined by 3-(4,5-Dimethylthiazol-2yl)-2,5-diphenyl-tetrazolium bromide) (MTT) assay. Statistical significance was assumed at p<0.05. Results: The antioxidant capacity of six fractions was negatively correlated with PC mDP (r2 = -0.81, p<0.05). All isolated fractions significantly reduced Caco-2 cell viability compared to control (p<0.05), although F2 and F3 were the most active fractions (immature F2 = 32%, F3 = 35% and mature F2 = 13% and F3 = 17%; percentage of viable cells remaining) on Caco-2 cells. When combined with 5-FU, immature seed fractions F1-F3 and mature seed fractions F1-F4 enhanced the growth-inhibitory effects of 5-FU by 27-73% and 60-83% (p<0.05; compared to 5-FU control), respectively. Moreover, some fractions were more potent at decreasing viability in Caco-2 cells (p<0.05; immature F2-F3 = 65-68%; mature F2-F4 = 83-87%) compared to 5-FU alone (37%). Conclusions: Procyanidin fractions of mDP 2-6 (immature F1-F3 and mature F1 and F4) exhibited synergistic effects on viability of Caco-2 cells when tested in combination with 5-FU. Concomitant use of grape seed PCs and 5-FU chemotherapy could represent a promising new approach for colon cancer chemoprevention.
Sa1966 Predictors of Right-Sided Polyp Growth in a Veteran Population Cynthia E. Quainoo, Sonia DeAlwis, Ching-Ho Huang, Vidushi Golla, Michael Haynes, Yasemin Aytaman, Albert Ndzengue, Kathlynn Caguiat, Ayse Aytaman, Manuel Martinez, Garrett Lawlor INTRODUCTION The clinical implications of right sided colon polyps is gaining interest though the exact difference of their pathogenesis as compared to left sided colon polyps remains unclear. Thus far, there is suggestion that right sided colon polyps carry increased risks of malignant transformation and this may be attributable to both environmental and host factors. The aim of our study is to better understand the factors which impact the rate of right sided colon polyp growth and pathogenesis. METHODS All patients undergoing two colonoscopies at least 12 months apart between 1996 and 2012 with detection and removal of a polyp on index colonoscopy, were identified using our endoscopic database to determine the incidence of polyps and adenomas. We recorded patient age, interval between colonoscopies, gender, race, BMI, HbA1C (>6.5, <6.4), HCV Ab positivity and statin use (Y/N), folate, Vitamin D use, Aspirin and NSAIDS. Patients were classified according to age (<49, 50-59, 60-69, 70-79, > 80yrs). From the endoscopic data, we recorded polyp histology, location, size and prep score. RESULTS Overall, 1056 patients underwent two colonoscopies at least 12 months apart (median interval 47 months) with removal of polyp on initial examination. Considering all patients, there were more right sided polyps among the white population (56%) compared to the black population (44.4%, P 0.0056). Statins were used by 57.9% of patient population; their use was associated with an increased percentage of right sided polyps (54.6% versus 46%, P 0.03). Aspirin use was found to increase the percentage of right sided polyps relative to left sided (56.5% versus 47.4% P 0.0219. Finally, Vitamin D supplementation was associated with a trend towards increased right sided polyps (OR 1.474, P 0.0716, CI 0.9648-2.25). CONCLUSION The pathogenesis and natural history of right -sided polyps appears to differ compared to left -sided polyps, though this mechanism is unclear. Our data suggest that race, statin use, aspirin, and vitamin D supplementation appear to impact right - sided polyp growth independent of age or median time between colonoscopies(while also impacting left-sided polyp growth). The effect of medication and non-colorectal disease on polyp growth will increasingly be recognized as differential between left and right-sided polyps, owing to their poorly understood, different mechanisms of growth.
Sa1964 Gastric Cancer Chemoprevention by Phenethyl Isothiocyanate-Containing Diet in Chemically - but Not Genetically - Induced Gastric Cancer in Mice Anders Øverby, Chun-Mei Zhao, Timothy C. Wang, Atle Bones, Duan Chen Background/aim: Vegetables containing isothiocyanates (ITCs) such as broccoli, cauliflower and watercress have been linked epidemiologically to decreased risk of gastric cancer. The aim of the present study was to demonstrate the chemopreventive effect of the naturally occurring aromatic phenethyl ITC (PEITC) in vivo in mouse models of gastric cancer and in vitro using human gastric cancer cell lines. Methods: FVB mice treated with N-nitrosoN-methylurea (MNU, orally for 10 weeks) and transgenic INS-GAS mice with spontaneous gastric cancer were employed. Tumorigenesis was assessed by measurement of tumor size using a point counting method. PEITC-containing diet (5 μmol/g PEITC) was given ad libitum to FVB mice that were either treated with MNU simultaneously or immediately after 10-week MNU treatment for 9 months. INS-GAS mice were fed the standard mouse diet with or without PEITC for 11.5 months (starting at preneoplastic stage, i.e., 4-5 weeks of age). For the in vitro study, the gastric cancer cell lines Kato-III and MKN74 were used. Cell proliferation was determined with the MTT-assay, while morphology and cell migration were analysed using a contrast microscope, cell cycle distribution was analysed with flow cytometry, and the effect of PEITC on microtubules was examined in transfected cells by confocal microscopy. Results: The PEITC-containing diet was without effect on the body weight development in all groups of mice. Both PEITC regimens whether it was administered prior or subsequent to MNU treatment yielded a significant reduced tumorigenesis in MNUtreated mice, with chemoprevention pre-MNU feeding being more potent than post-MNU feeding. There was no significant inhibitory effect of PEITC-containing diet on tumorigenesis in INS-GAS mice compared with age-matched INS-GAS mice without PEITC-containing diet. In vitro studies using Kato-III and MKN74 cells showed that PEITC inhibited cell proliferation in a time- and dose-dependent manner in the concentration range of 1-100 μM for 24, 48 and 72 hours, with Kato-III cells being more sensitive to PEITC than MKN74 cells. The reduction in cell survival coincided with changes in cell morphology and reduced cell migration. Furthermore, treatment of PEITC for 24 hours led to an accumulation of cells in G2/M-phase of the cell cycle. Kato-III cells expressing GFP-tagged microtubules showed that PEITC induced disruption of microtubules with the subsequent formation of apoptotic blebs. Conclusions: The present study demonstrates a chemopreventive effect of PEITC-containing diet in a mouse model of chemically-induced gastric cancer. The anticancer effect may involve disruption of microtubules, resulting in cell cycle arrest and apoptosis.
