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INTRODUCTION: In biomarker-driven clinical trials, translational strategies typically involve moving findings from animal experiments to human trials. Typically, the translation is static, using a fixed model derived from animal experiments for the duration of the trial. But Bayesian designs, capable of incorporating information external to the experiment, provide a dynamic translational strategy. The current article demonstrates an example of such a dynamic Bayesian strategy in a clinical trial. METHODS: This study explored the effect of a personalized dose of fish oil for reducing prostaglandin E(2), an inflammatory marker linked to colorectal cancer. A Bayesian design was implemented for the dose-finding algorithm that adaptively updated a dose–response model derived from a previously completed the animal study during the clinical trial. In the initial stages of the trial, the dose–response model parameters were estimated from the rodent data. The model was updated following a Bayesian algorithm after data on every ten to 15 subjects were obtained until the model stabilized. Subjects were enrolled in the study between 2013 and 2015, and the data analysis was carried out in 2016. RESULTS: Three dosing models were used for groups of 16, 15, and 15 subjects. The mean target dose significantly decreased from 6.63 g/day (Model 1) to 4.06 g/day (Model 3) (p=0.001). Compared with the static strategy of dosing with a single model, the dynamic modeling reduced the dose significantly by about 1.38 g/day, on average. CONCLUSIONS: A Bayesian design was effective in adaptively revising the dosing algorithm, resulting in a lower pill burden. TRIAL REGISTRATION: This study is registered at www.clinicaltrials.gov NCT01860352.

BACKGROUND: The intestinal microbiome is an important determinant of inflammatory balance in the colon that may affect response to dietary agents. OBJECTIVE: This is a secondary analysis of a clinical trial, the Fish Oil Study, to determine whether interindividual differences in colonic bacteria are associated with variability in the reduction of colonic prostaglandin E(2) (PGE(2)) concentrations after personalized supplementation with ω-3 (n–3) fatty acids. METHODS: Forty-seven healthy adults (17 men, 30 women, ages 26–75 y) provided biopsy samples of colonic mucosa and luminal stool brushings before and after personalized ω-3 fatty acid supplementation that was based on blood fatty acid responses. Samples were analyzed using 16S ribosomal RNA sequencing. The data analyses focused on changes in bacterial community diversity. Linear regression was used to evaluate factors that predict a reduction in colonic PGE(2). RESULTS: At baseline, increased bacterial diversity, as measured by the Shannon and Inverse Simpson indexes in both biopsy and luminal brushing samples, was positively correlated with dietary fiber intakes and negatively correlated with fat intakes. Dietary supplementation with ω-3 fatty acids increased the Yue and Clayton community dis-similarity index between the microbiome in luminal brushings and colon biopsy samples post-supplementation (P = 0.015). In addition, there was a small group of individuals with relatively high Prevotella abundance who were resistant to the anti-inflammatory effects of ω-3 fatty acid supplementation. In linear regression analyses, increases in diversity of the bacteria in the luminal brushing samples, but not in the biopsy samples, were significant predictors of lower colonic PGE(2) concentrations post-supplementation in models that included baseline PGE(2), baseline body mass index, and changes in colonic eicosapentaenoic acid–to–arachidonic acid ratios. The changes in bacterial diversity contributed to 6–8% of the interindividual variance in change in colonic PGE(2) (P = 0.001). CONCLUSIONS: Dietary supplementation with ω-3 fatty acids had little effect on intestinal bacteria in healthy humans; however, an increase in diversity in the luminal brushings significantly predicted reductions in colonic PGE(2). This trial was registered at www.clinicaltrials.gov as NCT 01860352.

The overall goal of this study was to determine whether Aquamin®, a calcium-, magnesium-, trace element-rich, red algae-derived natural product, would alter the expression of proteins involved in growth-regulation and differentiation in colon. Thirty healthy human subjects (at risk for colorectal cancer) were enrolled in a three-arm, 90-day interventional trial. Aquamin® was compared to calcium alone and placebo. Before and after the interventional period, colonic biopsies were obtained. Biopsies were evaluated by immunohistology for expression of Ki67 (proliferation marker) and for CK20 and p21 (differentiation markers). Tandem mass tag-mass spectrometry-based detection was used to assess levels of multiple proteins. As compared to placebo or calcium, Aquamin® reduced the level of Ki67 expression and slightly increased CK20 expression. Increased p21 expression was observed with both calcium and Aquamin®. In proteomic screen, Aquamin® treatment resulted in many more proteins being upregulated (including pro-apoptotic, cytokeratins, cell–cell adhesion molecules, and components of the basement membrane) or downregulated (proliferation and nucleic acid metabolism) than placebo. Calcium alone also altered the expression of many of the same proteins but not to the same extent as Aquamin®. We conclude that daily Aquamin® ingestion alters protein expression profile in the colon that could be beneficial to colonic health.

Background: Esophageal adenocarcinoma is a molecularly heterogeneous disease that is rising rapidly in incidence and has a poor prognosis. Current methods of endoscopic surveillance in patients with Barrett’s esophagus (BE) using white light illumination are not sensitive to pre-malignant lesions that are flat in appearance and patchy in distribution. We aim to demonstrate specific binding by a peptide heterodimer to Barrett’s neoplasia in human subjects. Methods: Peptide monomers specific for EGFR and ErbB2 were arranged in a heterodimer configuration and labeled with IRDye800. The near-infrared (NIR)-labeled heterodimer was synthesized using current good manufacturing practices (cGMP) methods and performed pharmacology/toxicology study in animals using good laboratory practices (GLP). The NIR contrast agent was topically administered in the distal esophagus of patients with history of neoplastic BE undergoing endoscopic therapy or surveillance. Fluorescence images were collected using a flexible fiber accessory passed through the instrument channel of an upper endoscope. Fluorescence images were collected from n = 31 BE patients, and a deep learning model was used to segment the target (T) and background (B) regions to calculate the T/B ratio. Findings: Between August 2018 and November 2020, 25 human subjects were enrolled for safety study and 31 patients for either evaluation or therapy of Barrett’s neoplasia were recruited for imaging study. No adverse events attributed to the heterodimer were found. The mean T/B ratio in patients for HGD and EAC were significantly greater than that for BE with low grade dysplasia, without dysplasia, or squamous mucosa. At a T/B ratio of 1.5, 94.1% sensitivity and 92.6% specificity for detection of Barrett’s neoplasia was achieved with an AUC = 0.95. The presence of Barrett’s neoplasia was validated by pathological assessment of resected specimens. Interpretation: This “first-in-human” clinical study demonstrates feasibility to detect early Barrett’s neoplasia using a NIR-labeled peptide heterodimer. Trial Registration: This study was registered online at ClinicalTrials.gov (NCT03643068) Funding National Institutes of Health. Declaration of Interest: Authors (JC, TDW) are inventors on patents filed by the University of Michigan on the peptide heterodimer used in this study. Author (EJS) is an inventor on patents filed by the University of Washington on the multimodal scanning fiber endoscope (mmSFE) used in this study. The other authors disclose no conflicts. Ethical Approval: All study protocols were reviewed and approved by the Institutional Review Board (IRB) at Michigan Medicine

