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Background & Aims 5-Fluorouracil (5-FU)–based adjuvant chemotherapy does not increase survival times of patients with colorectal tumors with microsatellite instability. We determined the response of patients with colorectal tumors with the CpG island methylator phenotype (CIMP) to 5-FU–based therapy. Methods We analyzed a population-based cohort of 302 patients with colorectal cancer (CRC) for a median follow-up time of 50.7 months. CIMP status was determined by analysis of the CACNAG1 , SOCS1 , RUNX3 , NEUROG1 , and MLH1 promoters; tumors were considered to be CIMP positive if at least 3 promoters were methylated. Results Tumors from 29.5% of patients (89/302) were CIMP positive; CIMP status did not influence disease-free survival (DFS; log-rank = 0.3). Of tumors of TNM stages II–III (n = 196), 32.7% were CIMP positive. Among patients with stages II–III CRC who did not receive adjuvant 5-FU chemotherapy, those with CIMP-positive tumors had longest times of DFS (log-rank = 0.04); In patients who received chemotherapy, those with CIMP-positive tumors had shorter times of DFS (log-rank = 0.02). In patients with CIMP-negative tumors, adjuvant 5-FU chemotherapy significantly increased time of DFS (log-rank = 0.00001). However, in patients with CIMP-positive tumors, adjuvant 5-FU chemotherapy did not affect time of DFS (log-rank = 0.7). Multivariate analysis showed a significant, independent interaction between 5-FU treatment and CIMP status (hazard ratio [HR], 0.6; 95% confidence interval [CI], 0.5–0.8). Among patients with CIMP-positive tumors, adjuvant chemotherapy was not an independent predictor of outcome (HR, 0.8; 95% CI, 0.3–2.0). In patients who did not receive adjuvant 5-FU chemotherapy, CIMP status was the only independent predictor of survival (HR, 2.0; 95% CI, 1.1–3.8). Conclusions Patients with CIMP-positive colorectal tumors do not benefit from 5-FU–based adjuvant chemotherapy.

G A A b st ra ct s gastric tissue samples harvested from an independent population ofH. pylori-infected persons in New Orleans to assess expression and methylation status of HIF-1α in vivo. Gastric tissue specimens from African-American subjects harboring an increased risk for gastric cancer exhibited marked decreases in methylation of HIF-1α with increasing disease severity. The highest levels of HIF-1α methylation were found in patients with non-atrophic gastritis and this decreased as disease progressed to atrophic gastritis (0.5-fold) and intestinal metaplasia (0.2-fold), versus gastritis alone. Consistent with decreasing HIF-1α methylation status, gastric tissue from African-American patients harbored increased levels of HIF-1α expression with increasing disease progression and this was not observed in Caucasian patients. Collectively, these data indicate that H. pylori induces HIF-1α in gastric epithelial cells and this is augmented under conditions of iron deficiency. HIF-1α expression in vivo increases in conjunction with decreased HIF-1α methylation and the development of premalignant lesions, which may provide a mechanism underpinning the link between high altitude, iron depletion, and increased gastric cancer rates within the context of H. pylori infection.

Comparison Between Universal Immunohistochemistry for Mismatch Repair Proteins Versus Revised Bethesda Guidelines in the Detection of Patients With Lynch Syndrome Lucia Perez-Carbonell, Carla Guarinos, Maria Rodriguez Soler, Cristina Sanchez-Fortun, Laura Sempere-Robles, Clara Ruiz-Ponte, Adela Castillejo, Alejandro Brea, Angel Carracedo, Victor M. Barbera, Estefania Rojas, Luis Bujanda, Joan Clofent, Montserrat Andreu, Xavier Llor, Antoni Castells, Jose-Luis Soto, Rodrigo Jover

Background & Aims Approximately half of the families that fulfill Amsterdam criteria for Lynch syndrome or hereditary nonpolyposis colorectal cancer (HNPCC) do not have evidence of the germline mismatch repair gene mutations that define this syndrome and result in microsatellite instability (MSI). The carcinogenic pathways and the best diagnostic approaches to detect microsatellite stable (MSS) HNPCC tumors are unclear. We investigated the contribution of epigenetic alterations to the development of MSS HNPCC tumors. Methods Colorectal cancers were divided into 4 groups: (1) microsatellite stable, Amsterdam-positive (MSS HNPCC) (N = 22); (2) Lynch syndrome cancers (identified mismatch repair mutations) (N = 21); (3) sporadic MSS (N = 92); and (4) sporadic MSI (N = 46). Methylation status was evaluated for CACNAG1 , SOCS1 , RUNX3 , NEUROG1 , MLH1 , and long interspersed nucleotide element-1 ( LINE-1 ). KRAS and BRAF mutation status was analyzed. Results MSS HNPCC tumors displayed a significantly lower degree of LINE-1 methylation, a marker for global methylation, than any other group. Although most MSS HNPCC tumors had some degree of CpG island methylation, none presented a high index of methylation. MSS HNPCC tumors had KRAS mutations exclusively in codon 12, but none harbored V600E BRAF mutations. Conclusions Tumors from Amsterdam-positive patients without mismatch repair deficiency (MSS HNPCC) have certain molecular features, including global hypomethylation, that distinguish them from all other colorectal cancers. These characteristics could have an important impact on tumor behavior or treatment response. Studies are underway to further assess the cause and effects of these features.