Sa1967 Lipid Profile and Polyp Size in Subjects With Colon Adenomas Sandar Linn, Carmen M. Stanca, Gregory Rozansky, Juan Tejada, Philip Xiao, Sury Anand BACKGROUND: Increased expression of LDL receptor mRNA has been described in colon cancer tissue. Malignant colon tumors are thought to be involved in cholesterol catabolism, and curative surgery would lead to changes in cholesterol levels. We examined the lipid profile of patients with colonic adenomas. METHODS: Adult patients who underwent polypectomy over a one year period were enrolled in this retrospective study. Controls included patients coming to the ambulatory clinic during the same period for routine visits. We recorded demographics, lipid levels, co-morbidities, and statin use in both study and control groups; we documented polyp size in patients undergoing polypectomy. Patients with concomitant colon or other malignancies were excluded. Data were analyzed using SPSS; results are shown as median (range). RESULTS: Ninety-four subjects with colon adenomas and one hundred and thirty controls were enrolled. Fifty-four percent of subjects were women and 88% were African American. The median age was 62 (23-94). There were no significant differences in terms of age, gender, race, BMI and statin use between the adenoma group and the controls. Significantly lower LDL-cholesterol levels were identified in the adenoma group (median 54 vs 114; p<0.001) in the absence of statin use. All the other
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also had 4-fold higher levels of miR-145 (n= 3, p<0.05). We could not demonstrate any EGFRESR2 physical interactions. Surprisingly, our preliminary results showed an upregulation in EGFR signals in ERB041 treated ESR2 transfectants. Summary: In AA with adenomas, reductions in ESR2 expression might contribute to higher colon cancer incidence and worse prognosis. The in vitro studies confirmed that ESR2 suppresses colon cancer cell proliferation. We postulate that the antiproliferative effects of ESR2 involve miR-145 dependent, EGFRindependent pathways. Our study suggests that ESR2 down-regulation might play an important role in colon cancer risk and contribute to racial differences in cancer progression.
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serum lipids had higher levels in the adenoma group compared to controls: total cholesterol (186 vs 146; p<0.001), triglycerides (104 vs 72; p=0.002), and HDL-cholesterol (105 vs 45; p=0.075 did not reach statistical significance). An inverse correlation was found between the total cholesterol level and the polyp size (Spearman's r=-0.343; p = 0.047). No statistically significant differences were identified in the serum lipid levels in the adenoma group after polypectomy. CONCLUSION: Subjects with colon adenomas have significantly lower LDLcholesterol levels, and their total cholesterol level inversely correlates with polyp size. Serum lipid levels do not change after polypectomy. Although the cause-effect relationship between lipid levels and colon cancer is not clearly defined, changes in the lipid profile seem to precede the development of colon cancer. This study offers additional insights in the complex relationship between lipid metabolism and colonic neoplasms. Sa1968 Mesalamine Exhibits a Protective DNA Damage Response in Normal Colon Epithelial Cells Under Oxidative Stress Vineeta Khare, Mario Asboth, Christoph Gasche
Figure: Kaplan-Meier survival for patients with native stage I and downstaged clinical stage III rectal cancer. Adj = adjuvant. CR = complete responder.