This clinical trial developed a personalized dosing model for reducing prostaglandin E2 (PGE2) in colonic mucosa using ω-3 fatty acid supplementation. The model utilized serum eicosapentaenoic acid (EPA, ω-3):arachidonic acid (AA, ω-6) ratios as biomarkers of colonic mucosal PGE2 concentration. Normal human volunteers were given low and high ω-3 fatty acid test doses for 2 weeks. This established a slope and intercept of the line for dose versus serum EPA:AA ratio in each individual. The slope and intercept was utilized to calculate a personalized target dose that was given for 12 weeks. This target dose was calculated on the basis of a model, initially derived from lean rodents, showing a log-linear relationship between serum EPA:AA ratios and colonic mucosal PGE2 reduction. Bayesian methods allowed addition of human data to the rodent model as the trial progressed. The dosing model aimed to achieve a serum EPA:AA ratio that is associated with a 50% reduction in colonic PGE2. Mean colonic mucosal PGE2 concentrations were 6.55 ng/mg protein (SD, 5.78) before any supplementation and 3.59 ng/mg protein (SD, 3.29) after 12 weeks of target dosing. In secondary analyses, the decreases in PGE2 were significantly attenuated in overweight and obese participants. This occurred despite a higher target dose for the obese versus normal weight participants, as generated by the pharmacodynamic predictive model. Large decreases also were observed in 12-hydroxyicosatetraenoic acids, and PGE3 increased substantially. Future biomarker-driven dosing models for cancer prevention therefore should consider energy balance as well as overall eicosanoid homeostasis in normal tissue. Cancer Prev Res; 10(12); 729–37. ©2017 AACR.

Preliminary single-institution data suggest that fluorescence in situ hybridization (FISH) may be useful for detecting high-grade dysplasia (HGD) and esophageal adenocarcinoma (EA) in patients with Barrett’s esophagus (BE). This multicenter study aims to validate the measurement of polysomy (gain of at least two loci) by FISH as a way to discriminate degrees of dysplasia in BE specimens. Tissue specimens were collected from four different hospitals and read by both the local pathology department (“Site diagnosis”) and a single central pathologist (“Review diagnosis”) at a separate institution. The specimens then underwent FISH analysis using probes 8q24 (MYC), 9p21 (CDKN2A), 17q12 (ERBB2), and 20q13 (ZNF217) for comparison. A total of 46 non-BE, 42 non-dysplastic specialized intestinal metaplasia (SIM), 23 indefinite-grade dysplasia (IGD), 10 low-grade dysplasia (LGD), 29 HGD, and 42 EA specimens were analyzed. We found that polysomy, as detected by FISH, was the predominant chromosomal abnormality present as dysplasia increased. Polysomy was also the best predictor for the presence of dysplasia or EA when comparing its area under the curve to that of other FISH abnormalities. We observed that if at least 10% of cells had polysomy within a specimen, the FISH probe was able to differentiate between EA/HGD and the remaining pathologies with a sensitivity of 80% and a specificity of 88%. This study demonstrates that using FISH to determine the percentage of cells with polysomy can accurately and objectively aid in the diagnosis of HGD/EA in BE specimens.

Aquamin is a calcium-, magnesium-, and multiple trace element–rich natural product with colon polyp prevention efficacy based on preclinical studies. The goal of this study was to determine the effects of Aquamin on colonic microbial community and attendant metabolomic profile. Thirty healthy human participants were enrolled in a 90-day trial in which Aquamin (delivering 800 mg of calcium per day) was compared with calcium alone or placebo. Before and after the intervention, colonic biopsies and stool specimens were obtained. All 30 participants completed the study without serious adverse event or change in liver and renal function markers. Compared with pretreatment values, intervention with Aquamin led to a reduction in total bacterial DNA (P = 0.0001) and a shift in the microbial community measured by thetaYC (θYC; P = 0.0087). Treatment with calcium also produced a decline in total bacteria, but smaller than seen with Aquamin, whereas no reduction was observed with placebo in the colon. In parallel with microbial changes, a reduction in total bile acid levels (P = 0.0375) and a slight increase in the level of the short-chain fatty acid (SCFA) acetate in stool specimens (P < 0.0001) from Aquamin-treated participants were noted. No change in bile acids or SCFAs was observed with calcium or placebo. We conclude that Aquamin is safe and tolerable in healthy human participants and may produce beneficial alterations in the colonic microbial community and the attendant metabolomic profile. Because the number of participants was small, the findings should be considered preliminary.

Background and study aims: To demonstrate the clinical use of a multimodal endoscope with a targeted fluorescently labeled peptide for quantitative detection of Barrett’s neoplasia. Patients and methods: We studied 50 patients with Barrett’s esophagus using a prototype multimodal endoscope with a fluorescently labeled peptide. Co-registered fluorescence and reflectance images were converted to ratios to correct for differences in distance and geometry over the image field of view. The ratio images were segmented using a unique threshold that maximized the variance between high and low intensities to localize regions of high grade dysplasia (HGD) and esophageal adenocarcinoma (EAC). Results: Early neoplasia (HGD and EAC) was identified with 94 % specificity and 96 % positive predictive value at a threshold of 1.49. The mean results for HGD and EAC were significantly greater than those for squamous/Barrett’s esophagus and low grade dysplasia by one-way analysis of variance (ANOVA). The receiver operator characteristic curve for detection of early neoplasia had an area under the curve of 0.884. No adverse events associated with the endoscope or peptide were found. Conclusion: A multimodal endoscope can quantify fluorescence images from targeted peptides to localize early Barrett’s neoplasia. (ClinicalTrials.gov number NCT01630798.)

