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Overweight and obesity have been linked to increased risk of several diseases, including colorectal cancer, but the underlying mechanisms are not fully known. An earlier study analyzed metabolically defined body size phenotypes in relation to colorectal cancer risk, using combinations of C-peptide (a marker of insulin resistance) and body mass index (BMI). Higher serum C-peptide concentrations were associated with higher colorectal cancer risk in both overweight and normal weight individuals compared to individuals with low levels of C-peptide and normal weight. The aim of the present study was to see if these results could be replicated and to further explore the role of metabolism and body size phenotypes in colorectal cancer subtypes. We conducted a nested case-control study of 1010 individuals with colorectal cancer and 1010 individually matched (age, sex, cohort, year of blood sampling and data collection, number of freeze-thaw cycles of plasma samples and fasting status at blood sampling) control participants from the population-based Northern Sweden Health and Disease Study. The blood samples and data used in our analyses were collected at a mean of 9.7 years prior to case diagnosis. Participants were categorized as (1) metabolically healthy/normal weight (BMI < 25 kg/m2), (2) metabolically healthy/overweight (BMI ≥ 25 kg/m2), (3) metabolically unhealthy/normal weight (BMI < 25 kg/m2), or (4) metabolically unhealthy/overweight (BMI ≥ 25 kg/m2), where metabolically healthy was defined as C-peptide in the lowest tertile, as in the previous study. We used multivariable conditional logistic regression to calculate odds ratios and confidence intervals and adjusted for tobacco smoking, recreational physical activity and alcohol consumption as potential confounders. In our preliminary results, metabolically unhealthy/overweight individuals had a significantly higher risk of developing colorectal cancer compared to metabolically healthy/normal weight individuals (adjusted odds ratio =1.44, 95% CI 1.13-1.85). The odds ratio for metabolically unhealthy/normal weight individuals was 1.23 (95% CI 0.92-1.65) and for metabolically healthy/overweight individuals 1.23 (95% CI 0.89-1.70). These results support the potential of a more nuanced analysis of metabolism and body size to better understand their etiological contribution to colorectal cancer development. To gain more knowledge about how the role of metabolism and body size might differ between tumor subtypes, we will also analyze body size phenotypes in relation to subtypes of colorectal cancer based on anatomical tumor site, KRAS and BRAF mutations and microsatellite instability status of the tumor. Citation Format: Linda Vidman, Simon Knekta, Björn Gylling, Carl Zingmark, Anna Löfgren-Burström, Richard Palmqvist, Sophia Harlid, Bethany Van Guelpen. Metabolically defined body size phenotypes in relation to subsequent colorectal cancer risk [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr LB142.

Despite advances in treatment, colorectal cancer remains one of the leading causes of cancer-specific death. To decrease colorectal cancer mortality, better and more cost-efficient tools for risk prediction and early detection are needed. One area that has shown promise is using protein biomarkers, measured in blood, to detect individuals with early-stage disease or at increased risk. The primary aim of this study was therefore to identify novel protein biomarkers associated with colorectal cancer risk and the secondary aim was to validate protein biomarkers that had been described in previous studies. We included data and samples from 185 incident colorectal cancer cases and 185 controls (matched on age, sex, sampling date, and the number of freeze-thaw cycles) from the Northern Sweden Health and Disease Study (NSHDS). Samples were collected between six years and three months prior to the diagnosis date of each case and concentrations of 1536 individual proteins were measured in plasma samples using the Olink Explore 1536 platform (Olink Proteomics). Prior to statistical analyses, 249 proteins were excluded, either due to not passing quality control or having more than 50% of the concentrations below the limit of detection. In addition, we also excluded extreme outliers. Conditional logistic regression was used to estimate Odds Ratios (ORs) and 95% confidence intervals (95% CI) for the association between protein concentrations and colorectal cancer risk. Multivariable models were adjusted for predetermined colorectal cancer risk factors, including body mass index, alcohol consumption, physical activity, smoking status, and level of education. When considering p Citation Format: Justin Harbs, Matilda Rentoft, Xijia Liu, Beatrice S. Melin, Sophia Harlid, Bethany Van Guelpen. Identifying colorectal cancer biomarkers using a targeted proteomics platform with extensive coverage [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3023.