Background: Ineffective DNA damage response such as cell cycle arrest and DNA repair contributes to colorectal cancer (CRC). Recently, our lab demonstrated that in colitis and colitis-associated cancer (CAC) oxidative stress causes DNA double strand breaks (DSB). Pharmacological manipulation of DNA damage response could be a potential mechanism for chemoprevention in CAC. Mesalamine (5-ASA) is commonly used as an anti-inflammatory compound in the treatment of UC and improves replication fidelity by activation of a replication checkpoint. Here we examined the effect of 5-ASA on DNA damage and DNA repair in normal colon epithelial cells (and a colorectal cancer cell line as control). Methods: Normal diploid human colon epithelial cells (HCEC-1CT) and HCT116 cells (CRC control) were treated with 5-ASA (5mM; 24h). Proteins implicated in DSB were analyzed by Western blot using various antibodies for DNA damage response proteins in the nuclear enriched cellular fraction. To assess the effect of 5-ASA on oxidative stress, cells were treated with H2O2 (100μM, 30 min) in the presence or absence of 5-ASA (5h pretreatment). Nuclear foci formation by 53BP1 and gamma-H2AX in response to oxidative stress was determined by immunocytochemistry. BrdU incorporation was measured to assess cell proliferation during recovery phase (24h). Results: DNA damage sensor proteins MRE11, γ-H2AX, and 53BP1 were strongly induced upon 5-ASA treatment in HCEC-1CT. Inversely, such proteins were highly expressed in HCT116 and were downregulated upon 5-ASA. In HCEC-1CT, 5-ASA resulted in diffuse nuclear localization of 53BP1 and γ- H2AX. Oxidative stress, however, resulted in redistribution of these proteins into distinct foci indicative of DNA damage. Pretreatment with 5-ASA reduced foci formation under oxidative stress. Compared to untreated HCEC-1CT, cell proliferation was significantly reduced upon H2O2 treatment (3% relative BrdU incorporation), however, 5-ASA pretreatment was protective (45%). HCT116 cells remained proliferative under oxidative stress and 5-ASA pretreatment showed no such effect (80% and 81.5%, respectively). Conclusion: 5-ASA treatment induces DNA damage response in normal colon epithelial cells and exhibits a protective effect under oxidative stress by induction of a double-strand break repair pathway impeding with CAC development. Advanced dysplastic lesions are rather unlikely to benefit from 5-ASA.
Sa1970 CD98 As a Ligand for Colitis-Targeted Delivery of Nanoparticle Bo Xiao, Emilie Viennois, Yuchen Zhang, Saravanan Ayyadurai, Hamed Laroui, Didier Merlin Background and Aims: Treatment strategies for inflammatory bowel disease (IBD) have been constrained by limited therapeutic efficacy and serious adverse effects owing to a lack of ligand for targeted drug delivery to the inflamed colon. Upon inflammation, CD98 expression is highly elevated in colonic epithelial cells and infiltrating immune cells. To investigate whether CD98 can be used as a colitis-targeted delivery ligand, we constructed CD98 Fabbearing quantum dots (QDs)-loaded nanoparticles (Fab'-NPs). Materials and Methods: CD98 Fab' was conjugated to the surfaces of QDs-loaded NPs mediated by bi-functional PEG derivative (MAL-PEG-NHS). Next, we evaluated their physicochemical properties (e.g., hydrodynamic particle size, zeta-potential, emission and absorption properties) in relation to their targeting capacity. We also investigated their targeting capacities to CD98 overexpressed colonic cells using flow cytometry. Finally, we tested their ability to target colitis tissue. Results: The resultant Fab'-NPs had desired particle size (~458 nm) with a narrow size distribution and zeta-potential (approximately +19 mV), low cytotoxicity, and excellent fluorescence properties. Electron microscopy images provided direct evidence for the welldispersed distribution of QDs within spherical Fab'-NPs. Cellular uptake experiments demonstrated that Fab'-NPs were efficiently internalized into Colon-26 and RAW 264.7 cells through the CD98-mediated endocytosis pathway, and showed that the targeting effect of CD98 Fab' markedly increased their cellular uptake efficiency compared with control pegylated QDs-loaded NPs (PEG-NPs). Furthermore, ex vivo studies showed much more effective accumulation of Fab'-NPs in colitis tissue than that of PEG-NPs. Conclusion: In the context of IBD therapy, one of the most relevant features of the drug formulation is that they can targetable deliver therapeutic molecules to specific cells. Our above findings suggest that active colitis-targeted delivery can be accomplished using NPs decorated with CD98 antibody based on the inflammation-dependent over-expression of CD98.