Systemic exposures to intestinal bacteria may play a role in the etiology of the chronic, low-grade inflammation that is associated with western diets. Production of lipopolysaccharide-binding protein (LBP) is one biomarker of increased exposures to intestinal bacteria. This study evaluated whether changes in diet quality could affect serum LBP.This was a randomized, controlled trial of Mediterranean and Healthy Eating diets over 6 months in 120 healthy subjects at increased risk of colon cancer. Blood samples obtained before and after intervention were analyzed for LBP, branched-chain fatty acids characteristic of intestinal bacteria, micronutrients and cytokines. Data were analyzed for changes in LBP over time and for predictors of LBP.Serum concentrations of branched-chain bacterial fatty acids declined significantly in both diet groups. However, there was no significant change in mean serum LBP concentrations with either diet intervention. In serum, LBP was positively associated with CRP and negatively associated with carotenoids both before and after intervention. After intervention, LBP was predicted positively by both CRP and bacterial fatty acid concentrations in serum, and negatively by serum carotenoids and the ω3/ω6 fatty acid ratio. This model accounted for 30 % of the inter-individual variation in serum LBP after intervention.These results indicate that dietary intervention over 6 months was insufficient to alter serum LBP. The relationships with inflammation-related markers, however, indicate that anti-inflammatory strategies other than changes in diet quality, such as weight loss or improved fitness, may have more potential for reducing systemic markers of LPS exposures in well-nourished populations.

Background Relatively high serum carotenoid levels are associated with reduced risks of chronic diseases, but inter-individual variability in serum carotenoid concentrations is modestly explained by diet. The bacterial community in the colon could contribute to the bioaccessibility of carotenoids by completing digestion of plant cells walls and by modulating intestinal permeability. Objective To evaluate whether colonic bacterial composition is associated with serum and colon carotenoid concentrations. Design The study was a randomized dietary intervention trial in healthy individuals who were at increased risk of colon cancer. Colon mucosal biopsy samples were obtained before and after 6 months of intervention without prior preparation of the bowels. Participants/setting Participants were recruited from Ann Arbor, MI, and nearby areas from July 2007 to November 2010. Biopsy data were available from 88 participants at baseline and 82 participants after 6 months. Methods Study participants were randomized to counseling for either a Mediterranean diet or a Healthy Eating diet for 6 months. Results At baseline, bacterial communities in biopsy samples from study participants in the highest vs the lowest tertile of total serum carotenoid levels differed by several parameters. Linear discriminant analysis effect size identified 11 operational taxonomic units that were significantly associated with higher serum carotenoid levels. In linear regression analyses, three of these accounted for an additional 12% of the variance in serum total carotenoid concentrations after including body mass index, smoking, and dietary intakes in the model. These factors together explained 36% of the inter-individual variance in serum total carotenoid concentrations. The bacterial community in the colonic mucosa, however, was resistant to change after dietary intervention with either a Mediterranean diet or Healthy Eating diet, each of which doubled fruit and vegetable intakes. Conclusions The colonic mucosal bacterial community was associated with serum carotenoid concentrations at baseline but was not appreciably changed by dietary intervention.

Early detection of precursor lesions for colorectal cancer can greatly improve survival. Pre-neoplasia can appear flat with conventional white light endoscopy. Sessile serrated adenomas (SSA) are precursor lesions found primarily in the proximal colon and frequently appear flat and indistinct. We performed a clinical study of n=37 patients using a multimodal endoscopy with a FITC-labeled peptide specific for SSA. Lesions were imaged with white light, reflectance and fluorescence. White light images were acquired before the peptide was applied and were used to help localize regions of abnormal tissues rightly. Co-registered fluorescence and reflectance images were combined to get ratio images thus the distance was corrected. We calculated the target/background ratio (T/B ratio) to quantify the images and found 2.3-fold greater fluorescence intensity for SSA compared with normal tissues. We found the T/B ratio for SSA to be significantly greater than that for normal colonic mucosa with 89.47% sensitivity and 91.67% specificity at the threshold of 1.22. An ROC curve for SSA and normal mucosa was also plotted with area under curve (AUC) of 0.93. The result also shows that SSA and adenoma are statistically significant and can be identified with 78.95% sensitivity and 90.48% specificity at the threshold of 1.66. An ROC curve was plotted with AUC of 0.88. Therefore, our result shows that the application of a multimodal endoscope with fluorescently labeled peptide can quantify images and works especially good for the detection of SSA which is a premalignant flat lesion conferring a high risk of subsequently leading to a colon cancer.

The available evidence indicates that γ-tocopherol has more potential for colon cancer prevention than α-tocopherol, but little is known about the effects of foods and supplements on tocopherol levels in human colon. This study randomized 120 subjects at increased colon cancer risk to either a Mediterranean or a Healthy Eating diet for six months. Supplement use was reported by 39% of the subjects, and vitamin E intake from supplements was 2-fold higher than that from foods. Serum α-tocopherol at baseline was positively predicted by dietary intakes of synthetic vitamin E in foods and supplements but not by natural α-tocopherol from foods. For serum γ-tocopherol, dietary γ-tocopherol was not a predictor, but dietary α-tocopherol was a negative predictor. Unlike with serum, the data supported a role for metabolic factors, and not a direct effect of diet, in governing concentrations of both α- and γ-tocopherol in colon. The Mediterranean intervention increased intakes of natural α-tocopherol, which is high in nuts, and decreased intakes of γ-tocopherol, which is low in olive oil. These dietary changes had no significant effects on colon tocopherols. The impact of diet on colon tocopherols therefore appears to be limited.

