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Identification of new genetic markers may improve the prediction of colorectal cancer prognosis. Our objective was to examine genome-wide associations of germline genetic variants with disease-specific survival in an analysis of 16,964 cases of colorectal cancer. We analyzed genotype and colorectal cancer-specific survival data from a consortium of 15 studies. Approximately 7.5 million SNPs were examined under the log-additive model using Cox proportional hazards models, adjusting for clinical factors and principal components. Additionally, we ran secondary analyses stratifying by tumor site and disease stage. We used a genome-wide p-value threshold of 5 × 10 -8 to assess statistical significance. No variants were statistically significantly associated with disease-specific survival in the full case analysis or in the stage-stratified analyses. Three SNPs were statistically significantly associated with disease-specific survival for cases with tumors located in the distal colon (rs698022, HR = 1.48, CI 1.30-1.69, p = 8.47 × 10 -9 ) and the proximal colon (rs189655236, HR = 2.14, 95% CI 1.65-2.77, p = 9.19 × 10 -9 and rs144717887, HR = 2.01, 95% CI 1.57-2.58, p = 3.14 × 10 -8 ), whereas no associations were detected for rectal tumors. Findings from this large genome-wide association study highlight the potential for anatomical-site-stratified genome-wide studies to identify germline genetic risk variants associated with colorectal cancer-specific survival. Larger sample sizes and further replication efforts are needed to more fully interpret these findings., (© 2022. The Author(s).)

Most risk prediction models for colorectal cancer (CRC) are based on questionnaires and show a modest discriminatory ability. Therefore, we aim to develop risk prediction models incorporating plasma biomarkers for CRC to improve discrimination. We assessed the predictivity of 11 biomarkers in 736 men in the Health Professionals Follow-up Study and 639 women in the Nurses' Health Study. We used stepwise logistic regression to examine whether a set of biomarkers improved the predictivity on the basis of predictors in the National Cancer Institute's (NCI) Colorectal Cancer Risk Assessment Tool. Model discrimination was assessed using C-statistics. Bootstrap with 500 randomly sampled replicates was used for internal validation. The models containing each biomarker generated a C-statistic ranging from 0.50 to 0.59 in men and 0.50 to 0.54 in women. The NCI model demonstrated a C-statistic (95% CI) of 0.67 (0.62-0.71) in men and 0.58 (0.54-0.63) in women. Through stepwise selection of biomarkers, the C-statistic increased to 0.70 (0.66-0.74) in men after adding growth/differentiation factor 15, total adiponectin, sex hormone binding globulin and tumor necrosis factor receptor superfamily member 1B (P for difference = 0.008); and increased to 0.62 (0.57-0.66) in women after further including insulin-like growth factor 1 and insulin-like growth factor-binding protein 3 (P for difference = .06). The NCI + selected biomarkers model was internally validated with a C-statistic (95% CI) of 0.73 (0.70-0.77) in men and 0.66 (0.61-0.70) in women. Circulating plasma biomarkers may improve the performance of risk factor-based prediction model for CRC., (© 2021 UICC.)

Background: Smoking is associated with colorectal cancer (CRC) risk. Previous studies suggested this association may be restricted to certain molecular subtypes of CRC, but large-scale comprehensive analysis is lacking., Methods: A total of 9789 CRC cases and 11 231 controls of European ancestry from 11 observational studies were included. We harmonized smoking variables across studies and derived sex study-specific quartiles of pack-years of smoking for analysis. Four somatic colorectal tumor markers were assessed individually and in combination, including BRAF mutation, KRAS mutation, CpG island methylator phenotype (CIMP), and microsatellite instability (MSI) status. A multinomial logistic regression analysis was used to assess the association between smoking and risk of CRC subtypes by molecular characteristics, adjusting for age, sex, and study. All statistical tests were 2-sided and adjusted for Bonferroni correction., Results: Heavier smoking was associated with higher risk of CRC overall and stratified by individual markers ( P trend < .001). The associations differed statistically significantly between all molecular subtypes, which was the most statistically significant for CIMP and BRAF . Compared with never-smokers, smokers in the fourth quartile of pack-years had a 90% higher risk of CIMP-positive CRC (odds ratio = 1.90, 95% confidence interval = 1.60 to 2.26) but only 35% higher risk for CIMP-negative CRC (odds ratio = 1.35, 95% confidence interval = 1.22 to 1.49; P difference = 2.1 x 10 -6 ). The association was also stronger in tumors that were CIMP positive, MSI high, or KRAS wild type when combined ( P difference < .001)., Conclusion: Smoking was associated with differential risk of CRC subtypes defined by molecular characteristics. Heavier smokers had particularly higher risk of CRC subtypes that were CIMP positive and MSI high in combination, suggesting that smoking may be involved in the development of colorectal tumors via the serrated pathway., (© The Author(s) 2021. Published by Oxford University Press.)

Background and Aims: The heterogeneity among colorectal tumors is probably due to differences in developmental pathways and might associate with patient survival times. We studied the relationship among markers of different subtypes of colorectal tumors and patient survival., Methods: We pooled data from 7 observational studies, comprising 5010 patients with colorectal cancer. All the studies collected information on microsatellite instability (MSI), CpG island methylator phenotype (CIMP), and mutations in KRAS and BRAF in tumors. Tumors with complete marker data were classified as type 1 (MSI-high, CIMP-positive, with pathogenic mutations in BRAF but not KRAS), type 2 (not MSI-high, CIMP-positive, with pathogenic mutations in BRAF but not KRAS), type 3 (not MSI-high or CIMP, with pathogenic mutations in KRAS but not BRAF), type 4 (not MSI-high or CIMP, no pathogenic mutations in BRAF or KRAS), or type 5 (MSI-high, no CIMP, no pathogenic mutations in BRAF or KRAS). We used Cox regression to estimate hazard ratios (HR) and 95% confidence intervals (CIs) for associations of these subtypes and tumor markers with disease-specific survival (DSS) and overall survival times, adjusting for age, sex, stage at diagnosis, and study population., Results: Patients with type 2 colorectal tumors had significantly shorter time of DSS than patients with type 4 tumors (HR DSS 1.66; 95% CI 1.33-2.07), regardless of sex, age, or stage at diagnosis. Patients without MSI-high tumors had significantly shorter time of DSS compared with patients with MSI-high tumors (HR DSS 0.42; 95% CI 0.27-0.64), regardless of other tumor markers or stage, or patient sex or age., Conclusions: In a pooled analysis of data from 7 observational studies of patients with colorectal cancer, we found that tumor subtypes, defined by combinations of 4 common tumor markers, were associated with differences in survival time. Colorectal tumor subtypes might therefore be used in determining patients' prognoses., (Copyright © 2020 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Background: Histological lymphocytic reaction is regarded as an independent prognostic marker in colorectal cancer. Considering the lack of adequate statistical power, adjustment for selection bias and comprehensive tumour molecular data in most previous studies, we investigated the strengths of the prognostic associations of lymphocytic reaction in colorectal carcinoma by utilising an integrative database of two prospective cohort studies., Methods: We examined Crohn's-like reaction, intratumoural periglandular reaction, peritumoural reaction and tumour-infiltrating lymphocytes in 1465 colorectal carcinoma cases. Using covariate data of 4420 colorectal cancer cases in total, inverse probability-weighted Cox proportional hazard regression model was used to control for selection bias (due to tissue availability) and potential confounders, including stage, MSI status, LINE-1 methylation, PTGS2 and CTNNB1 expression, KRAS, BRAF and PIK3CA mutations, and tumour neoantigen load., Results: Higher levels of each lymphocytic reaction component were associated with better colorectal cancer-specific survival (P trend  < 0.002). Compared with cases with negative/low intratumoural periglandular reaction, multivariable-adjusted HRs were 0.55 (95% CI, 0.42-0.71) in cases with intermediate reaction and 0.20 (95% CI, 0.12-0.35) in cases with high reaction. These relationships were consistent in strata of MSI status or neoantigen loads (P interaction  > 0.2)., Conclusions: The four lymphocytic reaction components are prognostic biomarkers in colorectal carcinoma.

Background & Aims: Human studies examining associations between circulating levels of insulin-like growth factor 1 (IGF1) and insulin-like growth factor binding protein 3 (IGFBP3) and colorectal cancer risk have reported inconsistent results. We conducted complementary serologic and Mendelian randomization (MR) analyses to determine whether alterations in circulating levels of IGF1 or IGFBP3 are associated with colorectal cancer development., Methods: Serum levels of IGF1 were measured in blood samples collected from 397,380 participants from the UK Biobank, from 2006 through 2010. Incident cancer cases and cancer cases recorded first in death certificates were identified through linkage to national cancer and death registries. Complete follow-up was available through March 31, 2016. For the MR analyses, we identified genetic variants associated with circulating levels of IGF1 and IGFBP3. The association of these genetic variants with colorectal cancer was examined with 2-sample MR methods using genome-wide association study consortia data (52,865 cases with colorectal cancer and 46,287 individuals without [controls]) RESULTS: After a median follow-up period of 7.1 years, 2665 cases of colorectal cancer were recorded. In a multivariable-adjusted model, circulating level of IGF1 associated with colorectal cancer risk (hazard ratio per 1 standard deviation increment of IGF1, 1.11; 95% confidence interval [CI] 1.05-1.17). Similar associations were found by sex, follow-up time, and tumor subsite. In the MR analyses, a 1 standard deviation increment in IGF1 level, predicted based on genetic factors, was associated with a higher risk of colorectal cancer risk (odds ratio 1.08; 95% CI 1.03-1.12; P = 3.3 × 10 -4 ). Level of IGFBP3, predicted based on genetic factors, was associated with colorectal cancer risk (odds ratio per 1 standard deviation increment, 1.12; 95% CI 1.06-1.18; P = 4.2 × 10 -5 ). Colorectal cancer risk was associated with only 1 variant in the IGFBP3 gene region (rs11977526), which also associated with anthropometric traits and circulating level of IGF2., Conclusions: In an analysis of blood samples from almost 400,000 participants in the UK Biobank, we found an association between circulating level of IGF1 and colorectal cancer. Using genetic data from 52,865 cases with colorectal cancer and 46,287 controls, a higher level of IGF1, determined by genetic factors, was associated with colorectal cancer. Further studies are needed to determine how this signaling pathway might contribute to colorectal carcinogenesis., (Copyright © 2020 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Background: We hypothesized that the risk of colorectal cancer in night-shift workers might be different according to insulin receptor substrate status., Methods: Among 77,470 eligible women having night work assessed in the Nurses' Health Study, we documented a total of 1,397 colorectal cancer cases, of which 304 or 308 had available data on IRS1 and IRS2 , respectively. We used duplication-method Cox proportional hazards regression analysis for competing risks to calculate HRs and 95% confidence intervals (CI) for each colorectal cancer subtype. We measured tumor IRS1 or IRS2 expression by immunohistochemistry (IHC)., Results: Compared with women who never worked night shifts, those working ≥15 years night shifts had a marginal trend of increased overall risk of colorectal cancer ( P trend = 0.06; multivariable HR = 1.20; 95% CI, 0.99-1.45). Longer duration of night-shift work was associated with a higher risk of IRS2 -positive tumors (multivariable HR = 2.69; 95% CI, 1.48-4.89; P trend = 0.001, ≥15 years night shifts vs. never) but not with IRS2 -negative tumors (multivariable HR = 0.90; 95% CI, 0.54-1.51; P trend = 0.72; P heterogeneity for IRS2 = 0.008). Similarly, the corresponding multivariable HRs were 1.81 for IRS1 -positive tumors (95% CI, 0.94-3.48; P trend = 0.06) and 1.13 for IRS1 -negative tumors (95% CI, 0.71-1.80; P trend = 0.56; P heterogeneity for IRS1 = 0.02)., Conclusions: Our molecular pathologic epidemiology data suggest a potential role of IRS in mediating carcinogenesis induced by night-shift work., Impact: Although these findings need validation, rotating night shift might increase colorectal cancer risk in women with abnormal insulin receptor pathways., (©2019 American Association for Cancer Research.)