Sa1969 Effects of Neoadjuvant and Adjuvant Therapy for Rectal Cancer on Long Term Survival Mohan K. Mallipeddi, Paul J. Speicher, Asvin M. Ganapathi, Brian R. Englum, John Migaly, Christopher R. Mantyh
Sa1971 High-Resolution Esophageal Impedance Topography (EIT) Parameters for Quantifying Bolus Retention Zhiyue Lin, Frédéric Nicodème, Chen-Yuan Lin, Benjamin Mogni, Laurel Friesen, Peter J. Kahrilas, John E. Pandolfino
Purpose: Pre-operative chemoradiation (C/XRT) for advanced rectal cancers often pathologically downstages tumors and significantly decreases local recurrence. Adjuvant chemotherapy is also recommended for advanced staged rectal cancer to reduce systemic recurrence. However, it is unknown what C/XRT downstaging effect has on overall long term survival. Also it is unclear what effect omission of adjuvant chemotherapy has on overall survival. Methods: Patients with clinical stage III rectal cancer who underwent pre-op C/XRT in 2006 were identified from the National Cancer Database (NCDB), which captures cancer site specific data from over 1500 centers across the US. Overlapping survival and chemotherapy data only were available for 2006. Patients who underwent resection with curative intent were included for analysis. Kaplan-Meier survival estimates were used to compare survival by pathological stage with native stage I patients and complete responders as references. Cox proportional hazards modeling was used to assess the impact of demographic, clinical, and pathological factors on survival as well as the interaction between adjuvant therapy and degree of neoadjuvant downstaging. Results: There were 818 patients with clinical stage III disease who received neoadjuvant therapy. Pathology revealed 403 ypStageIII patients, 193 ypStageII patients, and 186 ypStageI patients, and 36 complete responders (CRs). Overall, 231 patients received adjuvant chemotherapy, 25% of ->ypI patients vs 35% of ->ypIII patients (p<0.001). Five-year overall survival for all cIII->ypI patients compares favorably to native stage I disease at 83.3% versus 77.7%, respectively (Hazard Ratio: 0.98, CI: 0.63, p=0.909) (Figure). The subset of the cIII->ypI patients who received adjuvant therapy had significantly better unadjusted survival than native stage I patients (p=0.007). For ->CR, ->ypI, ->ypII, and ->ypIII patients receiving adjuvant therapy, 5-year survival was 94.4%, 92.8%, 90.5%, and 63.1%, respectively. Among patients who were downstaged to ypI, there was a trend towards a survival advantage with the addition of adjuvant chemotherapy (HR: 0.25, CI: 0.05-1.12, p=0.07), and adjuvant therapy provided a significantly larger survival benefit with increasing degree of downstaging (p=0.028). Yet, in the NCDB less than 50% of downstaged cStage III patients through 2011 were receiving adjuvant therapy. Conclusions: Patients with stage III rectal cancer that were downstaged to stage I had equivalent longterm survival compared to native stage I patients. Progressive downstaging by neoadjuvant C/XRT significantly improved survival after adjuvant C/XRT. Unfortunately, a minority of rectal cancer patients received adjuvant therapy which likely reduced overall survival. Future work should focus on improving utilization of adjuvant therapy in these patients.
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Background and Aim: The multichannel intraluminal impedance (MII) technique allows one to evaluate bolus transit without radiation. However, the line tracing analysis mode is limited by low spatial resolution and the higher resolution impedance color contour plot view (CCPV) mode does not provide quantitative analysis of bolus volume on MII data. This study aimed to develop a new method for quantifying bolus volume clearance using highresolution esophageal impedance topography (EIT) technique. Methods: The ability of highresolution EIT parameters to detect bolus volume clearance was validated and tested with concurrent fluoroscopic imaging and high-resolution impedance manometry (HRIM) studies (barium bolus) in 10 healthy subjects (4 males, mean age 28 years, range 21-47). HRIM data from each subject were analyzed using a MATLAB program customized for calculating EIT parameters for each swallow within a region of interest from lower limit of UES to upper limit of EGJ in distance and from onset of swallowing to 12 seconds later or until completion of peristalsis (red dashed rectangle in Figure). The impedance volume defined as the sum of all pixel volumes (i.e., impedance value less than 50% drop from the mean baseline value to the mean nadir impedance (white curves) multiplied by time interval and by spatial interval) were derived in area 1 prior to the contraction wave-front at 20 mmHg isobaric contour (black solid curves) as Z1 and in area 2 after the contraction as Z2, and presented as a ratio of Z2 area/Z1 area. Results: A total of 40 thin barium swallows (4 per subject; 2 supine and 2 upright) were analyzed from the 10 subjects with simultaneous fluoroscopic imaging and HRIM data. In 93% (37/40) of swallows, the results from the EIT method were in agreement with fluoroscopy results with one of three patterns: 1) 25 normal bolus transit, 2) 8 bolus stasis, and 3) 4 retrograde escape or reflux. Three swallows (8%) had slight retention at the transition zone or distal esophagus (Z2/Z1<0.3) identified by EIT, but no retention was detected by videofluoroscopy. There was a strong correlation between percent of bolus area retained in the esophagus detected by fluoroscopic imaging and percent of bolus area retention (Z2/Z1) after swallows with EIT method (r=0.96 for supine and r= 0.69 for upright position, p<0.001). Conclusions: There was a strong correlation between percent of areas retained in the esophagus detected by fluoroscopic imaging and the EIT method and thus, bolus volume retention can be estimated by the Z2/Z1 EIT parameter. Outcome-based prospective studies are needed to clarify whether the Z2/Z1 ratio is a better