This article provides an update for the primary care community on the evidence and recommendations for colorectal cancer screening in the adult population without symptoms at average and increased risk, excluding patients with high-risk genetic syndromes. The current and possible new screening strategies are reviewed, along with clinical wisdom related to the implementation of each method.

Background & Aims Many cancers in the proximal colon develop via from sessile serrated adenomas (SSAs), which have flat, subtle features that are difficult to detect with conventional white-light colonoscopy. Many SSA cells have the V600E mutation in BRAF. We investigated whether this feature could be used with imaging methods to detect SSAs in patients. Methods We used phage display to identify a peptide that binds specifically to SSAs, using subtractive hybridization with HT29 colorectal cancer cells containing the V600E mutation in BRAF and Hs738.St/Int cells as a control. Binding of fluorescently labeled peptide to colorectal cancer cells was evaluated with confocal fluorescence microscopy. Rats received intra-colonic 0.0086 mg/kg, 0.026 mg/kg, or 0.86 mg/kg peptide or vehicle and morbidity, mortality, and injury were monitored twice daily to assess toxicity. In the clinical safety study, fluorescently labeled peptide was topically administered, using a spray catheter, to the proximal colon of 25 subjects undergoing routine outpatient colonoscopies (3 subjects were given 2.25 μmol/L and 22 patients were given 76.4 μmol/L). We performed blood cell count, chemistry, liver function, and urine analyses approximately 24 hours after peptide administration. In the clinical imaging study, 38 subjects undergoing routine outpatient colonoscopies, at high risk for colorectal cancer, or with a suspected unresected proximal colonic polyp, were first evaluated by white-light endoscopy to identify suspicious regions. The fluorescently labeled peptide (76.4 μmol/L) was administered topically to proximal colon, unbound peptide was washed away, and white-light, reflectance, and fluorescence videos were recorded digitally. Fluorescence intensities of SSAs were compared with those of normal colonic mucosa. Endoscopists resected identified lesions, which were analyzed histologically by gastrointestinal pathologists (reference standard). We also analyzed the ability of the peptide to identify SSAs vs adenomas, hyperplastic polyps, and normal colonic mucosa in specimens obtained from the tissue bank at the University of Michigan. Results We identified the peptide sequence KCCFPAQ and measured an apparent dissociation constant of K d = 72 nM and an apparent association time constant of K = 0.174 min −1 (5.76 minutes). During fluorescence imaging of patients during endoscopy, regions of SSA had 2.43-fold higher mean fluorescence intensity than that for normal colonic mucosa. Fluorescence labeling distinguished SSAs from normal colonic mucosa with 89% sensitivity and 92% specificity. The peptide had no observed toxic effects in animals or patients. In the analysis of ex vivo specimens, peptide bound to SSAs had significantly higher mean fluorescence intensity than to hyperplastic polyps. Conclusions We have identified a fluorescently labeled peptide that has no observed toxic effects in animals or humans and can be used for wide-field imaging of lesions in the proximal colon. It distinguishes SSAs from normal colonic mucosa with 89% sensitivity and 92% specificity. This targeted imaging method might be used in early detection of premalignant serrated lesions during routine colonoscopies. ClinicalTrials.gov ID: NCT02156557.

The galactose oxidase-Schiff (GOS) reaction detects D-galactose-β-[1,3]-N-acetyl-D-galactosamine. This is a T-antigen expressed in mucus from malignant cells and colonic mucosa adjacent to cancer but not in normal mucosa. Previous studies using a qualitative GOS assay proved to be of limited value for the detection of colorectal neoplasia. We used a newly developed quantitative GOS assay to determine its potential as an early detection biomarker for colorectal cancer. We completed a multi-center, prospective, cross-sectional cohort validation study consisting of 70 normal controls, 23 high-risk normal patients (polyp history or family history of colorectal cancer (CRC) with currently normal colonoscopy), 137 patients with adenomatous polyps, and 69 with colorectal cancers. Prior to colonoscopy, two samples of stool were collected via a rectal exam: one for FOBT, and one for GOS. The area under the ROC curve (AUC) for detecting colonic adenomas and cancer for normal colons, computed with logistic regression was 0.69 for GOS, 0.62 for FOBT, and 0.73 for GOS combined with FOBT. Adding GOS to FOBT did not significantly change the ROC of FOBT alone. GOS does not appear to be a suitable marker of colorectal neoplasia.

Introduction: Previous murine studies have demonstrated that dietary Aquamin®, a calcium-rich, multi-mineral natural product, suppressed colon polyp formation and transition to invasive tumors more effectively than calcium alone when provided over the lifespan of the animals. Methods: In the present study, we compared Aquamin® to calcium for modulation of growth and differentiation in human colon adenomas in enteroid culture. Adenoma growth and differentiation were assessed at the light and electron microscopic levels and by immunostaining. Image quantitation of immunohistochemical markers was performed using Aperio Imagescope to compare these interventions. A proteomic assessment was done to compare protein expression patterns in treated cultures as compared to cultures maintained under control conditions (calcium 0.15 mM). Enteroids established from normal colonic tissue were examined in parallel. Results: Both calcium and Aquamin® provided at 1.5 mM calcium fostered differentiation in the adenoma enteroid cultures as compared to control, but Aquamin® was more effective. Even at a concentration providing only 0.15 mM calcium, Aquamin® induced differentiation in some individual enteroids. Aquamin® was more effective than calcium in inducing upregulation of proteins. There were 35 proteins upregulated with Aquamin as compared to 20 with calcium 1.5mM at 1.8-fold change or above (in 3 colonic adenomas) with less than 2% FDR. Both calcium and Aquamin® induced differences in the expression pattern of proteins known to be involved in differentiation (table). In addition, several keratins and histones, as well as merlin and Olfactomedin-4 were upregulated. Conclusion: These findings support that i) calcium (1.5 mM) has the capacity to modulate growth and differentiation in large human colon adenomas and ii) the additional trace elements provided along with calcium in Aquamin® can have effects on proliferation and differentiation at lower levels than observed with calcium at 1.5mM. Expression of Immunohistochemical Markers in enteroid cultures of Human Adenomas (Fold-Changes)IHC MarkersCalcium 0.15mMAquamin 0.15mMCalcium 1.5mMAquamin 1.5mMKi6710.750.59*0.49*CK2012.13*2.36*2.35*E-Cadherin11.051.57*1.57*NF2 (Merlin)12.44*#1.012.56*#Occludin11.23*1.25*1.29**reflects significance as compared to Ca 0.15mM. #reflects significance as compared to Ca 1.5mM Citation Format: Shannon D. McClintock, Justin A. Colacino, Durga Attili, Michael K. Dame, Areeba H. Rizvi, Aliah Richter, Anusha R. Reddy, Venkatesha Basrur, D. Kim Turgeon, James Varani, Muhammad N. Aslam. Differentiation of human colon adenomas in an enteroid culture: A treatment comparison [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1276.