Background: Prostaglandin-endoperoxide synthase 2 (PTGS2, cyclooxygenase-2, COX-2)-prostaglandin E 2 (PGE 2 ) pathway promotes tumour progression. Considering evidence suggesting increased PGE 2 synthesis by BRAF mutation in tumour cells, we hypothesised that the association of tumour PTGS2 (COX-2) expression with colorectal cancer mortality might be stronger in BRAF-mutated tumours than in BRAF-wild-type tumours., Methods: Using 1708 patients, including 1200 stage I-IV colorectal carcinoma cases in the Nurses' Health Study (NHS) and the Health Professionals Follow-up Study (HPFS) and 508 stage III colon cancer cases in a National Cancer Institute-sponsored randomised controlled trial of adjuvant therapy (CALGB/Alliance 89803), we evaluated tumour PTGS2 (COX-2) expression status using immunohistochemistry. We examined the prognostic association of PTGS2 (COX-2) expression in strata of BRAF mutation status by multivariable Cox proportional hazards regression models to adjust for potential confounders, including disease stage, tumour differentiation, microsatellite instability status and KRAS and PIK3CA mutations., Results: In NHS and HPFS, the association of PTGS2 (COX-2) expression with colorectal cancer-specific survival differed by BRAF mutation status (P interaction  = 0.0005); compared with PTGS2 (COX-2)-negative/low carcinomas, the multivariable-adjusted hazard ratios for PTGS2 (COX-2)-high carcinomas were 2.44 (95% confidence interval, 1.39-4.28) in BRAF-mutated cases and 0.82 (95% confidence interval, 0.65-1.04) in BRAF-wild-type cases. Differential prognostic associations of PTGS2 (COX-2) expression in strata of BRAF mutation status were similarly observed in CALGB/Alliance 89803 trial (P interaction  = 0.03)., Conclusions: The association of tumour PTGS2 (COX-2) expression with colorectal cancer mortality is stronger in BRAF-mutated tumours than in BRAF-wild-type tumours, supporting interactive roles of PTGS2 (COX-2) expression and BRAF mutation statuses in prognostication of patients with colorectal cancer; ClinicalTrials.gov Identifier, NCT00003835., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Purpose: SMAD4 has shown promise in identifying patients with colorectal cancer at high risk of recurrence or death. Experimental Design: A discovery cohort and independent validation cohort were classified by SMAD4 status. SMAD4 status and immune infiltrate measurements were tested for association with recurrence-free survival (RFS). Patient-derived xenografts from SMAD4-deficient and SMAD4-retained tumors were used to examine chemoresistance., Results: The discovery cohort consisted of 364 patients with stage I-IV colorectal cancer. Median age at diagnosis was 53 years. The cohort consisted of 61% left-sided tumors and 62% stage II/III patients. Median follow-up was 5.4 years (interquartile range, 2.3-8.2). SMAD4 loss, noted in 13% of tumors, was associated with higher tumor and nodal stage, adjuvant therapy use, fewer tumor-infiltrating lymphocytes (TIL), and lower peritumoral lymphocyte aggregate (PLA) scores (all P < 0.04). SMAD4 loss was associated with worse RFS ( P = 0.02). When stratified by SMAD4 and immune infiltrate status, patients with SMAD4 loss and low TIL or PLA had worse RFS ( P = 0.002 and P = 0.006, respectively). Among patients receiving 5-fluorouracil (5-FU)-based systemic chemotherapy, those with SMAD4 loss had a median RFS of 3.8 years compared with 13 years for patients with SMAD4 retained. In xenografted mice, the SMAD4-lost tumors displayed resistance to 5-FU. An independent cohort replicated our findings, in particular, the association of SMAD4 loss with decreased immune infiltrate, as well as worse disease-specific survival., Conclusions: Our data show SMAD4 loss correlates with worse clinical outcome, resistance to chemotherapy, and decreased immune infiltrate, supporting its use as a prognostic marker in patients with colorectal cancer., (©2018 American Association for Cancer Research.)

Evidence indicates a complex link between microbiota, tumor characteristics, and host immunity in the tumor microenvironment. In experimental studies, bifidobacteria appear to modulate intestinal epithelial cell differentiation. Accumulating evidence suggests that bifidobacteria may enhance the antitumor immunity and efficacy of immunotherapy. We hypothesized that the amount of bifidobacteria in colorectal carcinoma tissue might be associated with tumor differentiation and higher immune response to colorectal cancer. Using a molecular pathologic epidemiology database of 1313 rectal and colon cancers, we measured the amount of Bifidobacterium DNA in carcinoma tissue by a quantitative PCR assay. The multivariable regression model was used to adjust for potential confounders, including microsatellite instability status, CpG island methylator phenotype, long-interspersed nucleotide element-1 methylation, and KRAS, BRAF, and PIK3CA mutations. Intratumor bifidobacteria were detected in 393 cases (30%). The amount of bifidobacteria was associated with the extent of signet ring cells (P = 0.002). Compared with Bifidobacterium-negative cases, multivariable odd ratios for the extent of signet ring cells were 1.29 (95% CI, 0.74-2.24) for Bifidobacterium-low cases and 1.87 (95% CI, 1.16-3.02) for Bifidobacterium-high cases (P trend  = 0.01). The association between intratumor bifidobacteria and signet ring cells suggests a possible role of bifidobacteria in determining distinct tumor characteristics or as an indicator of dysfunctional mucosal barrier in colorectal cancer., (Copyright © 2018 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Background: Colorectal carcinoma evolves through a multitude of molecular events including somatic mutations, epigenetic alterations, and aberrant protein expression, influenced by host immune reactions. One way to interrogate the complex carcinogenic process and interactions between aberrant events is to model a biomarker correlation network. Such a network analysis integrates multidimensional tumor biomarker data to identify key molecular events and pathways that are central to an underlying biological process. Due to embryological, physiological, and microbial differences, proximal and distal colorectal cancers have distinct sets of molecular pathological signatures. Given these differences, we hypothesized that a biomarker correlation network might vary by tumor location., Results: We performed network analyses of 54 biomarkers, including major mutational events, microsatellite instability (MSI), epigenetic features, protein expression status, and immune reactions using data from 1380 colorectal cancer cases: 690 cases with proximal colon cancer and 690 cases with distal colorectal cancer matched by age and sex. Edges were defined by statistically significant correlations between biomarkers using Spearman correlation analyses. We found that the proximal colon cancer network formed a denser network (total number of edges, n = 173) than the distal colorectal cancer network (n = 95) (P < 0.0001 in permutation tests). The value of the average clustering coefficient was 0.50 in the proximal colon cancer network and 0.30 in the distal colorectal cancer network, indicating the greater clustering tendency of the proximal colon cancer network. In particular, MSI was a key hub, highly connected with other biomarkers in proximal colon cancer, but not in distal colorectal cancer. Among patients with non-MSI-high cancer, BRAF mutation status emerged as a distinct marker with higher connectivity in the network of proximal colon cancer, but not in distal colorectal cancer., Conclusion: In proximal colon cancer, tumor biomarkers tended to be correlated with each other, and MSI and BRAF mutation functioned as key molecular characteristics during the carcinogenesis. Our findings highlight the importance of considering multiple correlated pathways for therapeutic targets especially in proximal colon cancer.

Accumulating evidence suggests that post-diagnostic insulin levels may influence colorectal cancer (CRC) survival. Yet, no previous study has examined CRC survival in relation to a post-diagnostic diet rich in foods that increase post-prandial insulin levels. We hypothesized that glycemic and insulin scores (index or load; derived from food frequency questionnaire data) may be associated with survival from specific CRC subtypes sensitive to the insulin signaling pathway. We prospectively followed 1,160 CRC patients from the Nurses' Health Study (1980-2012) and Health Professionals Follow-Up Study (1986-2012), resulting in 266 CRC deaths in 10,235 person-years. CRC subtypes were defined by seven tumor biomarkers (KRAS, BRAF, PIK3CA mutations, and IRS1, IRS2, FASN and CTNNB1 expression) implicated in the insulin signaling pathway. For overall CRC and each subtype, hazard ratio (HR) and 95% confidence interval (95% CI) for an increase of one standard deviation in each of glycemic and insulin scores were estimated using time-dependent Cox proportional hazards model. We found that insulin scores, but not glycemic scores, were positively associated with CRC mortality (HR = 1.19, 95% CI = 1.02-1.38 for index; HR = 1.23, 95% CI = 1.04-1.47 for load). The significant positive associations appeared more pronounced among PIK3CA wild-type cases and FASN-negative cases, with HR ranging from 1.36 to 1.60 across insulin scores. However, we did not observe statistically significant interactions of insulin scores with PIK3CA, FASN, or any other tumor marker (p interaction > 0.12). While additional studies are needed for definitive evidence, a high-insulinogenic diet after CRC diagnosis may contribute to worse CRC survival., (© 2017 UICC.)