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Sa1963 Low Molecular Weight Procyanidins From Grape Seeds Enhance the Impact of 5-Fluorouracil Chemotherapy on Colon Cancer Cells Ker Y. Cheah, Gordon S. Howarth, Keren A. Bindon, James A. Kennedy, Suzanne Mashtoub, Susan E. Bastian Introduction: Grape seed procyanidins (PC) are flavan-3-ol oligomers and polymers known for their biological activity in the gut. Grape seed PCs have been reported to reduce intestinal injury in rat models of intestinal mucositis [Cheah et al. Cancer Biol Ther (2009)] and ulcerative colitis [Cheah et al. Dig Dis Sci (2013)]. We sought to determine the differential efficacy of purified PC fractions differing in mean degree of polymerization (mDP) on the viability of colon cancer cells (Caco-2) when combined with 5-Fluorouracil (5-FU) chemotherapy. Methods: Six PC fractions (F1-F6) were isolated from Cabernet Sauvignon seeds at two ripeness stages: pre-veraison unripe (immature) and ripe (mature), utilizing step gradient, low-pressure chromatography on a Sephadex LH-20 resin. Fractions were characterized by phloroglucinolysis and gel permeation chromatography (GPC). The antioxidant capacity of the fractions was determined by ferric reducing antioxidant power (FRAP) assay. Fractions were tested on Caco-2 cells, alone and in combination with 5-FU. Cell viability was determined by 3-(4,5-Dimethylthiazol-2yl)-2,5-diphenyl-tetrazolium bromide) (MTT) assay. Statistical significance was assumed at p<0.05. Results: The antioxidant capacity of six fractions was negatively correlated with PC mDP (r2 = -0.81, p<0.05). All isolated fractions significantly reduced Caco-2 cell viability compared to control (p<0.05), although F2 and F3 were the most active fractions (immature F2 = 32%, F3 = 35% and mature F2 = 13% and F3 = 17%; percentage of viable cells remaining) on Caco-2 cells. When combined with 5-FU, immature seed fractions F1-F3 and mature seed fractions F1-F4 enhanced the growth-inhibitory effects of 5-FU by 27-73% and 60-83% (p<0.05; compared to 5-FU control), respectively. Moreover, some fractions were more potent at decreasing viability in Caco-2 cells (p<0.05; immature F2-F3 = 65-68%; mature F2-F4 = 83-87%) compared to 5-FU alone (37%). Conclusions: Procyanidin fractions of mDP 2-6 (immature F1-F3 and mature F1 and F4) exhibited synergistic effects on viability of Caco-2 cells when tested in combination with 5-FU. Concomitant use of grape seed PCs and 5-FU chemotherapy could represent a promising new approach for colon cancer chemoprevention.
Sa1966 Predictors of Right-Sided Polyp Growth in a Veteran Population Cynthia E. Quainoo, Sonia DeAlwis, Ching-Ho Huang, Vidushi Golla, Michael Haynes, Yasemin Aytaman, Albert Ndzengue, Kathlynn Caguiat, Ayse Aytaman, Manuel Martinez, Garrett Lawlor INTRODUCTION The clinical implications of right sided colon polyps is gaining interest though the exact difference of their pathogenesis as compared to left sided colon polyps remains unclear. Thus far, there is suggestion that right sided colon polyps carry increased risks of malignant transformation and this may be attributable to both environmental and host factors. The aim of our study is to better understand the factors which impact the rate of right sided colon polyp growth and pathogenesis. METHODS All patients undergoing two colonoscopies at least 12 months apart between 1996 and 2012 with detection and removal of a polyp on index colonoscopy, were identified using our endoscopic database to determine the incidence of polyps and adenomas. We recorded patient age, interval between colonoscopies, gender, race, BMI, HbA1C (>6.5, <6.4), HCV Ab positivity and statin use (Y/N), folate, Vitamin D use, Aspirin and NSAIDS. Patients were classified according to age (<49, 50-59, 60-69, 70-79, > 80yrs). From the endoscopic data, we recorded polyp histology, location, size and prep score. RESULTS Overall, 1056 patients underwent two colonoscopies at least 12 months apart (median interval 47 months) with removal of polyp on initial examination. Considering all patients, there were more right sided polyps among the white population (56%) compared to the black population (44.4%, P 0.0056). Statins were used by 57.9% of patient population; their use was associated with an increased percentage of right sided polyps (54.6% versus 46%, P 0.03). Aspirin use was found to increase the percentage of right sided polyps relative to left sided (56.5% versus 47.4% P 0.0219. Finally, Vitamin D supplementation was associated with a trend towards increased right sided polyps (OR 1.474, P 0.0716, CI 0.9648-2.25). CONCLUSION The pathogenesis and natural history of right -sided polyps appears to differ compared to left -sided polyps, though this mechanism is unclear. Our data suggest that race, statin use, aspirin, and vitamin D supplementation appear to impact right - sided polyp growth independent of age or median time between colonoscopies(while also impacting left-sided polyp growth). The effect of medication and non-colorectal disease on polyp growth will increasingly be recognized as differential between left and right-sided polyps, owing to their poorly understood, different mechanisms of growth.
