AGA Abstracts downregulated in gastric cancer tissues relative to non-tumor gastric mucosae. Downregul- ation of miR-29c was associated with progression of gastric cancer, and was more prominent in advanced gastric cancers than in gastric adenomas and early gastric cancers (p<0.05). In addition, expression of the oncogene Mcl-1, a target of miR-29c, was significantly increased in gastric cancer tissues relative to non-tumor gastric mucosae. Activation of miR-29c by celecoxib induced suppression of Mcl-1 and apoptosis in gastric cancer cells. Celecoxib activated the expression of miR-29c, but not that of miR-29b, suggesting that celecoxib may modulate the binding of transcriptional factors between miR-29b and miR-29c. The database (www.gene-regulation.com/pub/programs.html), indicated that C/EBPα binding sites were located in the promoter region of miR-29c. The chromatin immunoprecipitation (ChIP) assay showed that immunoprecipitation with the C/EBPα antibody was significantly increased in AGS cells treated with celecoxib, indicating that celecoxib activates the expression of miR- 29c by enhancing the binding of C/EBPα to the promoter. Conclusions : Downregulation of the tumor suppressor miR-29c plays critical roles in the progression of gastric cancer, and that miR-29c is a novel therapeutic target for gastric cancer. Selective COX-2 inhibitors may have clinical promise for the chemoprevention of gastric cancer via activation of miR-29c. 668 Nanoparticle-Based Delivery of DCAMKL-1 SiRNA and DAPT Increases MicroRNA-144 and Inhibits Colorectal Cancer Tumor Growth via a Notch-1 Dependent Mechanism Sripathi M. Sureban, Randal May, Dongfeng Qu, Fadee G. Mondalek, Sima Asfa, Rama Ramanujam, Shrikant Anant, Courtney W. Houchen Background and Aim: Nanotechnology is an exciting new research field that combines chemistry, engineering and biology. It has enormous potential in cancer biology by providing early detection, precise diagnosis, and targeted therapeutic drug delivery. Nanoparticles (NP) are extremely small (<100 nanometers in size and can be used to create powerful interactions with biomolecules both on the surface of and inside of cancer cells. Development of an optimal delivery system capable of protecting and transporting siRNA through extracellular and intracellular barriers to reach the site of action in the cytosol has been elusive. We have previously reported that DCAMKL-1 is overexpressed in human colorectal cancers and that liposomal preparation of siDCAMKL-1 resulted in HCT116 human colorectal cancer tumor xenograft growth arrest. These findings were associated with an upregulation of the tumor suppressor miRNA let-7a and a corresponding decrease in the oncogene c-Myc. In this report we have combined the application of NP technology and siRNA biology in order to provide a target specific drug delivery platform to knockdown the potential key cancer regulatory molecule DCAMKL-1. Methods: mRNA and miRNA expressions were determined by real- time RT PCR, protein by Immunohistochemistry and western blot. siRNA-targeting DCAMKL-1 was encapsulated in Poly(lactide-co-glycolide) (PLGA)-based nanoparticles (NP- siDCAMKL-1) and transfected into HCT116 cells; injected directly to HCT116 tumor xenog- rafts in nude mice and tumor volumes were measured. Tumor xenografts were also treated with DAPT (γ-secretase inhibitor) alone or in combination with NP-siDCAMKL-1. HCT116 cells were transfected separately with plasmids encoding the firefly luciferase gene with a let-7a and miR-144 miRNA binding sites at the 3'UTR to measure let-7a and miR-144 expression. Results: NP delivered siDCAMKL-1 resulted in HCT116 tumor growth arrest, down regulated proto-oncogene c-Myc and Notch1 via let-7a and miR-144 miRNA dependent mechanism respectively. An increase in let-7a and miR-144 miRNA following NP-siDCAMKL- 1 was confirmed by a corresponding reduction in let-7a and miR-144 specific luciferase activity. Knockdown of DCAMKL-1 also induced miR-200a, an EMT inhibitor, along with down-regulation of EMT-associated transcription factors ZEB1, ZEB2, Snail, Slug and Twist. Furthermore, DAPT-mediated knockdown of Notch1 alone and in combination with NP- siDCAMKL-1 resulted in HCT116 tumor growth arrest and down regulation of Notch1 via miR-144 dependent mechanism. Conclusion: These findings confirm that nanoparticle-based delivery of siRNAs directed at critical targets may have profound therapeutic implications for solid tumor cancer treatment. Furthermore these results implicate regulation of miRNAs a potentially novel direct approach to treating cancer in the future. 