Purpose: Tumor levels of thymidylate synthase (TS), a target of 5-fluorouracil (5-FU)-based chemotherapy for colorectal cancer, have been studied as a predictive or prognostic biomarker with mixed results., Patients and Methods: Tumor TS levels were prospectively evaluated in two adjuvant therapy trials for patients with resected stage II or III colon cancer. TS expression was determined by standard immunohistochemistry and by automated quantitative analysis. Tumor mismatch repair deficiency (MMR-D) and BRAF c.1799T > A (p.V600E) mutation status were also examined. Relationships between tumor TS, MMR-D, and BRAF mutation status, overall survival (OS), and disease-free survival (DFS) were investigated in the subset of stage III patients., Results: Patients whose tumors demonstrated high TS expression experienced better treatment outcomes, with DFS hazard ratio (HR) = 0.67, 95% confidence interval (CI) = 0.53, 0.84; and OS HR = 0.68, 95% CI = 0.53, 0.88, for high versus low TS expression, respectively. No significant interaction between TS expression and stage was observed (DFS: interaction HR = 0.94; OS: interaction HR = 0.94). Tumors with high TS expression were more likely to demonstrate MMR-D (22.2% vs. 12.8%; p =  .0003). Patients whose tumors demonstrated both high TS and MMR-D had a 7-year DFS of 77%, compared with 58% for those whose tumors had low TS and were non-MMR-D (log-rank p =  .0006). Tumor TS expression did not predict benefit of a particular therapeutic regimen., Conclusion: This large prospective analysis showed that high tumor TS levels were associated with improved DFS and OS following adjuvant therapy for colon cancer, although tumor TS expression did not predict benefit of 5-FU-based chemotherapy. The Oncologist 2017;22:107-114Implications for Practice: This study finds that measurement of tumor levels of thymidylate synthase is not helpful in assigning specific adjuvant treatment for colorectal cancer. It also highlights the importance of using prospective analyses within treatment clinical trials as the optimal method of determining biomarker utility., (© AlphaMed Press 2016.)

Leptin is an adipokine involved in regulating energy balance, which has been identified as a potential biologic link in the development of obesity-associated cancers, such as pancreatic cancer. In this prospective, nested case-control study of 470 cases and 1,094 controls from five U.S. cohorts, we used conditional logistic regression to evaluate pancreatic cancer risk by prediagnostic plasma leptin, adjusting for race/ethnicity, diabetes, body mass index, physical activity, plasma C-peptide, adiponectin, and 25-hydroxyvitamin D. Because of known differences in leptin levels by gender, analyses were conducted separately for men and women. We also evaluated associations between 32 tagging SNPs in the leptin receptor (LEPR) gene and pancreatic cancer risk. Leptin levels were higher in female versus male control participants (median, 20.8 vs. 6.7 ng/mL; P < 0.0001). Among men, plasma leptin was positively associated with pancreatic cancer risk and those in the top quintile had a multivariable-adjusted OR of 3.02 [95% confidence interval (CI), 1.27-7.16; P trend = 0.02] compared with men in the bottom quintile. Among women, circulating leptin was not associated with pancreatic cancer risk (P trend = 0.21). Results were similar across cohorts (P heterogeneity = 0.88 for two male cohorts and 0.35 for three female cohorts). In genetic analyses, rs10493380 in LEPR was associated with increased pancreatic cancer risk among women, with an OR per minor allele of 1.54 (95% CI, 1.18-2.02; multiple hypothesis-corrected P = 0.03). No SNPs were significantly associated with risk in men. In conclusion, higher prediagnostic levels of plasma leptin were associated with an elevated risk of pancreatic cancer among men, but not among women. Cancer Res; 76(24); 7160-7. ©2016 AACR., Competing Interests: of Potential Conflicts of Interest No potential conflicts of interest were disclosed, (©2016 American Association for Cancer Research.)

Objective: Accumulating evidence links the intestinal microbiota and colorectal carcinogenesis. Fusobacterium nucleatum may promote colorectal tumour growth and inhibit T cell-mediated immune responses against colorectal tumours. Thus, we hypothesised that the amount of F. nucleatum in colorectal carcinoma might be associated with worse clinical outcome., Design: We used molecular pathological epidemiology database of 1069 rectal and colon cancer cases in the Nurses' Health Study and the Health Professionals Follow-up Study, and measured F. nucleatum DNA in carcinoma tissue. Cox proportional hazards model was used to compute hazard ratio (HR), controlling for potential confounders, including microsatellite instability (MSI, mismatch repair deficiency), CpG island methylator phenotype (CIMP), KRAS, BRAF, and PIK3CA mutations, and LINE-1 hypomethylation (low-level methylation)., Results: Compared with F. nucleatum-negative cases, multivariable HRs (95% CI) for colorectal cancer-specific mortality in F. nucleatum-low cases and F. nucleatum-high cases were 1.25 (0.82 to 1.92) and 1.58 (1.04 to 2.39), respectively, (p for trend=0.020). The amount of F. nucleatum was associated with MSI-high (multivariable odd ratio (OR), 5.22; 95% CI 2.86 to 9.55) independent of CIMP and BRAF mutation status, whereas CIMP and BRAF mutation were associated with F. nucleatum only in univariate analyses (p<0.001) but not in multivariate analysis that adjusted for MSI status., Conclusions: The amount of F. nucleatum DNA in colorectal cancer tissue is associated with shorter survival, and may potentially serve as a prognostic biomarker. Our data may have implications in developing cancer prevention and treatment strategies through targeting GI microflora by diet, probiotics and antibiotics., Competing Interests: Dr. Chan previously served as a consultant for Bayer Healthcare, Millennium Pharmaceuticals, Pozen Inc, and Pfizer Inc. This study was not funded by Bayer Healthcare, Millennium Pharmaceuticals, Pozen Inc, or Pfizer Inc. Dr. Meyerson applies a patent on Fusobacterium in colorectal cancer diagnosis, and has ownership interest in and is a consultant and advisory board member for Foundation Medicine. The other authors declare that they have no competing interests., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.)

Total circulating 25-hydroxyvitamin D [25(OH)D)] has been associated with lower risk of colorectal cancer. The physiologic mechanism, however, may be more directly related to the free or bioavailable fraction of 25(OH)D, which is influenced by levels of vitamin D binding protein (VDBP). We assessed the association of prediagnosis total, free, and bioavailable 25(OH)D and VDBP with colorectal cancer risk among predominantly white women in the Nurses' Health Study (NHS) who provided a blood specimen in 1989-1990. We documented 378 cases of colorectal cancer through 2011 and matched them to 689 controls according to age and time of blood draw. We genotyped two common polymorphisms in the gene coding VDBP and calculated free and bioavailable 25(OH)D levels based on total 25(OH)D, VDBP, albumin, and their estimated genotype-specific binding affinities. Total 25(OH)D was associated with lower colorectal cancer risk (P for trend = 0.01). Compared with women in the lowest quintile of total 25(OH)D, those in the highest quintile had a multivariable-adjusted odds ratio (OR) for colorectal cancer of 0.54 [95% confidence interval (CI), 0.33-0.87]. Comparing extreme quintiles, we did not find any significant association with risk of colorectal cancer for VDBP (OR, 0.98; 95% CI, 0.65-1.47), free 25(OH)D (OR, 0.71; 95% CI, 0.46-1.10), or bioavailable 25(OH)D (OR, 0.92; 95% CI, 0.60-1.42). In conclusion, prediagnosis levels of total, but not free or bioavailable 25(OH)D, were associated with lower colorectal cancer risk. Although our findings support an inverse association of vitamin D with colorectal cancer, this association does not appear to be due to the unbound or bioavailable fraction of circulating vitamin D. Cancer Prev Res; 9(8); 664-72. ©2016 AACR., (©2016 American Association for Cancer Research.)

Purpose: We hypothesized that adverse prognostic associations of specific tumor molecular factors vary by patient age at colorectal cancer diagnosis., Experimental Design: We examined the prognostic associations and interactions by age at colorectal cancer diagnosis (<60 vs. 60-74 vs. ≥75 years old) of key molecular factors-CpG island methylator phenotype (CIMP), microsatellite instability (MSI), KRAS, BRAF, and PIK3CA mutations, and nuclear CTNNB1 expression status-on colorectal cancer-specific survival (CSS) and overall survival (OS), using 1,280 incident colorectal cancer cases (median age, 69 years; range, 38-91 years) within the Nurses' Health Study and Health Professionals Follow-up Study cohorts., Results: MSI-high was associated with better survival, whereas BRAF mutation was associated with worse survival, but these associations did not appreciably differ by age group. Status of CIMP, KRAS mutation, or PIK3CA mutation was not associated with prognosis regardless of age. Nuclear CTNNB1 expression was associated with a trend toward worse prognosis among older adults [age ≥ 75 years; multivariate HR, 1.67; 95% confidence interval (CI), 0.89-3.13 (for CSS); multivariate HR, 1.44; 95% CI, 0.93-2.24 (for OS)] but not among younger patients, and there was a statistically significant interaction by age (Pinteraction = 0.03 for CSS; Pinteraction = 0.007 for OS)., Conclusions: Tumor nuclear CTNNB1 expression may be associated with higher mortality among older patients with colorectal cancer but not among younger patients. Our findings need to be confirmed in independent datasets. Detailed exploration of tumor molecular signatures in older patients with colorectal cancer in large populations is warranted., (©2015 American Association for Cancer Research.)

Background: High-level physical activity is associated with lower colorectal cancer (CRC) mortality, likely through insulin sensitization. Insulin receptor substrate 1 (IRS1) is a mediator of insulin and insulin-like growth factor (IGF) signaling pathways, and its down-regulation is associated with insulin resistance. Therefore, we hypothesized that tumor IRS1 expression status might modify cellular sensitivity to insulin and IGF, and the prognostic association of physical activity., Methods: We assessed IRS1 expression level in 371 stage I-III rectal and colon cancers in the Nurses' Health Study and the Health Professionals Follow-up Study by immunohistochemistry. In survival analysis, Cox proportional hazards model was used to assess an interaction between post-diagnosis physical activity (ordinal scale of sex-specific quartiles Q1 to Q4) and IRS1 expression (ordinal scale of negative, low, and high), controlling for potential confounders, including microsatellite instability, CpG island methylator phenotype, long interspersed nucleotide element-1 (LINE-1) methylation level, and KRAS, BRAF, and PIK3CA mutation status., Results: There was a statistically significant interaction between post-diagnosis physical activity and tumor IRS1 expression in CRC-specific mortality analysis (P interaction = 0.005). Multivariable hazard ratio (95% confidence interval) for higher post-diagnosis physical activity (Q3-Q4 vs. Q1-Q2) was 0.15 (0.02-1.38) in the IRS1-negative group, 0.45 (0.19-1.03) in the IRS1-low group, and 1.32 (0.50-3.53) in the IRS1-high group., Conclusions: The association of post-diagnosis physical activity with colorectal carcinoma patient survival may differ by tumor IRS1 expression level. If validated, tumor IRS1 expression status may serve as a predictive marker to identify subgroups of patients who might gain greater survival benefit from an increased level of exercise.