To gain further understanding of why some colorectal cancer patients develop sarcopenia (loss of skeletal muscle mass) we conducted a study of metabolic and inflammatory markers in pre-diagnostic samples from a well-defined patient cohort. The study population included 755 colorectal cancer patients from population-based prospective cohorts in northern Sweden. Eleven markers were measured in plasma samples collected prior to diagnosis (median 10.1 years), and their relation to sarcopenia at diagnosis (median 7 days before surgery) was assessed. The markers included were adiponectin, total cholesterol, C-peptide, CRP, acyl ghrelin, insulin, leptin, PYY, triglycerides and plasma glucose (both fasting and 2 hours after oral glucose tolerance test). Patients were classified as either sarcopenic or non-sarcopenic based on skeletal muscle index from peri-diagnostic CT scans. Skeletal muscle index was calculated by segmentation of skeletal muscle area at the transaxial level of the third lumbar vertebrae, adjusted to body height. Due to sex differences in muscle mass, all calculations were performed separately in men and women. Multivariable logistic regression was used to estimate associations between pre-diagnostic biomarker concentrations and the risk of sarcopenia. We made adjustment for plausible confounders including age at sampling, smoking, alcohol consumption, energy intake, physical activity, and pre-diagnostic body mass index (BMI). Our preliminary results indicate an association between higher pre-diagnostic cholesterol levels and increased risk of sarcopenia in men (age-adjusted odds ratio 1.30 95% CI 1.06 - 1.59), but not women. This association remained after adjusting for potential confounders. In women, higher levels of adiponectin were associated with an increased sarcopenia risk (age-adjusted OR 1.57 95% CI 1.02 - 2.41). This association remained when adjusting for most potential confounders and was attenuated and no longer statistically significant when BMI was added. Among the other biomarkers no clear associations with sarcopenia risk were observed. In conclusion, we performed a hypothesis-generating study of pre-diagnostic, circulating metabolic and inflammatory markers in relation to radiologically estimated sarcopenia at diagnosis in colorectal cancer patients. Positive associations between cholesterol and sarcopenia in men and between adiponectin and sarcopenia in women were shown. These results need to be validated in future studies. Citation Format: Johannes Färdeman, Björn Gylling, Richard Palmqvist, Jan Axelsson, Katrine Riklund, Sophia Harlid, Bethany Van Guelpen. Relationship between pre-diagnostic biomarkers and sarcopenia in colorectal cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 733.

Protective associations of fruits, vegetables, and fiber intake with colorectal cancer risk have been shown in many, but not all epidemiologic studies. One possible reason for study heterogeneity is that dietary factors may have distinct effects by colorectal cancer molecular subtypes. Here, we investigate the association of fruit, vegetables, and fiber intake with four well-established colorectal cancer molecular subtypes separately and in combination. Nine observational studies including 9,592 cases with molecular subtypes for microsatellite instability (MSI), CpG island methylator phenotype (CIMP), and somatic mutations in BRAF and KRAS genes, and 7,869 controls were analyzed. Both case-only logistic regression analyses and polytomous logistic regression analyses (with one control set and multiple case groups) were used. Higher fruit intake was associated with a trend toward decreased risk of BRAF-mutated tumors [OR 4th vs. 1st quartile = 0.82 (95% confidence interval, 0.65–1.04)] but not BRAF-wildtype tumors [1.09 (0.97–1.22); P difference as shown in case-only analysis = 0.02]. This difference was observed in case–control studies and not in cohort studies. Compared with controls, higher fiber intake showed negative association with colorectal cancer risk for cases with microsatellite stable/MSI-low, CIMP-negative, BRAF-wildtype, and KRAS-wildtype tumors (Ptrend range from 0.03 to 3.4e-03), which is consistent with the traditional adenoma-colorectal cancer pathway. These negative associations were stronger compared with MSI-high, CIMP-positive, BRAF-mutated, or KRAS-mutated tumors, but the differences were not statistically significant. These inverse associations for fruit and fiber intake may explain, in part, inconsistent findings between fruit or fiber intake and colorectal cancer risk that have previously been reported. Significance: These analyses by colorectal cancer molecular subtypes potentially explain the inconsistent findings between dietary fruit or fiber intake and overall colorectal cancer risk that have previously been reported.