669 The Tryptophan Catabolism Pathway: A Novel Therapeutic Target in Colitis Associated Cancer Matthew A. Ciorba, Ameet I. Thaker, Lynne R. Foster, Suprada Rao, William F. Stenson BACKGROUND: Colitis associated cancer is an important complication of the human inflammatory bowel diseases (IBD). Indoleamine 2,3 Dioxygenase (IDO), the rate limiting enzyme in tryptophan catabolism, is over-expressed in the epithelium of active IBD and some colon cancers. Specific inhibitors of IDO1 are under investigation for their ability to break tumor-induced immune tolerance induced by IDO1 mediated suppression of T- effector cell responses. IDO inhibition in acute colitis also reduces regenerative epithelial cell proliferation. AIM: Examine the role of IDO1 in colitis associated cancer models in the presence and absence of adaptive immunity. METHODS: Comparison groups were WT vs IDO-/- as well as Rag1-/- vs DbKO (Rag1-/- x IDO-/-, Double KO), all on the C57Bl/6 background. Co-caging was used in all experiments. Acute colitis experiments used 4% dextran-sodium-sulfate (DSS) for 5 days, followed by 5 days recovery. Colitis associated cancer induction used a standardized protocol of azoxymethane (AOM: 10mg/kg) IP prior to 3 week-long cycles of 2.5% DSS. Clinical Disease Activity Index (DAI) was followed and histology examined. Epithelial proliferation was assessed by BrDU incorporation. RESULTS: High IDO expression was identified in the malignant epithelium by IHC and RT-PCR. In acute DSS colitis, IDO-/- mice exhibited greater weight loss (-11% vs -21%, p<.01), higher disease activity index scores (3 vs 6.4, p<.05) during recovery phase and worse histology severity (mean ulcer length from anorectal junction 9.8 vs 14.3 mm, p<0.01). Relative epithelial proliferation index at ulcer margin was lower in IDO-/- vs WT mice (4.1 vs 8.3 fold, p<0.05). Over the 60 day AOM/DSS protocol, IDO-/- vs WT mice had higher disease activity index during the first and second, but not third DSS cycle. At sacrifice tumor number (4.1 vs 7.5, p<0.01) and total tumor area (13.7 vs 31.2 sq mm, p=0.017) were lower in IDO-/- vs WT mice. Correspondingly, the adenoma epithelial proliferation index was lower in IDO-/- vs WT mice (20.2% vs 41%, p=0.04). In mice lacking intact adaptive immunity S-122 AGA Abstracts (Rag1-/- vs DbKO), deletion of IDO resulted in no difference in DAI scores or histology severity. Surprisingly however, the reduced tumor phenotype persisted with DbKO mice developing fewer (avg # 5.4 vs 14, p<.001) and smaller (17.9 vs 39.1 sq mm, p=.001) tumors than Rag1-/- mice. CONCLUSIONS: These data show that IDO is an important regulator of colitis associated cancer progression where dual roles of promoting tolerance from the adaptive immune system and facilitated cancer cell proliferation are implicated. While IDO blockade worsens acute colitis, its effects in chronic colitis are less pronounced. Inhibition of IDO may be an effective therapeutic strategy for colon cancer, particularly inflammation associated colon cancer. 739 The Evaluation of Intraesophageal Air Kinetics of Esophageal Speech Measuring by Multichannel Intraesophageal Impedance in Patients With Total Laryngectomy Aykut Bozan, Rukiye Vardar, Serdar Akyildiz, Esra Belen, Tayfun Kirazli, Serhat Bor Aim: Total laryngectomy is a mutilating operation with lifelong functional and psychological consequences. Since removal of the larynx results in loss of the sound generator, postoperative voice rehabilitation has been of major concern. In some cases the cervical oesophagus could effectively act as a neoglottis. Although esophageal speech may not be easy to achieve, it has the advantages. However no data exist about the mechanisms of esophageal speech and kinetics of air within the upper gastrointestinal tract. We aimed to measure of air within the esophagus in patients with total laryngectomy using multichannel intraesophageal impedance (MII) and possible role of upper (UES) and lower esophageal sphincter (LES) pressures. METHOD: 23 patients with total laryngectomy because of laryngeal squamous cancer (all male, mean age 59 yo) evaluated. Esophageal motility was performed with a Dent-Sleeve catheter. An impedance catheter was placed 5cm above the LES. All patients were asked to say the same letters, words, sentences and classified according to the Wepman scale to measure the quality