Background: Loss of CDH1 (E-cadherin) expression in cancer cells may promote cell migration and invasion. Therefore, we hypothesised that loss of CDH1 expression in colorectal carcinoma might be associated with aggressive features and clinical outcome., Methods: Utilising molecular pathological epidemiology database of 689 rectal and colon cancer cases in the Nurses' Health Study and the Health Professionals Follow-up Study, we assessed tumour CDH1 expression by immunohistochemistry. Multivariate logistic regression analysis was conducted to assess association of CDH1 loss with tumour growth pattern (expansile-intermediate vs infiltrative) and lymph node metastasis and distant metastasis, controlling for potential confounders including microsatellite instability, CpG island methylator phenotype, LINE-1 methylation, and PIK3CA, BRAF and KRAS mutations. Mortality according to CDH1 status was assessed using Cox proportional hazards model., Results: Loss of tumour CDH1 expression was observed in 356 cases (52%), and associated with infiltrative tumour growth pattern (odds ratio (OR), 2.02; 95% confidence interval (CI), 1.23-3.34; P=0.006) and higher pN stage (OR, 1.73; 95% CI, 1.23-2.43; P=0.001). Tumour CDH1 expression was not significantly associated with distant metastasis or prognosis., Conclusions: Loss of CDH1 expression in colorectal cancer is associated with infiltrative tumour growth pattern and lymph node metastasis., Competing Interests: ATC previously served as a consultant for Bayer Healthcare, Millennium Pharmaceuticals, Pozen Inc and Pfizer Inc. This study was not funded by Bayer Healthcare, Millennium Pharmaceuticals, Pozen Inc, or Pfizer Inc. The remaining authors declare no conflicts of interest.

Background: Pancreatic tumors cause changes in whole-body metabolism, but whether prediagnostic circulating metabolites predict survival is unknown., Methods: We measured 82 metabolites by liquid chromatography-mass spectrometry in prediagnostic plasma from 484 pancreatic cancer case patients enrolled in four prospective cohort studies. Association of metabolites with survival was evaluated using Cox proportional hazards models adjusted for age, cohort, race/ethnicity, cancer stage, fasting time, and diagnosis year. After multiple-hypothesis testing correction, a P value of .0006 or less (.05/82) was considered statistically significant. Based on the results, we evaluated 33 tagging single-nucleotide polymorphisms (SNPs) in the ACO1 gene, requiring a P value of less than .002 (.05/33) for statistical significance. All statistical tests were two-sided., Results: Two metabolites in the tricarboxylic acid (TCA) cycle--isocitrate and aconitate--were statistically significantly associated with survival. Participants in the highest vs lowest quintile had hazard ratios (HRs) for death of 1.89 (95% confidence interval [CI] = 1.06 to 3.35, Ptrend < .001) for isocitrate and 2.54 (95% CI = 1.42 to 4.54, Ptrend < .001) for aconitate. Isocitrate is interconverted with citrate via the intermediate aconitate in a reaction catalyzed by the enzyme aconitase 1 (ACO1). Therefore, we investigated the citrate to aconitate plus isocitrate ratio and SNPs in the ACO1 gene. The ratio was strongly associated with survival (P trend < .001) as was the SNP rs7874815 in the ACO1 gene (hazard ratio for death per minor allele = 1.37, 95% CI = 1.16 to 1.61, P < .001). Patients had an approximately three-fold hazard for death when possessing one or more minor alleles at rs7874851 and high aconitate or isocitrate., Conclusions: Prediagnostic circulating levels of TCA cycle intermediates and inherited ACO1 genotypes were associated with survival among patients with pancreatic cancer., (© The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Evidence remains inconclusive about the association of systemic inflammatory markers with colorectal neoplasia. We investigated whether circulating inflammatory markers were associated with risk of advanced colorectal adenoma. We measured plasma macrophage inhibitory cytokine-1 (MIC-1), C-reactive protein (CRP), interleukin-6 (IL6), and soluble TNF receptor 2 (sTNFR-2) in blood samples drawn from 32,826 women in 1989 to 1990 in the Nurses' Health Study. Through 2008, we documented 757 cases of advanced colorectal adenomas (≥1 cm or any size with advanced histology); each case was matched by age and time of blood draw with one control randomly selected from participants who underwent lower endoscopy and did not have neoplasia. Plasma MIC-1 was associated with higher risk of advanced adenoma (Ptrend = 0.04), with an OR of 1.55 (95% confidence interval, 1.03-2.32) comparing extreme quintiles of MIC-1 after adjusting for colorectal cancer-risk factors and other inflammatory markers. Among cases, MIC-1 level was positively associated with the number of adenomas (P < 0.001) and gradually increased from adenomas located in the rectum, distal colon, and up to the proximal colon. There was a strong positive association between MIC-1 and risk of adenomas with multiplicity, ≥1 cm size and location in the proximal colon (all Ptrend < 0.05). CRP, IL6, or sTNFR-2 was not associated with adenoma risk. In conclusion, plasma MIC-1 was associated with higher risk of colorectal adenoma, especially multiple, large, and proximal adenomas. Our results provide further support for a role for MIC-1 in carcinogenesis and the potential for MIC-1 as an adjunctive biomarker for detection of advanced colorectal adenoma., (©2015 American Association for Cancer Research.)

Circulating adiponectin is inversely related to the risk of colorectal cancer. However, its influence on colorectal cancer survival is unclear. We conducted a prospective study to evaluate the association between prediagnostic plasma levels of adiponectin and mortality in patients with colorectal cancer. We identified 621 incident colorectal cancer cases who provided blood specimens prior to diagnosis within the Nurses' Health Study (NHS) and Health Professionals Follow-up Study (HPFS). Cox proportional hazards models were used to calculate HRs and 95% confidence intervals (CI). After a median follow-up of 9 years, there were 269 (43%) total deaths, of which 181 (67%) were due to colorectal cancer. Compared with participants in the lowest quartile of adiponectin, those in the highest quartile had multivariate HRs of 1.89 (95% CI, 1.21-2.97; P(trend) = 0.01) for colorectal cancer-specific mortality and 1.66 (95% CI, 1.15-2.39; P(trend) = 0.009) for overall mortality. The apparent increased risk in colorectal cancer-specific mortality was more pronounced in patients with metastatic disease (HR, 3.02: 95% CI, 1.50-6.08). Among patients with colorectal cancer, prediagnostic plasma adiponectin is associated with an increased risk of colorectal cancer-specific and overall mortality and is more apparent in patients with metastatic disease. Adiponectin may be a marker for cancers which develop through specific pathways that may be associated with worsened prognosis. Further studies are needed to validate these findings., (©2015 American Association for Cancer Research.)