Sa1964 Gastric Cancer Chemoprevention by Phenethyl Isothiocyanate-Containing Diet in Chemically - but Not Genetically - Induced Gastric Cancer in Mice Anders Øverby, Chun-Mei Zhao, Timothy C. Wang, Atle Bones, Duan Chen Background/aim: Vegetables containing isothiocyanates (ITCs) such as broccoli, cauliflower and watercress have been linked epidemiologically to decreased risk of gastric cancer. The aim of the present study was to demonstrate the chemopreventive effect of the naturally occurring aromatic phenethyl ITC (PEITC) in vivo in mouse models of gastric cancer and in vitro using human gastric cancer cell lines. Methods: FVB mice treated with N-nitrosoN-methylurea (MNU, orally for 10 weeks) and transgenic INS-GAS mice with spontaneous gastric cancer were employed. Tumorigenesis was assessed by measurement of tumor size using a point counting method. PEITC-containing diet (5 μmol/g PEITC) was given ad libitum to FVB mice that were either treated with MNU simultaneously or immediately after 10-week MNU treatment for 9 months. INS-GAS mice were fed the standard mouse diet with or without PEITC for 11.5 months (starting at preneoplastic stage, i.e., 4-5 weeks of age). For the in vitro study, the gastric cancer cell lines Kato-III and MKN74 were used. Cell proliferation was determined with the MTT-assay, while morphology and cell migration were analysed using a contrast microscope, cell cycle distribution was analysed with flow cytometry, and the effect of PEITC on microtubules was examined in transfected cells by confocal microscopy. Results: The PEITC-containing diet was without effect on the body weight development in all groups of mice. Both PEITC regimens whether it was administered prior or subsequent to MNU treatment yielded a significant reduced tumorigenesis in MNUtreated mice, with chemoprevention pre-MNU feeding being more potent than post-MNU feeding. There was no significant inhibitory effect of PEITC-containing diet on tumorigenesis in INS-GAS mice compared with age-matched INS-GAS mice without PEITC-containing diet. In vitro studies using Kato-III and MKN74 cells showed that PEITC inhibited cell proliferation in a time- and dose-dependent manner in the concentration range of 1-100 μM for 24, 48 and 72 hours, with Kato-III cells being more sensitive to PEITC than MKN74 cells. The reduction in cell survival coincided with changes in cell morphology and reduced cell migration. Furthermore, treatment of PEITC for 24 hours led to an accumulation of cells in G2/M-phase of the cell cycle. Kato-III cells expressing GFP-tagged microtubules showed that PEITC induced disruption of microtubules with the subsequent formation of apoptotic blebs. Conclusions: The present study demonstrates a chemopreventive effect of PEITC-containing diet in a mouse model of chemically-induced gastric cancer. The anticancer effect may involve disruption of microtubules, resulting in cell cycle arrest and apoptosis.
Sa1967 Lipid Profile and Polyp Size in Subjects With Colon Adenomas Sandar Linn, Carmen M. Stanca, Gregory Rozansky, Juan Tejada, Philip Xiao, Sury Anand BACKGROUND: Increased expression of LDL receptor mRNA has been described in colon cancer tissue. Malignant colon tumors are thought to be involved in cholesterol catabolism, and curative surgery would lead to changes in cholesterol levels. We examined the lipid profile of patients with colonic adenomas. METHODS: Adult patients who underwent polypectomy over a one year period were enrolled in this retrospective study. Controls included patients coming to the ambulatory clinic during the same period for routine visits. We recorded demographics, lipid levels, co-morbidities, and statin use in both study and control groups; we documented polyp size in patients undergoing polypectomy. Patients with concomitant colon or other malignancies were excluded. Data were analyzed using SPSS; results are shown as median (range). RESULTS: Ninety-four subjects with colon adenomas and one hundred and thirty controls were enrolled. Fifty-four percent of subjects were women and 88% were African American. The median age was 62 (23-94). There were no significant differences in terms of age, gender, race, BMI and statin use between the adenoma group and the controls. Significantly lower LDL-cholesterol levels were identified in the adenoma group (median 54 vs 114; p<0.001) in the absence of statin use. All the other
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also had 4-fold higher levels of miR-145 (n= 3, p<0.05). We could not demonstrate any EGFRESR2 physical interactions. Surprisingly, our preliminary results showed an upregulation in EGFR signals in ERB041 treated ESR2 transfectants. Summary: In AA with adenomas, reductions in ESR2 expression might contribute to higher colon cancer incidence and worse prognosis. The in vitro studies confirmed that ESR2 suppresses colon cancer cell proliferation. We postulate that the antiproliferative effects of ESR2 involve miR-145 dependent, EGFRindependent pathways. Our study suggests that ESR2 down-regulation might play an important role in colon cancer risk and contribute to racial differences in cancer progression.
AGA Abstracts
serum lipids had higher levels in the adenoma group compared to controls: total cholesterol (186 vs 146; p<0.001), triglycerides (104 vs 72; p=0.002), and HDL-cholesterol (105 vs 45; p=0.075 did not reach statistical significance). An inverse correlation was found between the total cholesterol level and the polyp size (Spearman's r=-0.343; p = 0.047). No statistically significant differences were identified in the serum lipid levels in the adenoma group after polypectomy. CONCLUSION: Subjects with colon adenomas have significantly lower LDLcholesterol levels, and their total cholesterol level inversely correlates with polyp size. Serum lipid levels do not change after polypectomy. Although the cause-effect relationship between lipid levels and colon cancer is not clearly defined, changes in the lipid profile seem to precede the development of colon cancer. This study offers additional insights in the complex relationship between lipid metabolism and colonic neoplasms. Sa1968 Mesalamine Exhibits a Protective DNA Damage Response in Normal Colon Epithelial Cells Under Oxidative Stress Vineeta Khare, Mario Asboth, Christoph Gasche
Figure: Kaplan-Meier survival for patients with native stage I and downstaged clinical stage III rectal cancer. Adj = adjuvant. CR = complete responder.