Background: Evidence suggests that, compared to later-onset colorectal cancer (CRC), early-onset CRC (diagnosed before 50 years of age) occur more frequently in rectal location and less frequently in proximal colon location. Studies also indicate heterogeneity of molecular characteristics within early-onset CRCs and between early-onset and later-onset CRCs. Understanding how tumor molecular features and biological aggressiveness of early-onset CRC differ by detailed tumor location is important in personalized patient management. Methods: Using 14,004 CRC cases including 3,089 early-onset cases from The Cancer Genome Atlas and the Genetics and Epidemiology of CRC Consortium (GECCO), we examined microsatellite instability (MSI), CpG island methylator phenotype (CIMP), and KRAS and BRAF mutations in cancers of the cecum, ascending colon, transverse colon, descending colon, sigmoid colon, and rectum. We further conducted CRC-specific survival analyses of detailed tumor location stratified by molecular characteristics using multivariable Cox proportional hazards models. Results: The proportions of MSI-high, CIMP-high, and BRAF-mutated early-onset tumors gradually increased from the rectum (8.8%, 3.4%, and 3.5%, respectively) to the ascending colon (46% MSI-high; 15% CIMP-high) or transverse colon (8.6% BRAF-mutated), followed by declines in the cecum for MSI-high (36%) and BRAF-mutated (4.6%) tumors. Notably, later-onset MSI-high tumors showed a continuous decrease in KRAS mutation prevalence from the rectum (36%) to ascending colon (9%; Ptrend Citation Format: Tomotaka Ugai, Yin Cao, Koichiro Haruki, Tabitha A. Harrison, Daniel D. Buchanan, Peter T. Campbell, Andrew T. Chan, Steven Gallinger, Marc J. Gunter, Michael Hoffmeister, Mark A. Jenkins, Roger L. Milne, Polly A. Newcomb, Rish K. Pai, Andrew J. Pellatt, Robert E. Schoen, Bethany Van Guelpen, Michael O. Woods, Amanda I. Phipps, Ulrike Peters, Shuji Ogino. Molecular characteristics and prognosis of early-onset colorectal cancer according to detailed anatomical locations: Comparison to later-onset cases [abstract]. In: Proceedings of the AACR Special Conference on Colorectal Cancer; 2022 Oct 1-4; Portland, OR. Philadelphia (PA): AACR; Cancer Res 2022;82(23 Suppl_1):Abstract nr A010.

Colorectal cancer (CRC) is the third most common cancer worldwide and its incidence is rising. Identifying etiologic pathways and early detection for CRC are key to prevention and reducing mortality as the prognosis for advanced CRC, despite recent improvements in therapy, remains poor. One area of potentially measurable CRC risk factors, that is still relatively unexplored, are circulating levels of endogenous sex hormones. The incidence of CRC is greater in men than in women and a proposed hypothesis for this sex disparity is that higher levels of endogenous estrogens found in women may be protective against CRC. In addition, androgens, including testosterone, have been hypothesized to confer protective effects for CRC in men. However, few studies have evaluated the association between testosterone and CRC in men, and the limited available data suggest that any observed protective association is likely confined to colon rather than rectal cancer. The aim of this study was therefore to evaluate the relation of colon cancer risk with circulating levels of sex hormones and sex hormone binding globulin (SHBG) in a male population. Preliminary analyses included data from 461 cases of incident colon cancer and 461 matched controls from the European Prospective Investigation into Cancer and Nutrition (EPIC) cohort. Sex hormones (estradiol, estrone, testosterone, androstenedione, dehydroepiandrosterone, and progesterone) were measured in prediagnostic plasma samples using liquid chromatography-tandem /mass spectrometry (LC/-MS/MS), and SHBG was measured using a solid phase “sandwich” immunoradiometric assay. To test potential associations between circulating levels of sex hormones and SHBG with the risk of developing colon cancer, conditional logistic regression models were applied to estimate odds ratios (OR) and 95% confidence intervals (95% CI). Multivariable models were adjusted for colon cancer risk factors, including anthropometric parameters, alcohol consumption, physical activity and smoking status. In addition, models including estrone, estradiol and testosterone were adjusted for SHBG and vice versa. Preliminary results indicated an inverse association for testosterone (multivariable OR per SD: 0.80 95% CI: 0.69 - 0.93) and SHBG (multivariable OR per SD: 0.84 95% CI: 0.72 - 0.99) with CRC risk. However, after additional adjustments to SHBG and sex hormone models respectively these associations were attenuated towards the null. No associations were identified for estradiol, estrone, androstenedione, dehydroepiandrosterone, and progesterone. These preliminary results do not support an association between endogenous estrogen and colon cancer in men, while a potential association between testosterone and SHBG and colon cancer will need to be further evaluated. Citation Format: Justin Harbs, Sabina Rinaldi, Audrey Gicquiau, Xijia Liu, Marc Gunter, Bethany Van Guelpen, Neil Murphy, Sophia Harlid, Epic Network. Associations between circulating sex hormone levels and risk of colon cancer among men from six European countries [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr LB-152.