A 27-year-old male presented with recurrent abdominal pain and high volume hematochezia despite undergoing extensive testing and a right hemicolectomy 3 years prior for a linear bleeding ulceration in the ascending colon. Studies at the University of Michigan included esophagogastroduodenoscopy (EGD), colonoscopy and video capsule endoscopy (VCE), revealing an arteriovenous malformation (AVM) in the terminal ileum. He was hospitalized for recurrent symptoms. His presentation suggested a small bowel source of obscure-overt GI bleeding based on prior non-diagnostic colonoscopy and EGD and a bilious nasogastric lavage. Tagged red blood cell scan localized bleeding to the right lower quadrant. Colonoscopy showed fresh blood in the terminal ileum without a clear source. Angiography showed no evidence of bleeding or terminal ileal AVM. A novel Meckel’s scan fused with SPECT imaging showed focal uptake in the terminal ileum. The patient underwent Meckel’s diverticulectomy with sparing of adjacent bowel and has remained asymptomatic for 19 months. This case illustrates that patients with obscure-overt GI bleeding require a step-wise multi-modality diagnostic work-up. Because Meckel’s scans are false-positive in 28% of adults, Meckel’s scan fused with SPECT imaging may offer an approach to refine diagnostic accuracy of either scan alone, but requires further investigation. Exploratory laparotomy should be reserved as a last option and is best performed with intraoperative endoscopy.

Human DNA fragments, as well a as whole colonocytes, can be found in stool and interrogated for mutations associated with carcinogenesis. Multiplexed panels of point mutations in fecal DNA improved the sensitivity of cancer detection (51.6%; 95% CI, 34.8%-68%) compared with fecal occult blood testing (12.9%; 95% CI, 5.1-28.9). However, the multiplex panels are not more sensitive than fecal occult blood testing in detecting advanced adenomas. Markers of epigenetic events, such as DNA methylation of gene promoters in genes not previously known to be associated with carcinogenesis or colorectal cancer, have recently been shown to be potential markers for early detection. Future approaches to improve sensitivity of fecal DNA detection of colorectal adenocarcinoma may require the inclusion of epigenetic biomarkers, the use of colonocytes isolated from feces, or both.

Esophageal adenocarcinoma (EAC) arises from Barrett’s esophagus, a columnar metaplasia of the epithelium caused by chronic acid reflux.1 This disease is growing faster than any other type of cancer in Western countries,2 and despite extensive efforts for prevention, the incidence remains high and the 5- year survival rate is poor. Early detection may allow for more effective interventions, and is critical to improving prognosis. Current methods of surveillance with white light endoscopy are limited in effectiveness because pre-malignant lesions (dysplasia) are flat in appearance and difficult to visualize.3 Molecular imaging is an emerging technology that can help physicians guide biopsy by generating image contrast based on unique biological properties tissue.4 Imaging agents that are specific for overexpressed molecular targets can improve diagnosis, staging, intervention, and management of this disease. Early imaging validation is needed to establish safety and evidence of efficacy.5

Starting with properly collected, processed, annotated, and stored human specimens is critical for successful research using unsupervised, high-throughput technologies for discovery, validation, and translation from such samples. Samples that are not reproducibly collected and managed undermine data generated in platforms, such as mass spectroscopy–based proteomic technologies. The key factors that determine quality of human biosamples are use of consistent methodologies for collection, handling, processing, and storing specimens; understanding the potential sources of bias; and thoroughly annotating the life-cycle of the specimen. These factors apply when a new biospecimen resource is being created or when previously collected or banked samples are being used. Standard operating procedures permit control of bias and ensure that high-quality biosamples result in reproducible data. Compliance with increasingly complex human subject protection regulations governing the collection, management, storage, sharing, and use of human biosamples is essential to ensure that data will be published in high-quality journals, that intellectual property is protected, and that data generated using biosamples can be used as part of submissions to governmental regulatory bodies for product approvals.

Interpatient differences in the kinetics of cyclosporine appear to result in part from interindividual differences in the catalytic activity of an enzyme termed P450 3A. We investigated the relationship between P450 3A activity, as measured by the erythromycin breath test (ERMBT), and the appropriate stable daily dose of cyclosporine as currently determined by physicians at our institution. The ERMBT was administered to kidney and heart allograft recipients who had attended at least two monthly clinic visits without having their daily cyclosporine dose changed. There was a significant positive correlation between the ERMBT result and the daily cyclosporine doses (in milligrams per kilogram) in both the heart (r = 0.68; p = 0.04; n = 9) and kidney (r = 0.68; p = 0.03; n = 10) recipients. To confirm our findings, we prospectively administered the ERMBT on multiple occasions to 20 patients who were undergoing kidney transplantation. Although the transplant physicians were blinded to the ERMBT results, the test predicted the stable daily doses of cyclosporine that they ultimately prescribed to the patients (r = 0.54; p = 0.015). When data from all 39 patients were pooled and subjected to multiple regression analysis, the ERMBT was the only variable examined that significantly correlated with the stable daily cyclosporine dose (r = 0.63; p < 0.001; n = 39). In the 20 patients prospectively studied, the prescribed daily dose of cyclosporine generally decreased during the months after surgery and the percentage changes in cyclosporine daily dose correlated with changes in P450 3A activity during this period (r = 0.47; p = 0.03). We conclude that interpatient and intrapatient differences in P450 3A activity in part account for the cyclosporine dosing practices of transplant physicians. Clinical Pharmacology and Therapeutics (1994) 56, 253–260; doi:10.1038/clpt.1994.135

Medical imaging is an invaluable tool for diagnosis, surgical guidance, and assessment of treatment efficacy. The Network for Translational Research (NTR) for Optical Imaging consists of four research groups working to “bridge the gap” between lab discovery and clinical use of fluorescence- and photoacoustic-based imaging devices used with imaging biomarkers. While the groups are using different modalities, all the groups face similar challenges when attempting to validate these systems for FDA approval and, ultimately, clinical use. Validation steps taken, as well as future needs, are described here. The group hopes to provide translational validation guidance for itself, as well as other researchers.