Background: Ineffective DNA damage response such as cell cycle arrest and DNA repair contributes to colorectal cancer (CRC). Recently, our lab demonstrated that in colitis and colitis-associated cancer (CAC) oxidative stress causes DNA double strand breaks (DSB). Pharmacological manipulation of DNA damage response could be a potential mechanism for chemoprevention in CAC. Mesalamine (5-ASA) is commonly used as an anti-inflammatory compound in the treatment of UC and improves replication fidelity by activation of a replication checkpoint. Here we examined the effect of 5-ASA on DNA damage and DNA repair in normal colon epithelial cells (and a colorectal cancer cell line as control). Methods: Normal diploid human colon epithelial cells (HCEC-1CT) and HCT116 cells (CRC control) were treated with 5-ASA (5mM; 24h). Proteins implicated in DSB were analyzed by Western blot using various antibodies for DNA damage response proteins in the nuclear enriched cellular fraction. To assess the effect of 5-ASA on oxidative stress, cells were treated with H2O2 (100μM, 30 min) in the presence or absence of 5-ASA (5h pretreatment). Nuclear foci formation by 53BP1 and gamma-H2AX in response to oxidative stress was determined by immunocytochemistry. BrdU incorporation was measured to assess cell proliferation during recovery phase (24h). Results: DNA damage sensor proteins MRE11, γ-H2AX, and 53BP1 were strongly induced upon 5-ASA treatment in HCEC-1CT. Inversely, such proteins were highly expressed in HCT116 and were downregulated upon 5-ASA. In HCEC-1CT, 5-ASA resulted in diffuse nuclear localization of 53BP1 and γ- H2AX. Oxidative stress, however, resulted in redistribution of these proteins into distinct foci indicative of DNA damage. Pretreatment with 5-ASA reduced foci formation under oxidative stress. Compared to untreated HCEC-1CT, cell proliferation was significantly reduced upon H2O2 treatment (3% relative BrdU incorporation), however, 5-ASA pretreatment was protective (45%). HCT116 cells remained proliferative under oxidative stress and 5-ASA pretreatment showed no such effect (80% and 81.5%, respectively). Conclusion: 5-ASA treatment induces DNA damage response in normal colon epithelial cells and exhibits a protective effect under oxidative stress by induction of a double-strand break repair pathway impeding with CAC development. Advanced dysplastic lesions are rather unlikely to benefit from 5-ASA.
Sa1970 CD98 As a Ligand for Colitis-Targeted Delivery of Nanoparticle Bo Xiao, Emilie Viennois, Yuchen Zhang, Saravanan Ayyadurai, Hamed Laroui, Didier Merlin Background and Aims: Treatment strategies for inflammatory bowel disease (IBD) have been constrained by limited therapeutic efficacy and serious adverse effects owing to a lack of ligand for targeted drug delivery to the inflamed colon. Upon inflammation, CD98 expression is highly elevated in colonic epithelial cells and infiltrating immune cells. To investigate whether CD98 can be used as a colitis-targeted delivery ligand, we constructed CD98 Fabbearing quantum dots (QDs)-loaded nanoparticles (Fab'-NPs). Materials and Methods: CD98 Fab' was conjugated to the surfaces of QDs-loaded NPs mediated by bi-functional PEG derivative (MAL-PEG-NHS). Next, we evaluated their physicochemical properties (e.g., hydrodynamic particle size, zeta-potential, emission and absorption properties) in relation to their targeting capacity. We also investigated their targeting capacities to CD98 overexpressed colonic cells using flow cytometry. Finally, we tested their ability to target colitis tissue. Results: The resultant Fab'-NPs had desired particle size (~458 nm) with a narrow size distribution and zeta-potential (approximately +19 mV), low cytotoxicity, and excellent fluorescence properties. Electron microscopy images provided direct evidence for the welldispersed distribution of QDs within spherical Fab'-NPs. Cellular uptake experiments demonstrated that Fab'-NPs were efficiently internalized into Colon-26 and RAW 264.7 cells through the CD98-mediated endocytosis pathway, and showed that the targeting effect of CD98 Fab' markedly increased their cellular uptake efficiency compared with control pegylated QDs-loaded NPs (PEG-NPs). Furthermore, ex vivo studies showed much more effective accumulation of Fab'-NPs in colitis tissue than that of PEG-NPs. Conclusion: In the context of IBD therapy, one of the most relevant features of the drug formulation is that they can targetable deliver therapeutic molecules to specific cells. Our above findings suggest that active colitis-targeted delivery can be accomplished using NPs decorated with CD98 antibody based on the inflammation-dependent over-expression of CD98.