Colorectal cancer (CRC) mortality is highly dependent on stage at diagnosis, and measures to increase the likelihood of early detection are urgently needed. Many countries have ongoing screening programs, but the addition of blood-based pre-screening biomarkers for risk stratification could reduce costs and increase compliance. In this study, we used prospectively collected plasma samples from the population-based Northern Sweden Health and Disease Study (NSHDS) to evaluate panels of protein biomarkers in relation to CRC risk. Using a two-tiered approach, we first included a unique set of 58 CRC cases and 58 matched controls with time-matched, repeated, prediagnostic samples (10 years apart, with the second sample collected three months to five years before diagnosis). Relative plasma concentrations of 161 proteins were analyzed by multiplex immunoassay (Olink proteomics, inflammation panel and oncology II panel). In the second tier, 15 of these proteins were selected for further analysis, together with six promising early detection protein markers from the literature. The second sample set included prediagnostic samples from 450 CRC cases and 450 matched controls, of which 33 case/control pairs had repeated samples. Conditional logistic regression and linear mixed models were used to estimate odds ratios for CRC risk. In the second-tier analyses, Lasso regression was used to develop multi-marker models, and the resulting protein combinations were tested using ROC analysis. Of the 15 proteins showing promise as predictive biomarkers in the first phase, none retained statistical significance in the second phase. The most promising marker candidate was FGF-21 (multivariable OR: 1.14 95% CI: 0.99-1.30), a metabolic hormone involved in stimulation of glucose uptake that has been associated with body mass index. FGF-21 was also the only protein consistently selected by the Lasso algorithm. Sensitivity analysis for FGF-21 in subgroups based on tumor site, stage and molecular subtypes including BRAF and KRAS mutations did not reach statistical significance. Our findings highlight the challenge of identifying cancer biomarkers that can be used in the prediagnostic window of opportunity for early detection. For early risk stratification, with the vision of achieving effective precision screening, single biomarkers or small marker panels may not be sufficient. Large studies using prospectively collected samples and machine learning to identify biomarker risk patterns may be a more promising approach. Citation Format: Sophia Harlid, Justin Harbs, Robin Myte, Carl Brunius, Marc Gunter, Xijia Liu, Richard Palmqvist, Bethany Van Guelpen. A two-tiered targeted proteomics approach to identify biomarkers of colorectal cancer risk [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2353.

Background: Antibiotic use has been hypothesized to modify colorectal cancer (CRC) risk by altering the composition of gut microbiota, with some suggestive evidence reported. The aim of the current study was to investigate the possible association between antibiotic use and CRC risk, taking into consideration daily dose, antibiotic type, and tumor location. Methods: This is a nation-wide, registry-based study with a matched case-control design. All primary CRC cases diagnosed during 2010 to 2016 were selected from the Swedish Colorectal Cancer Register to ensure availability of antibiotic prescription data and a minimum of 5 years of follow-up. Five cancer-free controls were matched to each case for age, sex, and county using the Register of the Total Population. Defined daily doses of antibiotics were extracted from the Swedish Prescribed Drug Register (with complete data from 2005), antibiotic prophylaxis use was estimated using surgical procedural codes from the Swedish National Patient Register, and potential confounders related to socioeconomic status (level of education, country of birth and marital status) were obtained from the LISA database. Multivariable conditional logistic regression was performed to investigate associations between antibiotic use (divided into 5 categories ranging from non-use to very high use (>6months)) and CRC risk. Results: Our study included 40,545 cases and 202,720 matched controls. We identified a positive, linear association between antibiotic use and CRC risk (adjusted OR for very high users vs non-users = 1.11 (95% CI: 1.03-1.19), p-trend=0.014), which was pronounced for proximal colon cancer (OR = 1.42 (95% CI: 1.28-1.58), p-trend = 1.20 × 10−15), less robust for distal colon and null for rectal cancer. The association was strongest for broad-spectrum beta lactams (adjusted OR =1.50 (95% CI, 1.01-2.23), p-trend=3.12 × 10−6). In contrast, Hiprex, an antibiotic used for treatment and prevention of urinary tract infections, with no known effect on gut microbiota, was not associated with CRC risk (OR = 0.92 (95% CI, 0.81-1.05), p-trend=0.217). Conclusion: In this registry-based study covering the entire population of Sweden, we identified a robust association between antibiotic use, particularly broad-spectrum beta-lactams, and an increased risk of colon cancer, particularly proximal colon cancer, and a null relationship for rectal cancer. The null findings for Hiprex, which does not affect gut microbiota, lend support to the microbiota hypothesis for the putative influence of antibiotic use in colonic carcinogenesis. Citation Format: Sai San Moon Lu, Zahraa Mohammed, Christel Häggström, Robin Myte, Elisabeth Lindquist, Åsa Gylfe, Bethany Van Guelpen, Sophia Harlid. Antibiotic use and risk of colorectal cancer: A Swedish population-based registry study [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1055.