It has been shown recently that cyclosporine is largely metabolized by P450IIIA (CYP3A), an enzyme whose catalytic activity varies significantly among patients. To determine whether heterogeneity in P450IIIA activity contributes to interpatient differences in cyclosporine dosing requirements, the oral pharmacokinetics of the drug were determined in 20 stable kidney transplant recipients. P450IIIA activity was then measured in each patient by use of the erythromycin breath test. In the 16 patients who were at steady state, the logarithm of the apparent oral clearance of cyclosporine correlated significantly with the rate of 14CO2 exhaled in breath after intravenous administration of [14C N-methyl]erythromycin (r = 0.55, p = 0.03). No significant correlations existed between apparent oral clearance and age, high-density lipoprotein cholesterol or low-density lipoprotein cholesterol, or hematocrit in these patients. We conclude that heterogeneity in P450IIIA activity significantly contributes to interpatient differences in dosing requirements of cyclosporine in kidney transplant patients.

There are significant interpatient differences in the activity of a major drug metabolizing enzyme termed P450IIIA. Because P450IIIA uniquely catalyzes the N-demethylation of erythromycin, we have proposed that the P450IIIA activity of a patient may be determined from the rate of 14CO2 production in the breath after an intravenous infusion of a test dose of [14C-N-methyl]erythromycin. However, direct evidence that this erythromycin breath test selectively measures P450IIIA and not other major human liver P450s in patients has been lacking. We therefore administered the erythromycin breath test to nine patients with severe liver disease who were awaiting liver transplantation. Microsomes were prepared from liver samples obtained during surgery and the concentration of P450IA2, P450IIC8, P450IIC9, P450IIE1, and P450IIIA were determined immunochemically. We found a significant correlation between patients' erythromycin breath test results and their liver P450IIIA levels (r2 = 0.56, p = 0.02). In contrast, there was no correlation at all between the erythromycin breath test result and the microsomal levels of any of the other four P450s assayed. The correlation of the erythromycin breath test and P450IIIA did not appear related to the extent of liver disease because neither correlated with prothrombin time or albumin or bilirubin levels. These data provide the best evidence to date that the erythromycin breath test is a specific assay of in vivo P450IIIA activity in patients.

Conventional colonoscopy misses some neoplastic lesions. We compared the sensitivity of chromoendoscopy and colonoscopy with intensive inspection for detecting adenomatous polyps missed by conventional colonoscopy. Fifty subjects with a history of colorectal cancer or adenomas underwent tandem colonoscopies at one of five centers of the Great Lakes New England Clinical Epidemiology and Validation Center of the Early Detection Research Network. The first exam was a conventional colonoscopy with removal of all visualized polyps. The second exam was randomly assigned as either pan-colonic indigocarmine chromoendoscopy or standard colonoscopy with intensive inspection lasting >20 minutes. Size, histology, and numbers of polyps detected on each exam were recorded. Twenty-seven subjects were randomized to a second exam with chromoendoscopy and 23 underwent intensive inspection. Forty adenomas were identified on the first standard colonoscopies. The second colonoscopies detected 24 additional adenomas: 19 were found using chromoendoscopy and 5 were found using intensive inspection. Chromoendoscopy found additional adenomas in more subjects than did intensive inspection (44% versus 17%) and identified significantly more missed adenomas per subject (0.7 versus 0.2, P < 0.01). Adenomas detected with chromoendoscopy were significantly smaller (mean size 2.66 ± 0.97 mm) and were more often right-sided. Chromoendoscopy was associated with more normal tissue biopsies and longer procedure times than intensive inspection. After controlling for procedure time, chromoendoscopy detected more adenomas and hyperplastic polyps compared with colonoscopy using intensive inspection alone. Chromoendoscopy detected more polyps missed by standard colonoscopy than did intensive inspection. The clinical significance of these small missed lesions warrants further study.

Background and Aims: Lynch syndrome (also known as hereditary nonpolyposis colon cancer) is associated with an increased risk for colorectal cancer, which can arise despite frequent colonoscopic exams. We evaluated the adenoma miss rate of conventional colonoscopy in patients with Lynch syndrome, and compared the sensitivity of chromoendoscopy versus intensive inspection for detecting polyps missed by conventional colonoscopy. Methods: Fifty-four subjects with Lynch syndrome underwent tandem colonoscopies at four centers of the Great Lakes-New England Clinical Epidemiology and Validation Center of the Early Detection Research Network. All participants first had a conventional colonoscopy with removal of all visualized polyps. The second endoscopy was randomly assigned as either pancolonic indigo carmine chromoendoscopy or standard colonoscopy with intensive inspection lasting >20 minutes. Size, histology, and number of polyps detected on each exam were recorded. Results: After undergoing standard colonoscopy, 28 individuals were randomized to a second exam with chromoendoscopy and 26 underwent intensive inspection. The mean interval since last colonoscopy was 17.5 months. Seventeen polyps (10 adenomas and 7 hyperplastic polyps) were identified on the first standard colonoscopies. Twenty-three additional polyps (12 adenomas and 11 hyperplastic polyps) were found on the second exams, yielding an adenoma miss rate of 55%. Fifteen polyps (5 adenomas and 10 hyperplastic polyps) were found in subjects who had chromoendoscopy and 8 polyps (7 adenomas and 1 hyperplastic polyp) in those who had intensive inspection. Chromoendoscopy was associated with more normal tissue biopsies (11 versus 5) and longer procedure times compared with intensive inspection (29.8 ± 9.5 versus 25.3 ± 5.8 minutes; P = 0.04). Controlling for age, number of previous colonoscopies, procedure time, and prior colonic resection, chromoendoscopy detected more polyps (P = 0.04), but adenoma detection was not significantly different compared with intensive inspection (P = 0.27). Conclusions: Small adenomas are frequently missed in patients with Lynch syndrome. Although chromoendoscopy did not detect more missed adenomas than intensive inspection in this pilot study, larger trials are needed to determine optimal surveillance techniques in this high-risk population.