Background: Low levels of adiponectin (ADIPOQ; HGNC ID; HGNC:13633), an adipokine, are associated with obesity, adiposity, excess energy balance, and increased risk of colorectal neoplasia. Given the reported association of increased body mass index (BMI) and low-level physical activity with KRAS-mutated colorectal tumor, we hypothesized that low-level plasma adiponectin might be associated with increased risk of KRAS-mutant colorectal carcinoma but not with risk of KRAS wild-type carcinoma., Methods: We conducted molecular pathological epidemiology research using a nested case-control study design (307 incident rectal and colon cancer case patients and 593 matched control individuals) within prospective cohort studies, the Nurses' Health Study (152 case patients and 297 control individuals, with blood collection in 1989-1990) and the Health Professionals Follow-up Study (155 case patients and 296 control individuals, with blood collection in 1993-1995). Multivariable conditional logistic regression models and two-sided likelihood ratio tests were used to assess etiologic heterogeneity of the associations., Results: The association of low-level plasma adiponectin with colorectal cancer risk statistically significantly differed by KRAS mutation status (P heterogeneity = .004). Low levels of plasma adiponectin were associated with KRAS-mutant colorectal cancer (for the lowest vs highest tertile: multivariable odds ratio [OR] = 2.83, 95% confidence interval [CI] = 1.50 to 5.34, P trend = .002) but not with KRAS wild-type cancer (for the lowest vs highest tertile: multivariable OR = 0.83, 95% CI = 0.49 to 1.43, P trend = .48). In secondary analyses, the association between plasma adiponectin and colorectal cancer did not appreciably differ by BRAF or PIK3CA oncogene mutation status., Conclusions: Low-level plasma adiponectin is associated with KRAS-mutant colorectal cancer risk but not with KRAS wild-type cancer risk., (© The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Background & Aims: Patients with colorectal cancer (CRC) have high circulating levels of macrophage inhibitory cytokine-1 (MIC1 or growth differentiation factor 15), a marker of inflammation that might be involved in carcinogenesis. We analyzed blood samples collected from individuals before they were diagnosed with CRC to determine whether levels of MIC1 were associated with mortality., Methods: We collected data on survival of 618 participants diagnosed with CRC who provided prediagnosis blood specimens in 1990 (Nurses' Health Study) and 1994 (Health Professionals' Follow-up Study) and were followed through 2010. Levels of MIC1 were measured by enzyme-linked immunosorbent assay and then were categorized into quartiles based on the known distribution of MIC1 levels among previously matched individuals without CRC (controls) within each cohort. We then examined the association of MIC1 levels with overall and CRC-specific mortality using Cox proportional hazards models, with adjustments for mortality-associated risk factors and other plasma markers of inflammation. We also assessed the relationship between levels of MIC1 and levels of prostaglandin-endoperoxide synthase 2 expression (PTGS2 or cyclooxygenase-2), measured in 245 tumor samples by immunohistochemistry., Results: Compared with participants in the lowest quartile for plasma level of MIC1, the multivariate hazard ratio for CRC-specific death for participants in the highest quartile of MIC1 level was 2.40 (95% confidence interval: 1.33-4.34; P for linear trend = .009). The association of MIC1 with survival varied with level of PTGS2 expression in tumor samples (Pinteraction = .04). For individuals with PTGS2-positive tumors, the hazard ratio for CRC-specific death among those with high levels of MIC1 (equal to or greater than the median) was 2.13 (95% confidence interval: 0.99-4.58) compared with participants with low levels of MIC1 (below the median). In individuals with PTGS2-negative CRC, a high level of MIC1 was not associated with an increased risk of CRC-specific death (multivariate hazard ratio = 0.61; 95% confidence interval: 0.13-2.93)., Conclusions: Based on an analysis of blood and colorectal tumor samples from 2 large studies, high plasma levels of MIC1 (growth differentiation factor 15) before diagnosis of CRC are associated with greater CRC-specific mortality, particularly in individuals with PTGS2-positive tumors., (Copyright © 2015 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Background & Aims: Risk for colorectal cancer (CRC) can be greatly reduced through screening. To aid in the development of screening strategies, we refined models designed to determine risk of CRC by incorporating information from common genetic susceptibility loci., Methods: By using data collected from more than 12,000 participants in 6 studies performed from 1990 through 2011 in the United States and Germany, we developed risk determination models based on sex, age, family history, genetic risk score (number of risk alleles carried at 27 validated common CRC susceptibility loci), and history of endoscopic examinations. The model was validated using data collected from approximately 1800 participants in the Prostate, Lung, Colorectal, and Ovarian Cancer Screening Trial, conducted from 1993 through 2001 in the United States., Results: We identified a CRC genetic risk score that independently predicted which patients in the training set would develop CRC. Compared with determination of risk based only on family history, adding the genetic risk score increased the discriminatory accuracy from 0.51 to 0.59 (P = .0028) for men and from 0.52 to 0.56 (P = .14) for women. We calculated age- and sex-specific 10-year CRC absolute risk estimates based on the number of risk alleles, family history, and history of endoscopic examinations. A model that included a genetic risk score better determined the recommended starting age for screening in subjects with and without family histories of CRC. The starting age for high-risk men (family history of CRC and genetic risk score, 90%) was 42 years, and for low-risk men (no family history of CRC and genetic risk score, 10%) was 52 years. For men with no family history and a high genetic risk score (90%), the starting age would be 47 years; this is an intermediate value that is 5 years earlier than it would be for men with a genetic risk score of 10%. Similar trends were observed in women., Conclusions: By incorporating information on CRC risk alleles, we created a model to determine the risk for CRC more accurately. This model might be used to develop screening and prevention strategies., (Copyright © 2015 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Background: Colorectal carcinoma (CRC) represents a group of histopathologically and molecularly heterogeneous diseases, which may contain signet-ring cell component and/or mucinous component to a varying extent under pathology assessment. However, little is known about the prognostic significance of those components, independent of various tumor molecular features., Methods: Utilizing a molecular pathological epidemiology database of 1,336 rectal and colon cancers in the Nurses' Health Study and the Health Professionals Follow-up Study, we examined patient survival according to the proportion of signet-ring cell and mucinous components in CRCs. Cox proportional hazards models were used to compute hazard ratio (HR) for mortality, adjusting for potential confounders including stage, microsatellite instability, CpG island methylator phenotype, LINE-1 methylation, and KRAS, BRAF, and PIK3CA mutations., Results: Compared to CRC without signet-ring cell component, 1-50 % signet-ring cell component was associated with multivariate CRC-specific mortality HR of 1.40 [95 % confidence interval (CI) 1.02-1.93], and >50 % signet-ring cell component was associated with multivariate CRC-specific mortality HR of 4.53 (95 % CI 2.53-8.12) (P trend < 0.0001). Compared to CRC without mucinous component, neither 1-50 % mucinous component (multivariate HR 1.04; 95 % CI 0.81-1.33) nor >50 % mucinous component (multivariate HR 0.82; 95 % CI 0.54-1.23) was significantly associated with CRC-specific mortality (P trend < 0.57)., Conclusions: Even a minor (50 % or less) signet-ring cell component in CRC was associated with higher patient mortality, independent of various tumor molecular and other clinicopathological features. In contrast, mucinous component was not associated with mortality in CRC patients.

Background: Smoothened, frizzled family receptor (SMO) is an important component of the hedgehog signaling pathway, which has been implicated in various human carcinomas. However, clinical, molecular, and prognostic associations of SMO expression in colorectal cancer remain unclear., Methods: Using a database of 735 colon and rectal cancers in the Nurse's Health Study and the Health Professionals Follow-up Study, we examined the relationship of tumor SMO expression (assessed by immunohistochemistry) to prognosis, and to clinical, pathological, and tumor molecular features, including mutations of KRAS, BRAF, and PIK3CA, microsatellite instability, CpG island methylator phenotype (CIMP), LINE-1 methylation, and expression of phosphorylated AKT and CTNNB1., Results: SMO expression was detected in 370 tumors (50 %). In multivariate logistic regression analysis, SMO expression was independently inversely associated with phosphorylated AKT expression [odds ratio (OR) 0.48; 95 % confidence interval (CI) 0.34-0.67] and CTNNB1 nuclear localization (OR 0.48; 95 % CI 0.35-0.67). SMO expression was not significantly associated with colorectal cancer-specific or overall survival. However, in CIMP-high tumors, but not CIMP-low/0 tumors, SMO expression was significantly associated with better colorectal cancer-specific survival (log-rank P = 0.012; multivariate hazard ratio, 0.36; 95 % CI 0.13-0.95; P interaction = 0.035, for SMO and CIMP status)., Conclusions: Our data reveal novel potential associations between the hedgehog, the WNT/CTNNB1, and the PI3K (phosphatidylinositol-4,5-bisphosphonate 3-kinase)/AKT pathways, supporting pivotal roles of SMO and hedgehog signaling in pathway networking. SMO expression in colorectal cancer may interact with tumor CIMP status to affect patient prognosis, although confirmation by future studies is needed.

Of more than 150 000 published studies evaluating new biomarkers, fewer than 100 biomarkers have been implemented for patient care. One reason for this is lack of rigorous testing by the medical community to validate claims for biomarker clinical relevance, and potential reluctance to publish negative results when confirmation is not obtained. Here we sought to determine the utility and reproducibility of immunohistochemical detection of hepatocyte growth factor (HGF) in melanoma tissue, an approach of potential assistance in defining patients with innate resistance to BRAF inhibitor therapy. To this end, a published and a revised method that retained sensitivity but with greater specificity for HGF detection, were evaluated in cells known to endogenously express HGF, and in models where HGF is upregulated via cytokine induction and via overexpression by gene transfection. Consequent patient evaluation in collaboration with the Melanoma Institute Australia of a cohort of 41 melanoma specimens with extensive clinical annotation failed to validate HGF immunohistochemistry as a predictor of response to BRAF inhibitors. Targeted therapies for advanced melanoma and other cancers show great promise, and rigorous validation studies are thus indicated for approaches that seek to personalize such therapies to maximize therapeutic efficacy.

Aim: To investigate insulin-like growth factor 2 (IGF2) differentially methylated region (DMR)0 hypomethylation in relation to clinicopathological and molecular features in colorectal serrated lesions., Methods: To accurately analyze the association between the histological types and molecular features of each type of serrated lesion, we consecutively collected 1386 formalin-fixed paraffin-embedded tissue specimens that comprised all histological types [hyperplastic polyps (HPs, n = 121), sessile serrated adenomas (SSAs, n = 132), traditional serrated adenomas (TSAs, n = 111), non-serrated adenomas (n = 195), and colorectal cancers (CRCs, n = 827)]. We evaluated the methylation levels of IGF2 DMR0 and long interspersed nucleotide element-1 (LINE-1) in HPs (n = 115), SSAs (n = 120), SSAs with cytological dysplasia (n = 10), TSAs (n = 91), TSAs with high-grade dysplasia (HGD) (n = 15), non-serrated adenomas (n = 80), non-serrated adenomas with HGD (n = 105), and CRCs (n = 794). For the accurate quantification of the relative methylation levels (scale 0%-100%) of IGF2 DMR0 and LINE-1, we used bisulfite pyrosequencing method. Tumor specimens were analyzed for microsatellite instability, KRAS (codons 12 and 13), BRAF (V600E), and PIK3CA (exons 9 and 20) mutations; MLH1 and MGMT methylation; and IGF2 expression by immunohistochemistry., Results: The distribution of the IGF2 DMR0 methylation level in 351 serrated lesions and 185 non-serrated adenomas (with or without HGD) was as follows: mean 61.7, median 62.5, SD 18.0, range 5.0-99.0, interquartile range 49.5-74.4. The IGF2 DMR0 methylation level was divided into quartiles (Q1 ≥ 74.5, Q2 62.6-74.4, Q3 49.6-62.5, Q4 ≤ 49.5) for further analysis. With regard to the histological type, the IGF2 DMR0 methylation levels of SSAs (mean ± SD, 73.1 ± 12.3) were significantly higher than those of HPs (61.9 ± 20.5), TSAs (61.6 ± 19.6), and non-serrated adenomas (59.0 ± 15.8) (P < 0.0001). The IGF2 DMR0 methylation level was inversely correlated with the IGF2 expression level (r = -0.21, P = 0.0051). IGF2 DMR0 hypomethylation was less frequently detected in SSAs compared with HPs, TSAs, and non-serrated adenomas (P < 0.0001). Multivariate logistic regression analysis also showed that IGF2 DMR0 hypomethylation was inversely associated with SSAs (P < 0.0001). The methylation levels of IGF2 DMR0 and LINE-1 in TSAs with HGD (50.2 ± 18.7 and 55.7 ± 5.4, respectively) were significantly lower than those in TSAs (61.6 ± 19.6 and 58.8 ± 4.7, respectively) (IGF2 DMR0, P = 0.038; LINE-1, P = 0.024)., Conclusion: IGF2 DMR0 hypomethylation may be an infrequent epigenetic alteration in the SSA pathway. Hypomethylation of IGF2 DMR0 and LINE-1 may play a role in TSA pathway progression.