Background and Objectives: While there is a clear association between many lifestyle-related factors and colorectal cancer (CRC) risk, the protective associations of fruit, vegetables, and fiber intake have been shown in many, but not all epidemiological studies. One possible reason for study heterogeneity is that dietary factors may have distinct effects by CRC molecular subtypes. To explore this hypothesis, we investigated the association between fruit, vegetables, and fiber intake and four well-established CRC molecular characteristics. Methods: We analyzed 9 observational studies including 8,783 CRC cases with molecular tumor markers for microsatellite instability (MSI), the CpG island methylator phenotype (CIMP), and somatic mutations in BRAF (V600E) and KRAS (codon 12 or 13), and 7,869 controls. We used case-only and polytomous logistic regression analyses to assess the association between dietary intake of fruit, vegetables, and fiber with CRC molecular subtypes defined by each marker separately and in combination. Results: We found that higher fruit intake was associated with a decreased risk of BRAF-mutated tumors [Odds ratio (OR) 4th vs. 1st quartile = 0.72 (95% confidence interval (CI) = 0.57-0.91)], but not with BRAF-wildtype tumor [OR 4th vs. 1st quartile = 1.01 (95%CI = 0.90-1.13); p-difference = 3e-03]. Higher fiber intake showed significant negative association with MSS/MSI-low, CIMP-negative, BRAF-wildtype, and KRAS-wildtype when using these markers separately and in combination (p-trend range from 2e-03 to 4e-05). These negative associations were stronger compared with MSI-high, CIMP-positive, BRAF-mutated or KRAS-mutated tumors, but the differences were not statistically significant. Conclusion: Higher fruit intake may be associated with a decreased risk of BRAF-mutated CRC but not with BRAF-wildtype CRC. Additionally, higher fiber intake may be associated with a decreased risk of MSS/MSI-low, CIMP-negative, BRAF-wildtype, and KRAS-wildtype tumors. These results may explain in part the inconsistent findings between fruit or fiber intake and overall CRC risk that have previously been reported. Citation Format: Akihisa Hidaka, Harrison A. Tabitha, Daniel D. Buchanan, Michael Hoffmeister, Marc J. Gunter, Xiaoliang Wang, Yi Lin, Wei Sun, Syed H. Zaidi, Martha L. Slattery, Bethany Van Guelpen, Steven J. Gallinger, Michael O. Woods, Mark A. Jenkins, Shuji Ogino, Polly A. Newcomb, Peter T. Campbell, Li Hsu, Ulrike Peters. Intake of dietary fruit, vegetables, and fiber and risk of colorectal cancer according to molecular subtypes: A pooled analysis [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4652.