Preclinical studies of chemoprevention drugs given in combination at low doses show remarkable efficacy in preventing adenomas with little additional toxicities, suggesting a strategy to improve risk to benefit ratios for preventing recurrent adenomas. Three hundred seventy-five patients with history of resected (≥3 mm) adenomas were randomly assigned to receive oral difluoromethylornithine (DFMO) 500 mg and sulindac 150 mg once daily or matched placebos for 36 months, stratified by use of low-dose aspirin (81 mg) at baseline and clinical site. Follow-up colonoscopy was done 3 years after randomization or off-study. Colorectal adenoma recurrence was compared among the groups with log-binomial regression. Comparing the outcome in patients receiving placebos to those receiving active intervention, (a) the recurrence of one or more adenomas was 41.1% and 12.3% (risk ratio, 0.30; 95% confidence interval, 0.18-0.49; P < 0.001); (b) 8.5% had one or more advanced adenomas, compared with 0.7% of patients (risk ratio, 0.085; 95% confidence interval, 0.011-0.65; P < 0.001); and (c) 17 (13.2%) patients had multiple adenomas (>1) at the final colonoscopy, compared with 1 (0.7%; risk ratio, 0.055; 0.0074-0.41; P < 0.001). Serious adverse events (grade ≥3) occurred in 8.2% of patients in the placebo group, compared with 11% in the active intervention group (P = 0.35). There was no significant difference in the proportion of patients reporting hearing changes from baseline. Recurrent adenomatous polyps can be markedly reduced by a combination of low oral doses of DFMO and sulindac and with few side effects.

This year, the American Cancer Society reported that the rate of decline in both the incidence and mortality of colorectal cancer has increased over the last two decades. This success is felt to be attributable to the early detection and treatment of colonic adenomas and early-stage colorectal cancers. However, the current recommended 'menu of options' for screening is limited by poor patient acceptance, low sensitivity, and both high cost and poor accessibility for application to a large general screening population (colonoscopy). Computerized tomography and magnetic resonance colonography offer an alternative method for the identification of polyps and early lesions in certain patients, but have cost, access, and acceptance limitations that are similar to those of colonoscopy; thus, they present similar barriers to their use in broad population screening. These limitations provide a strong rationale for the development of early colorectal cancer detection biomarkers that are simple to use and are cost effective. A successful biomarker or biomarker panel, coupled with the colonoscopic follow-up of only those patients with positive results, would reduce the burden and morbidity associated with the screening of colonoscopy. This would most likely result in enhanced adherence to colorectal screening, as well as a dramatic reduction in the incidence and mortality rates of colorectal cancer. In this paper, we review recent advances in the discovery of potential colorectal cancer biomarkers. Their applicability to clinical population screening will require large prospective validation.

Alosetron hydrochloride (Lotronex®, GlaxoSmithKline, Inc) is a safe and effective agent for selective patients with severe irritable bowel syndrome when prescribed as recommended. We describe the first reported case of acute liver injury in a 39-year-old white woman who developed symptomatic hepatitis 28 days after starting alosetron. All other competing causes of acute hepatitis were excluded by radiologic and laboratory studies and the liver injury resolved after drug discontinuation. Although the mechanism of alosetron hepatotoxicity is unknown, metabolic idiosyncrasy is suspected since the drug is known to be extensively metabolized by cytochrome-P450 enzymes. Clinicians prescribing alosetron should be aware of this potential side effect if unexplained abdominal pain, jaundice,or abnormal liver biochemistries are encountered in a treated patient.

Polyethylene glycol-electrolyte lavage solutions are widely used to prepare the colon for colonoscopy. Unfortunately, some patients find this preparation difficult to complete. Recent studies of a sodium phosphate-based laxative have shown both good patient tolerance and good bowel preparation. In these studies, the laxative has generally been prescribed in two doses, with the second dose taken early the morning of colonoscopy. Because the morning dose is inconvenient for many patients, we compared giving a common polyethylene glycol-based electrolyte lavage solution the day before colonoscopy with our method of giving both doses of sodium phosphate-based laxative the day before colonoscopy: one dose at 4 PM and the second dose at 8 PM. We judged efficacy by an assessment of residual liquid and fecal matter in the colon and judged tolerance by the results of a symptom questionnaire completed by each patient immediately before the procedure. Our results in more than 200 patients showed similar efficacy ratings and similar symptom scores for both preparations, but patients rated the sodium phosphate-based preparation as easier to tolerate. In conclusion, in selected patients this new dosing method for sodium phosphate is preferable to large-volume, whole-gut lavage solutions.

Background: Curcumin, a dietary polyphenol derivative of turmeric, has potent cancer reductive activity in in vivo colon carcinogenesis models. We administered curcumin at doses of 2 g or 4 g to separate groups of 20 healthy human smokers with > 8 aberrant crypt foci (ACF). The 2 g dose showed no ACF reduction while the 4 g dose showed a 46% reduction (p Methods: Curcuminoid products were extracted from homogenized biopsy tissue and ultacentrifuged plasma and evaporated under argon. The separation was performed on a Symmetry® C18 column. The mobile phase used under gradient conditions was 25% acetonitrile with 74.9% containing 0.1% acetic acid (25:74.9:0.1; v/v/v; A) and 100% acetonitrile containing 0.1% acetic acid (99.9:0.1; v/v; B) The flow rate was 0.3 mL/min and injection volume for all samples was 10 µL. The lower limit of detection for curcumin from human plasma was 5 ng/mL and from human colonic mucosa was 50 ng/5mg tissue. Results: See Table below (summarized by pre and post treatment dose for plasma and mucosal concentrations) Conclusions: Native curcumin is not detectable in the majority of biopsy and plasma samples. Conjugated curcumin is detectable in all plasma samples and the majority of mucosal biopsies with a significant increase in plasma concentrations at 4g. We postulate the conjugates detected in the rectal mucosa are delivered systemically from absorption and conjugation higher in the gastrointestinal tract and may have biologic activity. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1286. doi:10.1158/1538-7445.AM2011-1286