Prostaglandin E2 (PGE2) promotes colorectal carcinogenesis. Overall, systemic PGE2 production can be assessed by measuring its major metabolite, PGE-M, in urine. We examined the potential role of PGE-M as a biomarker for colorectal adenoma risk and chemopreventive response to anti-inflammatory drugs. We conducted a prospective case-control study nested within the Nurses' Health Study. Among women who previously provided a urine sample, we identified 420 cases diagnosed with colorectal adenoma during follow-up and matched them to 420 endoscopy-negative controls. We measured urinary PGE-M using an LC/MS assay. Compared with women in the lowest quartile of urinary PGE-M, women in the highest quartile had a multivariate OR of 1.40 (95% confidence interval (CI), 0.92-2.14) for any adenoma; 0.91 (95% CI, 0.48-1.72) for low-risk adenoma (solitary adenoma <1 cm in greatest diameter with tubular/unspecified histology); and 1.66 (95% CI, 1.04-2.67) for high-risk adenoma (adenoma ≥1 cm in greatest diameter and/or tubulovillous, villous or high-grade dysplasia histology or multiple adenomas of any size or histology). Regular use of anti-inflammatory drugs (≥2 standard tablets of aspirin/NSAIDs per week) was associated with a significant reduction in adenoma risk (multivariate OR, 0.61; 95% CI, 0.43-0.87) in women with high baseline PGE-M (quartiles 2-4), but not low PGE-M (quartile 1).Urinary PGE-M is associated with an increased risk of high-risk adenoma. Anti-inflammatory drugs seem to reduce adenoma risk among women with high, but not low PGE-M. Urinary PGE-M may serve as a biomarker to define subsets of the population who may obtain differential chemopreventive benefit from anti-inflammatory drugs., (©2014 American Association for Cancer Research.)

The American Joint Committee on Cancer/Union Internationale Contre le Cancer (AJCC/UICC) TNM staging system provides the most reliable guidelines for the routine prognostication and treatment of colorectal carcinoma. This traditional tumour staging summarizes data on tumour burden (T), the presence of cancer cells in draining and regional lymph nodes (N) and evidence for distant metastases (M). However, it is now recognized that the clinical outcome can vary significantly among patients within the same stage. The current classification provides limited prognostic information and does not predict response to therapy. Multiple ways to classify cancer and to distinguish different subtypes of colorectal cancer have been proposed, including morphology, cell origin, molecular pathways, mutation status and gene expression-based stratification. These parameters rely on tumour-cell characteristics. Extensive literature has investigated the host immune response against cancer and demonstrated the prognostic impact of the in situ immune cell infiltrate in tumours. A methodology named 'Immunoscore' has been defined to quantify the in situ immune infiltrate. In colorectal cancer, the Immunoscore may add to the significance of the current AJCC/UICC TNM classification, since it has been demonstrated to be a prognostic factor superior to the AJCC/UICC TNM classification. An international consortium has been initiated to validate and promote the Immunoscore in routine clinical settings. The results of this international consortium may result in the implementation of the Immunoscore as a new component for the classification of cancer, designated TNM-I (TNM-Immune)., (© 2013 The Authors. Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.)

Adipokines are adipocyte-secreted hormones that may mediate the etiologic link between obesity and colorectal cancer; however, the evidence from large prospective studies is limited. We prospectively evaluated the association of plasma adiponectin and soluble leptin receptor (sOB-R) with colorectal cancer risk within the Nurses' Health Study (1990-2008) and the Health Professionals Follow-up Study (1994-2008) among 616 incident colorectal cancer cases and 1,205 controls selected using risk-set sampling and matched on age and date of blood draw. In unconditional logistic regression with adjustment for matching factors and multiple risk factors, plasma adiponectin was significantly associated with reduced risk of colorectal cancer among men, but not among women. Compared with men in the lowest quartile of adiponectin, men in the highest quartile had a relative risk (RR) for colorectal cancer of 0.55 [95% confidence interval (CI), 0.35-0.86; Ptrend = 0.02]. The corresponding RR in women was 0.96 (95% CI, 0.67-1.39; Ptrend = 0.74). Plasma sOB-R was not associated with overall colorectal cancer risk in either men or women. A significant heterogeneity was noted in the association between sOB-R and colorectal cancer by subsite in women (Pheterogeneity = 0.004); sOB-R was significantly associated with increased risk of rectal cancer but not colon cancer. These findings support a role for adiponectin in colorectal carcinogenesis in men. Further studies are warranted to confirm these associations and elucidate potential underlying mechanisms.

Background: Higher levels of physical activity are associated with lower colorectal carcinoma incidence and mortality, perhaps through influencing energy balance, cellular prosta7 systemic inflammation. Although evidence suggests interactive effects of energetics, sedentary lifestyle, and tumor CTNNB1 (β-catenin) or CDKN1B (p27) status on colon cancer prognosis, interactive effects of physical activity and tumor PTGS2 (the official symbol for COX-2) status on clinical outcome remain unknown., Methods: Using molecular pathological epidemiology database of 605 stage I-III colon and rectal cancers in two prospective cohort studies (the Nurse's Health Study and the Health Professionals Follow-up Study), we examined patient survival according to postdiagnosis physical activity and tumor PTGS2 status (with 382 PTGS2-positive and 223 PTGS2-negative tumors by immunohistochemistry). Cox proportional hazards models were used to calculate colorectal cancer-specific mortality HR, adjusting for clinical and other tumor variables including microsatellite instability status., Results: Among PTGS2-positive cases, compared with the least active first quartile, the multivariate HRs (95% confidence interval) were 0.30 (0.14-0.62) for the second, 0.38 (0.20-0.71) for the third, and 0.18 (0.08-0.41) for the fourth quartile of physical activity level (Ptrend = 0.0002). In contrast, among PTGS2-negative cases, physical activity level was not significantly associated with survival (Ptrend = 0.84; Pinteraction = 0.024, between physical activity and tumor PTGS2 status)., Conclusions: Postdiagnosis physical activity is associated with better survival among patients with PTGS2-positive tumors but not among patients with PTGS2-negative tumors., Impact: Immunohistochemical PTGS2 expression in colorectal carcinoma may serve as a predictive biomarker in pathology practice, which may predict stronger benefit from exercise.

Chronic inflammation may play a role in the development of pancreatic cancer. However, few prospective studies have examined the association between plasma inflammatory markers and pancreatic cancer risk. Therefore, we investigated the association of prediagnostic circulating C-reactive protein (CRP), interleukin-6 (IL-6), and TNF-α receptor II (TNF-αR2) with subsequent pancreatic cancer risk in a prospective, nested case-control study of 470 cases and 1,094 controls from Health Professionals Follow-up Study, Nurses' Health Study, Physicians' Health Study, Women's Health Initiative, and Women's Health Study. The median follow-up time of cases was 7.2 years (range 1-26 years). No association was observed between plasma CRP, IL-6, and TNF-αR2 and the risk of pancreatic cancer. Comparing extreme quintiles, the multivariate ORs were 1.10 [95% confidence interval (CI), 0.74-1.63; Ptrend = 0.81] for CRP, 1.19 (95% CI, 0.81-1.76; Ptrend = 0.08) for IL-6, and 0.88 (95% CI, 0.58-1.33; Ptrend = 0.57) for TNF-αR2. In conclusion, prediagnostic levels of circulating CRP, IL-6, and TNF-αR2 were not associated with the risk of pancreatic cancer, suggesting that systemic inflammation as measured by circulating inflammatory factors is unlikely to play a major role in the development of pancreatic cancer.

The incidence of esophageal adenocarcinoma (EAC) has risen 600% over the last 30 years. With a 5-year survival rate of ~15%, the identification of new therapeutic targets for EAC is greatly important. We analyze the mutation spectra from whole-exome sequencing of 149 EAC tumor-normal pairs, 15 of which have also been subjected to whole-genome sequencing. We identify a mutational signature defined by a high prevalence of A>C transversions at AA dinucleotides. Statistical analysis of exome data identified 26 significantly mutated genes. Of these genes, five (TP53, CDKN2A, SMAD4, ARID1A and PIK3CA) have previously been implicated in EAC. The new significantly mutated genes include chromatin-modifying factors and candidate contributors SPG20, TLR4, ELMO1 and DOCK2. Functional analyses of EAC-derived mutations in ELMO1 identifies increased cellular invasion. Therefore, we suggest the potential activation of the RAC1 pathway as a contributor to EAC tumorigenesis.

The minor allele (G) of rs4939827, a SMAD7 (18q21) intronic variant, is associated with a lower risk of developing colorectal cancer (CRC) and poorer survival after diagnosis. Our objective was to evaluate the associations of this variant with different tumor phenotype and intratumoral molecular characteristics. We evaluated 1509 CRC cases and 2307 age-matched controls nested within the Nurses' Health Study and the Health Professionals Follow-up Study. We used the TaqMan assay to genotype rs4939827 and logistic regression to assess the association of rs4939827 with risk of CRC according to different phenotypic and molecular characteristics. We found that the minor allele (G) in rs4939827 (SMAD7, 18q21) was associated with a lower risk of developing tumor stage pT1 or pT2 CRC [multivariate odds ratio (OR), 0.73; 95% confidence interval (CI) 0.62-0.87] but not tumor stage pT3 or pT4 (multivariate OR, 1.07; 95% CI 0.93-1.23, P for heterogeneity = 1.2 × 10(-4)). The association between rs4939827 and CRC also significantly differed by methylation of RUNX3 (P for heterogeneity = 0.005). Among those with CRC, the minor allele (G) in rs4939827 was significantly associated with poorer overall survival (hazards ratio, 1.20; 95% CI, 1.02-1.42). We can conclude that the minor allele (G) of the germline intronic SMAD7 variant rs4939827 is associated with a lower risk of CRC with earlier tumor stage and CRC without methylation of the tumor suppressor RUNX3. These findings suggest that individuals with this SMAD7 variant that develop CRC are more probably to have tumors with greater invasiveness and methylation of RUNX3, which potentially contributes to their poorer observed survival.