Comparison Between Routine Immunohistochemistry for Mismatch Repair Proteins Versus Revised Bethesda Guidelines in the Diagnosis of Lynch Syndrome in a Non-Selected Population of Colorectal Cancer Patients Lucia Perez-Carbonell, Clara Ruiz-Ponte, Xavier Bessa, Jose-Luis Soto, Adela Castillejo, Victor Barbera, Alejandro Brea, Laura Sempere, Cristina Sanchez-Fortun, Sergi CastellviBel, Francesc Balaguer, Rosa M Xicola, Xavier Llor, Anna Abuli, Montserrat Andreu, Cristina Alenda, Artemio Paya, Angel Carracedo, Antoni Castells, Rodrigo Jover

control study of eight methylation biomarkers in BE. Progressors (Ps) were defined as BE patients with index biopsies showing no dysplasia, indefinite for dysplasia, or low-grade dysplasia (LGD) at endoscopy performed ≥6 months prior to a diagnosis of either highgrade dysplasia or EAC. Nonprogressors (NPs) were defined as patients undergoing at least 3 surveillance endoscopic examinations who never progressed beyond LGD. Ps were considered as a single combined group and in two tiers: progression within 2 years or 4 years. Methylation was assessed in 145 NPs and 50 Ps using real-time quantitative methylation-specific PCR. RESULTS: Ps were significantly older than NPs (70.6 vs. 62.5 years, p < 0.001). Normalized methylation values of HPP1, p16 and RUNX3 were significantly higher in Ps than in NPs (0.456, 0.138, and 0.104 vs. 0.273, 0.069 and 0.063; p = 0.0025, 0.0066 and 0.0002, respectively). We also evaluated a linear combination of the 8 markers using coefficients from a multivariate logistic regression analysis. Based on this model, areas under the ROC curve (AUCs) were high in the 2-year, 4-year and combined models (0.843, 0.829 and 0.840; p

Introduction: The intestinal epithelium undergoes a constant and fast renewal that is maintained in check thanks to a tight equilibrium between cell death and proliferation. An alteration of this equilibrium in the colonic epithelium can result in colorectal cancer development. Nutrients capable of either inducing programmed cell death or decreasing the proliferative rate of this epithelium could presumably be useful as either chemo preventive or even adjuvant therapeutic elements in colorectal cancer. Fish oil, through its main fatty acids, Docosahexaenoic (DHA) and Eicosahexaenoic (EPA) has shown some promising effects in colorectal cancer both, epidemiologically and experimentally. We have previously shown that DHA and EPA induce apoptosis in colorectal cancer cells. We also showed that both, the intrinsic and extrinsic pathways are involved though the down-regulation of two key elements: FLIP and XIAP. Aims: The aims of this study were: to determine if the n-3 fatty acids DHA and EPA can regulate colorectal cancer cell proliferation; to determine if this regulation is dependent on the type of molecular alterations present in these cells and; to determine if any phase of the cell cycle is affected in this potentially anti neoplastic effect. Methods: Non-confluent HCT116, LoVo, SW480, Caco-2 and HT-29 human colorectal cancer cells were grown in their regular media and, 24 h before the experiments, they were synchronized by feeding them with sera-free media supplemented with DHA, EPA or vehicle at final concentrations of 30 to 50 uM according to the type of cell studied. Cell proliferation was assessed by MTT analysis at 24 and 48h after fatty acid supplementation. Cell cycle phases were analyzed by flow citometry through propidium iodide staining. Results: DHA and EPA supplementation resulted in a decrease in the proliferation rate of most colorectal cancer cells, independently of APC, p53 or mismatch repair status. This coincided with a significant decrease in the DNA synthesis of these cells. Neither DHA nor EPA was able to stop proliferation of the metastatic LoVo cells. DNA synthesis was preserved in the latter cells as opposed to the non-metastatic cells. Conclusions: DHA and EPA can exert an anti neoplastic effect in colorectal cancer not only through a significant apoptosis induction but also through a significant decrease in cell proliferation and in DNA synthesis. These effects are independent of the responsible underlying molecular pathways. Metastatic colorectal cancer cells may have developed mechanisms that make these cells resistant to the anti proliferative effects of DHA and EPA.