Background: Higher folate intake has been reported to be associated with modestly lower risk of colorectal cancer, but the overall state of the evidence is inconclusive. Revisiting putative and established lifestyle-related risk factors from the perspective of intertumoral heterogeneity is warranted, as risk relationships for a molecular subtype may be attenuated toward the null when colorectal cancer is investigated as a single disease. Aim: To investigate folate and folic acid intakes in relation to the risk of molecular subtypes of colorectal cancer. Methods: We pooled individual-level observational data from 7542 colorectal cancer cases and 7066 controls within the collaborative framework of the Genetics and Epidemiology of Colorectal Cancer Consortium (GECCO) and the Colon Cancer Family Registry (CCFR). Odds ratios per sex- and study-specific quartile increase in dietary and total folate intake, and for folic acid supplement use (yes/no), were estimated using logistic regression for case-only analyses and multinomial models for case-control analyses. Minimally adjusted analyses included sex, age, study and total energy intake as covariates. Tumor marker variables included microsatellite instability (MSI) status, CpG island methylator phenotype (CIMP), and BRAF and KRAS mutations. Results: In case-only analyses, we observed no heterogeneity in associations between folate intake, with or without supplemental folic acid (taking into consideration folic acid fortification when relevant), or with folic acid supplement use, and the risk of any subtype of colorectal cancer based on individual molecular tumor markers (lowest p for heterogeneity 0.073). In case-control analyses, higher dietary and total folate intake and folic acid supplement use were associated with a lower risk of most molecular tumor subtypes (all odds ratios were below one, and most were statistically significant). Adjustment for a larger set of potential confounders had no material effect on risk estimates. Conclusion: In this large, pooled analysis, higher dietary and total folate intakes and folic acid supplement use were all associated with a lower risk of colorectal cancer, regardless of individual molecular tumor markers including MSI status, CIMP, and BRAF and KRAS mutations. Citation Format: Bethany Van Guelpen, Björn Gylling, Sophia Harlid, Anna Winkvist, Hermann Brenner, Daniel D. Buchanan, Peter T. Campbell, Andrew T. Chan, Jenny Chang-Claude, Steven J. Gallinger, Graham G. Giles, Marc J. Gunter, Michael Hoffmeister, Li Hsu, Mark A. Jenkins, Roger L. Milne, Polly A. Newcomb, Shuji Ogino, John D. Potter, Conghui Qu, Lori C. Sakoda, Robert E. Schoen, Martha L. Slattery, Mikael O. Woods, Tabitha A. Harrison, Ulrike Peters. Folate and folic acid intake in relation to molecular subtypes of colorectal cancer; a pooled analysis of 7542 cases [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2355.

Background: A diet comprising low fiber intake, high consumption of red meat, and alcohol drinking, are all established risk factors for colorectal cancer (CRC). Identifying metabolic fingerprints associated with data-driven dietary patterns may provide insight into potential mechanisms linking diet and risk of CRC. Objective: The aim of this study was to first identify robust data-driven dietary factors derived from food frequency data and to assess their relationships with CRC risk. Second, to investigate plasma metabolic profiles in relation to CRC-associated diet factors. Methods: We used self-reported food frequency data and fasting plasma samples from 860 incident CRC cases (median 12 years from sampling to diagnosis) individually matched to controls by cohort, sex, age at diagnosis, year of blood draw and data collection (+/-1yr), fasting status and number of sample freeze thaw cycles from the population-based Northern Sweden Health and Disease Study (NSHDS). Diet factors were identified from combined exploratory/confirmatory factor analysis on random half-splits of the dietary data. Factors that appeared reproducible between repeated half-splits were retained, and their associations with CRC risk were assessed by multivariable conditional logistic regression, adjusted for age, BMI, smoking, physical activity, total energy intake, education and marital status. Factors associated with CRC risk were related to LC-MS metabolic profiles using random forest regression. Preliminary Results: We identified 12 dietary factors that were robustly associated with CRC risk: red meat, alcohol, snacks, fish, healthy breakfast, fast food, spread, full fat, low-fat, smoked fish/meat, vegetables, and an additional factor consisting of rice and fruit/syrup soup. Both alcohol (OR 1.16, 95% CI 1.00-1.34, p=0.047, fdr=0.283) and the healthy breakfast factor (OR 0.89, 95%CI 0.80-0.99, p=0.030, fdr=0.283) were associated with CRC in the adjusted model. Of these two factors, alcohol was also weakly associated with the metabolic profile (Q2=0.19). Conclusion: The data-driven factor analysis successfully identified dietary exposures, of which several corresponded to established CRC risk factors. Alcohol and healthy breakfast habits associated with CRC risk after adjustment for potential confounders. We also identified a metabolic profile in plasma potentially linked to alcohol consumption which merits further investigation. Citation Format: Stina Bodén, Rikard Landberg, Sophia Harlid, Ingegerd Johansson, Marc J. Gunter, Richard Palmqvist, Rui Zheng, Bethany Van Guelpen, Carl Brunius. Data-driven dietary patterns and their association with metabolic fingerprints and risk of colorectal cancer - a nested case-control study from Northern Sweden [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr LB-156.