Background: Evidence suggests that insulin-like growth factor 2 messenger RNA binding protein 3 (IGF2BP3, also known as IMP3) represents a promising cancer biomarker. However, the clinical, pathological, molecular and prognostic features of IGF2BP3-positive colorectal cancers remain uncertain., Materials and Methods: We evaluated IGF2BP3 expression by immunohistochemistry in 671 rectal and colon cancer cases that form part of a molecular pathological epidemiology database. Cox proportional hazards regression models were used to compute mortality hazard ratio (HR), adjusting for clinical, pathological and molecular features, including microsatellite instability, the CpG island methylator phenotype, LINE-1 methylation and KRAS, BRAF and PIK3CA mutations., Results: Among 671 colorectal cancers, 234 (35%) tumours were positive for IGF2BP3. In contrast, normal colorectal epithelium was negative for IGF2BP3 in all 403 specimens of normal mucosa adjacent to carcinoma. IGF2BP3 positivity was associated with poor differentiation (p=0.0003), stage III-IV disease (p=0.0081), BRAF mutation (p=0.031), and LINE-1 hypomethylation (p=0.020). IGF2BP3 positivity was significantly associated with shorter colorectal cancer-specific [log-rank p<0.0001; multivariate HR, 1.37; 95% confidence interval (CI), 1.02-1.84] and overall survival (log-rank p=0.0004; multivariate HR, 1.32; 95% CI, 1.05-1.66)., Conclusions: IGF2BP3 expression in colorectal cancer is associated with adverse clinical outcome. Our findings support a role for IGF2BP3 as a diagnostic and/or prognostic biomarker in colorectal cancer., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Background: Regular use of aspirin after a diagnosis of colon cancer has been associated with a superior clinical outcome. Experimental evidence suggests that inhibition of prostaglandin-endoperoxide synthase 2 (PTGS2) (also known as cyclooxygenase-2) by aspirin down-regulates phosphatidylinositol 3-kinase (PI3K) signaling activity. We hypothesized that the effect of aspirin on survival and prognosis in patients with cancers characterized by mutated PIK3CA (the phosphatidylinositol-4,5-bisphosphonate 3-kinase, catalytic subunit alpha polypeptide gene) might differ from the effect among those with wild-type PIK3CA cancers., Methods: We obtained data on 964 patients with rectal or colon cancer from the Nurses' Health Study and the Health Professionals Follow-up Study, including data on aspirin use after diagnosis and the presence or absence of PIK3CA mutation. We used a Cox proportional-hazards model to compute the multivariate hazard ratio for death. We examined tumor markers, including PTGS2, phosphorylated AKT, KRAS, BRAF, microsatellite instability, CpG island methylator phenotype, and methylation of long interspersed nucleotide element 1., Results: Among patients with mutated-PIK3CA colorectal cancers, regular use of aspirin after diagnosis was associated with superior colorectal cancer-specific survival (multivariate hazard ratio for cancer-related death, 0.18; 95% confidence interval [CI], 0.06 to 0.61; P<0.001 by the log-rank test) and overall survival (multivariate hazard ratio for death from any cause, 0.54; 95% CI, 0.31 to 0.94; P=0.01 by the log-rank test). In contrast, among patients with wild-type PIK3CA, regular use of aspirin after diagnosis was not associated with colorectal cancer-specific survival (multivariate hazard ratio, 0.96; 95% CI, 0.69 to 1.32; P=0.76 by the log-rank test; P=0.009 for interaction between aspirin and PIK3CA variables) or overall survival (multivariate hazard ratio, 0.94; 95% CI, 0.75 to 1.17; P=0.96 by the log-rank test; P=0.07 for interaction)., Conclusions: Regular use of aspirin after diagnosis was associated with longer survival among patients with mutated-PIK3CA colorectal cancer, but not among patients with wild-type PIK3CA cancer. The findings from this molecular pathological epidemiology study suggest that the PIK3CA mutation in colorectal cancer may serve as a predictive molecular biomarker for adjuvant aspirin therapy. (Funded by The National Institutes of Health and others.).

Background: Infiltrative growth pattern at the tumor margin has been associated with shorter patient survival. However, little is known about the prognostic significance of tumor growth pattern, independent of tumoral molecular alterations and other histologic features., Methods: Utilizing a database of 1139 colon and rectal cancer patients in two prospective cohort studies, histologic features including tumor growth pattern, tumor differentiation, lymphocytic reaction, mucinous component, and signet ring cell component were recorded by a single pathologist. Cox proportional hazard model was used to compute mortality hazard ratio, adjusting for clinical, pathologic, and tumor molecular features, including microsatellite instability, the CpG island methylator phenotype, long interspersed nucleotide element 1 (LINE-1) methylation, and KRAS, BRAF, and PIK3CA mutations., Results: Among 1139 colorectal cancers, we observed expansile growth pattern in 372 tumors (33%), intermediate growth pattern in 610 tumors (54%), and infiltrative growth pattern in 157 tumors (14%). Compared to patients with expansile growth pattern, those with infiltrative growth pattern experienced shorter cancer-specific survival (log rank P < 0.0001; multivariate hazard ratio 1.74; 95% confidence interval 1.22-2.47) and overall survival (log rank P < 0.0001; multivariate hazard ratio 1.78; 95% confidence interval 1.33-2.39). The prognostic association of infiltrative growth pattern was confined to patients with stage I-III disease (P (interaction) with stage = 0.0001)., Conclusions: Infiltrative growth pattern was associated with worse prognosis among stage I-III colorectal cancer patients, independent of other clinical, pathologic, and molecular characteristics.

The molecular analysis of biomarkers in oncology is rapidly advancing, but the incorporation of new molecular tests into clinical practice will require a greater understanding of the genetic changes that drive malignancy, the assays used to measure the resulting phenotypes and genotypes, and the regulatory processes that new molecular biomarkers must face to be accepted for clinical use. To address these issues and provide an overview of current molecular testing in 6 major malignancies, including glioma, breast cancer, colon cancer, lung cancer, prostate cancer, and acute myelogenous leukemia, an NCCN Task Force was convened on the topic of evaluating the clinical utility of tumor markers in oncology. The output of this meeting, contained within this report, describes the ways biomarkers have been developed and used; defines common terminology, including prognostic, predictive, and companion diagnostic markers, and analytic validity, clinical validity, and clinical utility; and proposes the use of a combination level of evidence score to aid in the evaluation of novel biomarker tests as they arise. The current state of regulatory oversight and anticipated changes in the regulation of molecular testing are also addressed.

Background & Aims: The insulin-like growth factor 2 (IGF2) gene is normally imprinted. Constitutive loss of imprinting (LOI) of IGF2 has been associated with increased risks of colon cancer and adenoma, indicating its role in carcinogenesis. The conventional LOI assay relies on a germline polymorphism to distinguish between 2 allelic expression patterns but results in many uninformative cases. IGF2 LOI correlates with hypomethylation at the differentially methylated region (DMR)-0. An assay for methylation of the DMR0 could overcome the limitations of the conventional IGF2 LOI assay., Methods: We measured methylation at the IGF2 DMR0 using a bisulfite-pyrosequencing assay with 1178 paraffin-embedded colorectal cancer tissue samples from 2 prospective cohort studies. A Cox proportional hazard model was used to calculate mortality hazard ratio (HR); calculations were adjusted for microsatellite instability; the CpG island methylator phenotype; LINE-1 methylation; and KRAS, BRAF, and PIK3CA mutations., Results: Methylation at the IGF2 DMR0 was successfully measured in 1105 (94%) of 1178 samples. Colorectal tumors had significantly less methylation at the DMR0 compared with matched, normal colonic mucosa (P < .0001; N = 51). Among 1033 patients eligible for survival analysis, hypomethylation of the IGF2 DMR0 was significantly associated with higher overall mortality (log-rank P = .0006; univariate HR, 1.41; 95% confidence interval, 1.16-1.71; P = .0006; multivariate HR, 1.33; 95% confidence interval, 1.08-1.63; P = .0066)., Conclusions: A bisulfite-pyrosequencing assay to measure methylation of the IGF2 DMR0 is robust and applicable to paraffin-embedded tissue. IGF2 DMR0 hypomethylation in colorectal tumor samples is associated with shorter survival time, so it might be developed as a prognostic biomarker., (Copyright © 2010 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Background and Aims: Proteases play a critical role in tumorigenesis and are upregulated in colorectal cancer and neoplastic polyps. In animal models, cathepsin B (CTSB)-activatable imaging agents show high enzyme activity within intestinal tumors., Methods: We conducted a prospective cohort study of 558 men and women with colon cancer with tumors that were accessible for immunohistochemical assessment. We used Cox proportional hazards models, stratified by stage, to compute colon cancer-specific and overall mortality according to tumoral expression of CTSB., Results: Among 558 participants, 457 (82%) had tumors that expressed CTSB (CTSB positive) and 101 (18%) had tumors that did not express CTSB (CTSB negative). CTSB expression was not associated with disease stage (P = 0.19). After a median follow-up of 11.6 years, there were 254 total and 155 colon cancer-specific deaths. Compared with participants with CTSB-negative tumors, participants with CTSB-positive tumors experienced a multivariate hazard ratio for colon cancer-specific mortality of 1.99 (95% confidence interval, 1.19-3.34) and overall mortality of 1.71 (95% confidence interval, 1.16-2.50). CTSB expression was independently associated with KRAS (P = 0.01) and BRAF mutation (P = 0.04), but not microsatellite instability status, CpG island methylator phenotype status, PIK3CA mutation, LINE-1 methylation, TP53 expression, or PTGS2 (cyclooxygenase-2) expression. Among 123 individuals with adenomas, 91% expressed CTSB., Conclusions: As assessed by immunohistochemistry, CTSB is expressed in the vast majority of colon cancers, independent of stage, and is significantly associated with higher risk of colon cancer-specific and overall mortality., Impact: These results support the potential of CTSB a target for image detection of neoplastic lesions in humans., (©2010 AACR.)

Background: Genome-wide DNA hypomethylation plays a role in genomic instability and carcinogenesis. LINE-1 (L1 retrotransposon) constitutes a substantial portion of the human genome, and LINE-1 methylation correlates with global DNA methylation status. LINE-1 hypomethylation in colon cancer has been strongly associated with poor prognosis. However, whether LINE-1 hypomethylators constitute a distinct cancer subtype remains uncertain. Recent evidence for concordant LINE-1 hypomethylation within synchronous colorectal cancer pairs suggests the presence of a non-stochastic mechanism influencing tumor LINE-1 methylation level. Thus, it is of particular interest to examine whether its wide variation can be attributed to clinical, pathologic or molecular features., Design: Utilizing a database of 869 colorectal cancers in two prospective cohort studies, we constructed multivariate linear and logistic regression models for LINE-1 methylation (quantified by Pyrosequencing). Variables included age, sex, body mass index, family history of colorectal cancer, smoking status, tumor location, stage, grade, mucinous component, signet ring cells, tumor infiltrating lymphocytes, CpG island methylator phenotype (CIMP), microsatellite instability, expression of TP53 (p53), CDKN1A (p21), CTNNB1 (beta-catenin), PTGS2 (cyclooxygenase-2), and FASN, and mutations in KRAS, BRAF, and PIK3CA., Results: Tumoral LINE-1 methylation ranged from 23.1 to 90.3 of 0-100 scale (mean 61.4; median 62.3; standard deviation 9.6), and distributed approximately normally except for extreme hypomethylators [LINE-1 methylation < 40; N = 22 (2.5%), which were far more than what could be expected by normal distribution]. LINE-1 extreme hypomethylators were significantly associated with younger patients (p = 0.0058). Residual plot by multivariate linear regression showed that LINE-1 extreme hypomethylators clustered as one distinct group, separate from the main tumor group. The multivariate linear regression model could explain 8.4% of the total variability of LINE-1 methylation (R-square = 0.084). Multivariate logistic regression models for binary LINE-1 hypomethylation outcomes (cutoffs of 40, 50 and 60) showed at most fair predictive ability (area under receiver operator characteristics curve < 0.63)., Conclusions: LINE-1 extreme hypomethylators appear to constitute a previously-unrecognized, distinct subtype of colorectal cancers, which needs to be confirmed by additional studies. Our tumor LINE-1 methylation data indicate enormous epigenomic diversity of individual colorectal cancers.