of speech; nine good speakers (≤3; group I) and 13 poorly speakers (>3; group II). UES, LES pressures, MII findings were evaluated by two blinded specialists. FINDINGS: Mean postoperative follow-up was 45 months (8-154). Motility of esophageal body was normal in all cases. Mean UES pressure was 18.1 vs 21.2 mmHg (p=0.26) and mean LES pressure was 18 vs 15.8 mmHg for group I and II respectively (p=0.55). MII findings revealed that group I could keep the air within the esophagus. Short and fast words were related with a rapid swallows into the upper esophagus and regurgitate immediately. Long speeches were consistent with a swallow of air deep into the lower esophagus and stomach followed by upper esophagus was filled from this reservoir. No air has been observed in group II patients. Two independent observer could easily differentiate the quality of the speech from MII tracings. RESULTS: This is the first study showing that esophageal speech was achieved when air was swallowed and regurgitated in a controlled manner. Short sounds were produced using the air from upper esophagus and long speeches were needed to fill of whole the esophagus and possibly upper part of the stomach. The ability of speech following total laryngectomy was not related with UES or LES pressures even UES pressures were lower than normal values because of surgery. This study raised the possibility that whether MII might be use as an objective tool to show the patients how to swallow and regurgitate the air in terms of a successful speech. 740 The Contractile Deceleration Point (CDP) Occurs in Synchrony With the Maximum Length of the Contracting Segment During Normal Peristalsis in Esophageal Pressure Topography (EPT) Studies John E. Pandolfino, Zhiyue Lin, Sabine Roman, Monika A. Kwiatek, Peter J. Kahrilas Background: The conventional depiction of peristalsis is of pressure variation over time at a fixed luminal position along the horizontal axis. With EPT it is equally feasible to depict pressure variation along the vertical axis of the esophagus at a fixed time (spatial pressure variation plots). Mechanistically, spatial pressure variation plots delineate the instantaneous length and location of the contracting segment. As peristalsis approaches the end of the esophagus, the leading edge of the contraction slows, defining the CDP, which is indicative of the transition from peristaltic transport to ampullary emptying. We hypothesized that the length of the contracting segment of the esophagus during normal peristalsis achieves a maximum at the CDP because thereafter the unslowed tail begins to “catch up” with the slowed leading edge (Figure). METHODS: EPT studies of 72 normal subjects (10 swallows each) were analyzed. Data were exported from ManoView™ (Sierra) to MATLAB™. A coordinate-based strategy was used to normalize both the length of the esophagus for each subject and the time frame for all swallows as a 100 x 200 pixel grid extending from the pharynx to the stomach for a 20s period beginning 1s before upper esophageal sphincter (UES) relaxation. A composite simulation of EPT for each subject was computed based on the median pressure value within each pixel for the 10 test swallows. A spatial pressure variation function of the greatest contractile pressure along the esophagus was calculated to identify the proximal (P) and the distal troughs (D) on each EPT. A fixed threshold of 30 mmHg was used to control the search process between the P and D both in time (s) and in distance (cm) to find the maximal contracting segment length. The CDP is by localized by manually fitting two tangential lines to the initial and terminal portions of the 30-mm Hg isobaric contours and noting intersection of the lines (Figure). RESULTS: The median timing (from UES relaxation) of the CDP position closely matched that of the maximal contracting segment length (Table) (r=0.90, p<0.001 by Pearson correlation analysis). The maximal contracting length averaged 65% of the span from P to D equating to an average of 9.3 cm. CONCLUSIONS: The timing of the CDP during normal peristalsis exhibits excellent correlation with the timing at which there is a maximal length of contracting segment as derived from spatial pressure variation plots of EPT studies, which maximizes at an average of 65% of the P-D span. These findings suggest an alternative strategy for localizing the CDP, a physiological landmark that may prove crucial in the understanding of esophageal motor disorders.