The production of 14CO2 in the breath from an intravenous dose of [14C-N-methyl]-erythromycin (the erythromycin breath test [ERMBT]) and the measurement of the ratio of 6-beta-cortisol to free cortisol (6-beta-F/FF) in the urine have each been proposed as means of measuring hepatic P450IIIA catalytic activity in patients. We found that there was a significant correlation between the results of each test (r = 0.59, p less than 0.001) in 47 patients who were without liver disease and who were not taking medications believed to influence P450IIIA catalytic activity. In the 24 of these patients who were subsequently treated with the P450IIIA substrate cyclosporine, the ERMBT result was highly correlated with the mean trough cyclosporine blood level observed; however, there was no correlation between urinary 6-beta-F/FF and the cyclosporine blood levels. In a separate study of a patient during the anhepatic phase of liver transplantation surgery, the ERMBT result decreased by greater than 85%, whereas urinary 6-beta-F/FF decreased by just 50%. We conclude that the ERMBT and urinary 6-beta-F/FF do not always provide similar information about P450IIIA catalytic activity in patients, possibly because of extrahepatic production of 6-beta-F. Of the two tests, the ERMBT appears to provide the most relevant information for cyclosporine administration.

B135 Epidemiological and animal studies indicate that the major components of the traditional Cretan-Mediterranean diet have great promise for the prevention of colon cancer while improving overall health. We designed an exchange-list diet that would target 1) an increase in amount and variety of fruits and vegetables, 2) an increase in whole grains and 3) a decrease in n-6 fatty acids with increased n-3 and n-9 fatty acids. The exchange list approach provided flexibility in food choices for meeting nutrient goals. In this ongoing study we are recruiting study participants at increased risk for colon cancer. Subjects are being randomized to a modified Mediterranean diet or a Healthy People 2010 diet for 6 months, each of which involves intensive dietary counseling. Dietary intake data, blood samples and biopsies of normal colonic mucosa from flexible sigmoidoscopy are being collected and analyzed. Thus far 45 subjects have been enrolled with a mean age of 52 years, mean BMI of 28 kg/m2 and 67% were female. Two thirds of the subjects had a family history of colon cancer and the remainder had a personal history of adenomas. Dietary intakes of mono-unsaturated fats increased by about 70% and fruit and vegetables intakes doubled, but caloric intake did not change appreciably in the Mediterranean arm. This indicates that the exchange list diet was an effective tool. HPLC-MS-MS methods were developed to profile prostaglandins and leukotrienes in granulocytes and colonic mucosa using a single chromatographic run. Thus far granulocyte samples from 23 subjects have been analyzed there was a trend for decreases in several eicosanoids in samples from the Mediterranean arm while changes in the Healthy Eating arm were more variable. These preliminary data indicate that subjects can make large dietary changes and that this may impact on inflammation. Effects on biomarkers in the colonic mucosa are ongoing and will be important to evaluate the potential preventive effects of a Mediterranean type of diet. Supported by NIH grant 1RO1CA120381. Citation Information: Cancer Prev Res 2008;1(7 Suppl):B135.

Introduction: Epidemiologic and experimental data suggests that colon cancer should be largely preventable through dietary or chemical intervention. Polyamines are actively involved in proliferation and differentiation of many tissues. Lowering of intracellular levels via inhibition of the initial step (ornithine decarboxylase) in the polyamine synthesis pathway by the specific enzyme-activated inhibitor DFMO results in a decrease of adenomas and cancers in many preclinical organ-site cancer models, most notably in the colon. Methods: From 1989-1997 we conducted a series of pilot and Phase IIa (short term) and Phase IIb (long term) trials in which we determined the lowest dose of DFMO at which polyamine content of the colonic flat mucosa would be lowered, with the goal of finding a non-toxic dose. In 1997 we debated whether to do a definitive trial with DFMO alone or to combine it with another chemoprevention compound. We settled on combining DFMO with the NSAID sulindac at low dose for a variety of mechanistic, experimental, and clinical reasons and began a Phase IIb trial in which the recurrence of adenomas would be correlated with polyamine changes in the flat mucosa. Several years later we expanded the trial to a full Phase III trial in which recurrence of adenomas became the primary endpoint. Results: Based on the recommendation of the Data Safety and Monitoring Board an accelerated second analysis was done after 70% of patients had completed their 3 year or off-study colonoscopy and the study was stopped for efficacy as the study goals had been met. Overall, a highly significant reduction (70%) of all adenomas, advanced adenomas (92% reduction), and multiple adenomas (95% reduction) was seen (all P

This study recruited persons at increased risk of colon cancer to an intensive dietary intervention study that required biopsies of the colon by flexible sigmoidoscopy at baseline and after six months of intervention. A total of 1314 individuals contacted the study, and only 16 individuals indicated that the sigmoidoscopy procedure was an obstacle to study participation. A total of 270 individuals completed a screening visit and signed a screening consent form. Inquiries about the study tended to be fewer in the winter and late summer. Failure to return food records was the most common reason for exclusion. Dietary recall at enrollment indicated that subjects were consuming significantly more vegetables, lower sodium and a lower glycemic load on the day before starting the study vs. during the eligibility phase which might have an impact on biomarker measures. This makes it important to capture dietary changes in the period between determination of eligibility and enrollment. Subjects (n = 120) were randomized to follow a Healthy Eating or a Mediterranean Diet, each of which required substantial dietary record-keeping. The study completion rate was 78%, and subjects reported high satisfaction with study participation. Of the 93 individuals who completed the study, only one refused the flexible sigmoidoscopy at the final visit. These findings suggest that flexible sigmoidoscopy does not appear to be a barrier for recruitment of high-risk individuals to an intensive dietary intervention trial, but that completing food records can be.