Sa1969 Effects of Neoadjuvant and Adjuvant Therapy for Rectal Cancer on Long Term Survival Mohan K. Mallipeddi, Paul J. Speicher, Asvin M. Ganapathi, Brian R. Englum, John Migaly, Christopher R. Mantyh
Sa1971 High-Resolution Esophageal Impedance Topography (EIT) Parameters for Quantifying Bolus Retention Zhiyue Lin, Frédéric Nicodème, Chen-Yuan Lin, Benjamin Mogni, Laurel Friesen, Peter J. Kahrilas, John E. Pandolfino
Purpose: Pre-operative chemoradiation (C/XRT) for advanced rectal cancers often pathologically downstages tumors and significantly decreases local recurrence. Adjuvant chemotherapy is also recommended for advanced staged rectal cancer to reduce systemic recurrence. However, it is unknown what C/XRT downstaging effect has on overall long term survival. Also it is unclear what effect omission of adjuvant chemotherapy has on overall survival. Methods: Patients with clinical stage III rectal cancer who underwent pre-op C/XRT in 2006 were identified from the National Cancer Database (NCDB), which captures cancer site specific data from over 1500 centers across the US. Overlapping survival and chemotherapy data only were available for 2006. Patients who underwent resection with curative intent were included for analysis. Kaplan-Meier survival estimates were used to compare survival by pathological stage with native stage I patients and complete responders as references. Cox proportional hazards modeling was used to assess the impact of demographic, clinical, and pathological factors on survival as well as the interaction between adjuvant therapy and degree of neoadjuvant downstaging. Results: There were 818 patients with clinical stage III disease who received neoadjuvant therapy. Pathology revealed 403 ypStageIII patients, 193 ypStageII patients, and 186 ypStageI patients, and 36 complete responders (CRs). Overall, 231 patients received adjuvant chemotherapy, 25% of ->ypI patients vs 35% of ->ypIII patients (p<0.001). Five-year overall survival for all cIII->ypI patients compares favorably to native stage I disease at 83.3% versus 77.7%, respectively (Hazard Ratio: 0.98, CI: 0.63, p=0.909) (Figure). The subset of the cIII->ypI patients who received adjuvant therapy had significantly better unadjusted survival than native stage I patients (p=0.007). For ->CR, ->ypI, ->ypII, and ->ypIII patients receiving adjuvant therapy, 5-year survival was 94.4%, 92.8%, 90.5%, and 63.1%, respectively. Among patients who were downstaged to ypI, there was a trend towards a survival advantage with the addition of adjuvant chemotherapy (HR: 0.25, CI: 0.05-1.12, p=0.07), and adjuvant therapy provided a significantly larger survival benefit with increasing degree of downstaging (p=0.028). Yet, in the NCDB less than 50% of downstaged cStage III patients through 2011 were receiving adjuvant therapy. Conclusions: Patients with stage III rectal cancer that were downstaged to stage I had equivalent longterm survival compared to native stage I patients. Progressive downstaging by neoadjuvant C/XRT significantly improved survival after adjuvant C/XRT. Unfortunately, a minority of rectal cancer patients received adjuvant therapy which likely reduced overall survival. Future work should focus on improving utilization of adjuvant therapy in these patients.
AGA Abstracts
Background and Aim: The multichannel intraluminal impedance (MII) technique allows one to evaluate bolus transit without radiation. However, the line tracing analysis mode is limited by low spatial resolution and the higher resolution impedance color contour plot view (CCPV) mode does not provide quantitative analysis of bolus volume on MII data. This study aimed to develop a new method for quantifying bolus volume clearance using highresolution esophageal impedance topography (EIT) technique. Methods: The ability of highresolution EIT parameters to detect bolus volume clearance was validated and tested with concurrent fluoroscopic imaging and high-resolution impedance manometry (HRIM) studies (barium bolus) in 10 healthy subjects (4 males, mean age 28 years, range 21-47). HRIM data from each subject were analyzed using a MATLAB program customized for calculating EIT parameters for each swallow within a region of interest from lower limit of UES to upper limit of EGJ in distance and from onset of swallowing to 12 seconds later or until completion of peristalsis (red dashed rectangle in Figure). The impedance volume defined as the sum of all pixel volumes (i.e., impedance value less than 50% drop from the mean baseline value to the mean nadir impedance (white curves) multiplied by time interval and by spatial interval) were derived in area 1 prior to the contraction wave-front at 20 mmHg isobaric contour (black solid curves) as Z1 and in area 2 after the contraction as Z2, and presented as a ratio of Z2 area/Z1 area. Results: A total of 40 thin barium swallows (4 per subject; 2 supine and 2 upright) were analyzed from the 10 subjects with simultaneous fluoroscopic imaging and HRIM data. In 93% (37/40) of swallows, the results from the EIT method were in agreement with fluoroscopy results with one of three patterns: 1) 25 normal bolus transit, 2) 8 bolus stasis, and 3) 4 retrograde escape or reflux. Three swallows (8%) had slight retention at the transition zone or distal esophagus (Z2/Z1<0.3) identified by EIT, but no retention was detected by videofluoroscopy. There was a strong correlation between percent of bolus area retained in the esophagus detected by fluoroscopic imaging and percent of bolus area retention (Z2/Z1) after swallows with EIT method (r=0.96 for supine and r= 0.69 for upright position, p<0.001). Conclusions: There was a strong correlation between percent of areas retained in the esophagus detected by fluoroscopic imaging and the EIT method and thus, bolus volume retention can be estimated by the Z2/Z1 EIT parameter. Outcome-based prospective studies are needed to clarify whether the Z2/Z1 ratio is a better
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