Background: Smoking has been associated with colorectal cancer (CRC) risk; but limited evidence has shown the association between smoking and molecular subtypes of CRC. Methods: We analyzed 9,422 CRC cases and 9,950 controls from 10 observational studies. Multinomial logistic regression analysis was performed to assess the association between sex-study-specific quartiles of pack years of smoking and risk of CRC molecular subtypes, using non-smokers as reference group, adjusting for age, sex, and study. Known oncogenic mutations in four somatic colorectal tumor markers were assessed individually and in combination, including BRAF mutations, KRAS mutations, CpG island methylator phenotype (CIMP), and microsatellite instability (MSI) status. Case-only analysis was also performed to estimate heterogeneity in risk of molecular subtypes of CRC. Results: Compared with controls, higher pack years of smoking were statistically significantly associated with increased CRC risk when stratified, individually, by all four markers, and the associations got stronger with higher quartiles pack years (p-trend Conclusion: In this largest study with a total of 19,372 subjects, we found that heavier pack years of smoking was associated with increased risk of all CRC molecular subtypes. Smokers with heavier pack-years of smoking had particularly higher risk of CRC subtypes with BRAF mutation and CIMP-high, suggesting smoking may be particularly involved in the development of these subtypes of colorectal tumor. Citation Format: Xiaoliang Wang, Efrat Amitay, Barbara L. Banbury, Hermann Brenner, Daniel D. Buchanan, Peter T. Campbell, Jenny Chang-Claude, Steven J. Gallinger, Graham G. Giles, Tabitha A. Harrison, John L. Hopper, Mark A. Jenkins, Yi Lin, Reiko Nishihara, Polly A. Newcomb, Shuji Ogino, Lori C. Sakoda, Robert E. Schoen, Martha L. Slattery, Steven N. Thibodeau, Bethany Van Guelpen, Michael O. Woods, Li Hsu, Michael Hoffmeister, Ulrike Peters. Smoking is associated with risks of molecular subtypes of colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 630.

Over 50 genetic variants have been associated with colorectal cancer (CRC) risk through genome-wide association studies (GWAS), yet these variants represent only a fraction of the total estimated heritability. CRC is a heterogenous disease with diverse tumor etiology. Assessing genetic risk in molecular subtypes may help to identify novel loci and characterize genetic risk among tumor subtypes. We used microsatellite instability (MSI), an established CRC classifier with etiological and therapeutic relevance, to define CRC subtypes for GWAS analyses. We conducted a case-case analysis to estimate odds ratios (OR) and 95% confidence intervals (CI) for association of genome-wide variants with microsatellite stable (MSS) versus unstable (MSI) carcinomas. We ran an inverse-variance weighted fixed-effects meta-analysis across GWAS in a discovery set of 4,163 population-based CRC cases with harmonized microsatellite instability (MSI) marker and imputed genotype data. For each analysis, we used log-additive logistic regression, adjusting for age, sex, and principal components to account for population substructure. We then followed up with replication of 102 SNPs that reached p-values less than 5x10-6 in 1,698 cases. A total of 845 (20.3%) cancer cases were microsatellite unstable in the discovery population and 174 (10.2%) were unstable in the replication population. No variants reached the genome-wide significance level of 5x10-8 in the discovery set. However, we identified two variants that reached a Bonferroni corrected p-value of 4.0x10-4 in the replication set. This included one variant in MLH1 (Replication: OR=1.74, 95% CI=1.53-1.98, p=1.63x10-5; Discovery+Replication: OR=1.45, 95% CI=1.37-1.54, p=9.76x10-11) and one variant in LOC105377645 (Replication: OR=1.70, 95% CI=1.49-1.94, p=5.13x10-5; Discovery+Replication: OR=1.45, 95% CI=1.37-1.54, p=9.76 x 10-11). The MLH1 gene is a DNA mismatch repair gene implicated in Lynch Syndrome, the hallmark of which is microsatellite instability. This is the first genome-wide scan to identify a common variant in MLH1 that is associated with CRC. This variant (minor allele frequency, MAF = 23% in this all European ancestry population) is located in the 5'-untranslated region of MLH1 and is thought to act as a long-range regulator of DCLK3, a potential tumor driver gene. The second variant, located in LOC105377645 with an MAF of 22%, is in an uncharacterized region of the genome and has not previously been implicated in cancer development. These findings suggest that accounting for molecular heterogeneity is important for discovery and characterization of genetic variants associated with CRC risk. We plan to run polytomous regression analyses, increase our sample size, and further investigate CRC subtypes by CIMP, BRAF mutation, KRAS mutation status. Citation Format: Tabitha A. Harrison, Yiwen Lu, Chenjie Zeng, Flora Qu, Kristin Anderson, Hermann Brenner, Daniel D. Buchanan, Peter T. Campbell, Andrew T. Chan, Jenny Chang-Claude, Graham G. Giles, Bethany Van Guelpen, Michael Hoffmeister, Mark A. Jenkins, Noralane M. Lindor, Roger L. Milne, Polly A. Newcomb, Reiko Nishihara, Michael O. Woods, Shuji Ogino, John D. Potter, Martha L. Slattery, Wei Sun, Stephen N. Thibodeau, Li Hsu, Ulrike Peters. Genome-wide association study by colorectal carcinoma subtype [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 229.