Purpose: Physical activity in colon cancer survivors has been associated with lower cancer recurrences and improved survival. Whether molecular features of the tumor portend more or less likelihood for benefit from exercise is unknown., Experimental Design: Using two large prospective cohort studies with physical activity assessments after colon cancer diagnosis, we examined expression of fatty acid synthase, p53, p21, and p27 and mutational status of K-ras and phosphatidylinositol 3-kinase(PI3KCA). We calculated hazard ratios (HR) of colon cancer-specific mortality, adjusted for tumor and patient characteristics, and tested for molecular interactions with exercise., Results: In a cohort of 484 men and women with stage I to III colon cancer, patients who engaged in at least 18 metabolic equivalent task (MET)-hours per week after diagnosis had an adjusted HR for colon cancer-specific mortality of 0.64 [95% confidence interval (95% CI), 0.33-1.23] and for overall mortality of 0.60 (95% CI, 0.41-0.86). A statistically significant interaction was detected based on p27 expression (P = 0.03). For tumors with loss of p27 (n = 195), physical activity of > or =18 MET-hours/week led to a HR for colon cancer mortality of 1.40 (95% CI, 0.41-4.72), compared with those with <18 MET-hours/week. However, for tumors with expression of p27 (n = 251), the adjusted HR was 0.33 (95% CI, 0.12-0.85). Molecular status of fatty acid synthase, K-ras, p53, p21, and PI3KCA did not influence the association between exercise and colon cancer-specific or overall mortality., Conclusion: The benefit of physical activity on outcomes in patients with stage I to III colon cancer may be influenced by p27 status. Further studies are warranted to confirm these findings.

Context: Aspirin reduces risk of colorectal neoplasia in randomized trials and inhibits tumor growth and metastases in animal models. However, the influence of aspirin on survival after diagnosis of colorectal cancer is unknown., Objective: To examine the association between aspirin use after colorectal cancer diagnosis on colorectal cancer-specific and overall survival., Design, Setting, and Participants: Prospective cohort study of 1279 men and women diagnosed with stage I, II, or III colorectal cancer. Participants were enrolled in 2 nationwide health professional cohorts in 1980 and 1986 prior to diagnosis and followed up through June 1, 2008., Main Outcome Measure: Colorectal cancer-specific and overall mortality., Results: After a median follow-up of 11.8 years, there were 193 total deaths (35%) and 81 colorectal cancer-specific deaths (15%) among 549 participants who regularly used aspirin after colorectal cancer diagnosis, compared with 287 total deaths (39%) and 141 colorectal cancer-specific deaths (19%) among 730 participants who did not use aspirin. Compared with nonusers, participants who regularly used aspirin after diagnosis experienced a multivariate hazard ratio (HR) for colorectal cancer-specific mortality of 0.71 (95% confidence interval [CI], 0.53-0.95) and for overall mortality of 0.79 (95% CI, 0.65-0.97). Among 719 participants who did not use aspirin before diagnosis, aspirin use initiated after diagnosis was associated with a multivariate HR for colorectal cancer-specific mortality of 0.53 (95% CI, 0.33-0.86). Among 459 participants with colorectal cancers that were accessible for immunohistochemical assessment, the effect of aspirin differed significantly according to cyclooxygenase 2 (COX-2) expression (P for interaction = .04). Regular aspirin use after diagnosis was associated with a lower risk of colorectal cancer-specific mortality among participants in whom primary tumors overexpressed COX-2 (multivariate HR, 0.39; 95% CI, 0.20-0.76), whereas aspirin use was not associated with lower risk among those with primary tumors with weak or absent expression (multivariate HR, 1.22; 95% CI, 0.36-4.18)., Conclusion: Regular aspirin use after the diagnosis of colorectal cancer is associated with lower risk of colorectal cancer-specific and overall mortality, especially among individuals with tumors that overexpress COX-2.

Methylation of CpG islands in gene promoter regions is a major molecular mechanism of gene silencing and underlies both cancer development and progression. In molecular oncology, testing for the CpG methylation of tissue DNA has emerged as a clinically useful tool for tumor detection, outcome prediction, and treatment selection, as well as for assessing the efficacy of treatment with the use of demethylating agents and monitoring for tumor recurrence. In addition, because CpG methylation occurs early in pre-neoplastic tissues, methylation tests may be useful as markers of cancer risk in patients with either infectious or inflammatory conditions. The Methylation Working Group of the Clinical Practice Committee of the Association of Molecular Pathology has reviewed the current state of clinical testing in this area. We report here our summary of both the advantages and disadvantages of various methods, as well as the needs for standardization and reporting. We then conclude by summarizing the most promising areas for future clinical testing in cancer molecular diagnostics.

AURKA (the official symbol for Aurora-A, STK15, or BTAK) regulates the function of centrosomes, spindles, and kinetochores for proper mitotic progression. AURKA overexpression is observed in various cancers including colon cancer, and a link between AURKA and chromosomal instability (CIN) has been proposed. However, no study has comprehensively examined AURKA expression in relation to CIN or prognosis using a large number of tumors. Using 517 colorectal cancers in two prospective cohort studies, we detected AURKA overexpression (by immunohistochemistry) in 98 tumors (19%). We assessed other molecular events including loss of heterozygosity (LOH) in 2p, 5q, 17q, and 18q, the CpG island methylation phenotype (CIMP), and microsatellite instability (MSI). Prognostic significance of AURKA was evaluated by Cox regression and Kaplan-Meier method. In both univariate and multivariate logistic regressions, AURKA overexpression was significantly associated with CIN (defined as the presence of LOH in any of the chromosomal segments; multivariate odds ratio, 2.97; 95% confidence interval, 1.40-6.29; P = .0045). In multivariate analysis, AURKA was associated with cyclin D1 expression (P = .010) and inversely with PIK3CA mutation (P=.014), fatty acid synthase expression (P=.028), and family history of colorectal cancer (P = .050), but not with sex, age, body mass index, tumor location, stage, CIMP, MSI, KRAS, BRAF, BMI, LINE-1 hypomethylation, p53, p21, beta-catenin, or cyclooxygenase 2. AURKA was not significantly associated with clinical outcome or survival. In conclusion, AURKA overexpression is independently associated with CIN in colorectal cancer, supporting a potential role of Aurora kinase-A in colorectal carcinogenesis through genomic instability (rather than epigenomic instability).

Background & Aims: The peroxisome proliferator-activated receptor gamma (PPARG, PPARgamma) is a nuclear receptor that regulates expression of mediators of lipid metabolism and the inflammatory response. There is controversy over the pro-oncogenic or antioncogenic effects of PPARG, and little is known about its prognostic significance in colon cancer., Methods: Among 470 patients with colorectal cancer (stages I-IV) identified in 2 independent prospective cohorts, PPARG expression was detected in 102 tumors (22%) by immunohistochemistry. Cox proportional hazards models were used to compute hazard ratios (HRs) of colorectal cancer-specific and overall mortalities, adjusted for patient characteristics and molecular features including cyclooxygenase 2, fatty acid synthase, KRAS, BRAF, PIK3CA, p53, p21, beta-catenin, LINE-1 hypomethylation, microsatellite instability (MSI), and the CpG island methylation phenotype (CIMP)., Results: Compared with patients with PPARG-negative tumors, patients with PPARG-positive tumors had significantly lower overall mortality, determined by Kaplan-Meier analysis (P=.0047), univariate Cox regression (HR, 0.55; 95% confidence interval [CI], 0.37-0.84; P=.0053), and multivariate analysis (adjusted HR, 0.43; 95% CI, 0.27-0.69; P=.0004). Patients with PPARG-positive tumors experienced lower colorectal cancer-specific mortality (adjusted HR, 0.44; 95% CI, 0.25-0.79; P=.0054). The relationship between PPARG and lower mortality did not appear to be significantly modified by MSI, CIMP, LINE-1, or the other clinical and molecular variables examined (all P(interaction)>.05)., Conclusions: Tumor expression of PPARG is independently associated with longer survival of patients. PPARG expression appears to mark an indolent subset of colorectal cancers.

Purpose: PIK3CA mutation and subsequent activation of the AKT pathway play an important role in colorectal carcinogenesis. However, little is known about the prognostic role of PIK3CA mutation in colon cancer., Patients and Methods: Using 450 resectable colon cancers (stage I to III) in two independent prospective cohorts, we detected PIK3CA mutation in 82 tumors (18%) by pyrosequencing. Cox proportional hazards models were used to calculate hazard ratios (HRs) of colon cancer-specific and overall mortalities, adjusted for patient characteristics and tumoral molecular features, including the CpG island methylation phenotype, microsatellite instability (MSI), LINE-1 hypomethylation, and p53, CIMP, KRAS and BRAF mutation., Results: Compared with patients with PIK3CA wild-type tumors, those with PIK3CA-mutated tumors experienced an increase in colon cancer-specific mortality according to univariate analysis (HR = 1.64; 95% CI, 0.95 to 2.86), which persisted after adjusting for other known or potential risk factors for cancer recurrence (including MSI; multivariate HR = 2.23; 95% CI, 1.21 to 4.11). The effect of PIK3CA mutation on cancer survival seemed to differ according to KRAS mutational status. Among patients with KRAS wild-type tumors, the presence of PIK3CA mutation was associated with a significant increase in colon cancer-specific mortality (HR = 3.80; 95% CI, 1.56 to 9.27). In contrast, PIK3CA mutation conferred no significant effect on mortality among patients with KRAS-mutated tumors (HR = 1.25; 95% CI, 0.52 to 2.96)., Conclusion: Among patients who undergo a curative resection of colon cancer, PIK3CA mutation is associated with shorter cancer-specific survival. The adverse effect of PIK3CA mutation may be potentially limited to patients with KRAS wild-type tumors.