Background: Pyridoxal 5’-phospate (PLP) is the most commonly used marker of circulating vitamin B6 status. In prospective case-control studies, individuals with higher plasma concentrations of PLP had a 30-50% decreased risk of colorectal cancer (CRC). This may partly be explained by decreased PLP due to inflammatory activity in persons at an increased risk of CRC. We aimed to investigate different markers of vitamin B6 in relation to CRC risk in a population with a very long follow-up time between screening and diagnosis. Methods: This was a prospective case-control study of 613 incident CRC cases and 1190 matched controls nested within the population-based Northern Sweden Health and Disease Study. PLP, pyridoxal (PL), pyridoxic acid (PA), 3-hydoxykynurenine (HK) and xanthurenic acid (XA) were measured in plasma by LC-MS/MS. Besides PLP, we investigated the recently introduced metabolite ratios PAr (PA/(PLP+PL)) and HK:XA as markers of vitamin B6 status in relation to CRC risk. Results: PAr, a marker of vitamin B6 catabolism during inflammation, was linearly associated with CRC risk, with a 50% higher risk in the highest vs. lowest quartile. In the group with a follow up time between screening and diagnosis over 10.5 years no association was observed, while in the groups with a follow up time between 5.8-10.5 years and below 5.8 years risk was approximately twofold in most quartiles vs the lowest quartile. A 50% risk decrease was observed in the in the third vs the lowest quartile of plasma PLP and a similar risk increase was observed in the highest vs the lowest quartile of the functional vitamin B6 marker HK:XA. Conclusions: PAr may influence later stages of tumor development, but possibly not early tumorigenesis. Poor functional vitamin B6 status and inflammatory activity might influence both the risk of CRC and plasma concentrations of PLP. Our results underscore the need to use several biomarkers for vitamin B6 status together with the already well established plasma PLP. This may be generalizable not only to CRC but also other diseases with an inflammatory aspect. Citation Format: Björn Gylling, Robin Myte, Jörn Schneede, Göran Hallmans, Jenny Häggström, Ingegerd Johansson, Arve Ulvik, Øivind Midttun, Per Magne Ueland, Bethany van Guelpen, Richard Palmqvist. Vitamin B6 biomarkers and colorectal cancer: Modifications by time. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4291.

Background: Colorectal cancer (CRC) is a heterogeneous disease caused by a number of distinct pathways defined by genetic and epigenetic events. These molecular subtypes differ in prognosis and treatment options, and recent studies also imply differences in etiology. One-carbon metabolism has been extensively studied in relation to incident CRC risk because of its involvement in nucleotide synthesis and methylation reactions, yet only a few studies have evaluated if associations differ over CRC subtypes. In this study, we investigated a targeted panel of circulating metabolites and a set of single nucleotide polymorphisms involved in one-carbon metabolism in relation to the risk of overall CRC and CRC subtypes approximated by mutations in the BRAF and KRAS oncogenes, microsatellite instability (MSI), or CpG island methylator phenotype (CIMP). Methods: This was a prospective population-based nested case control study of 613 CRC cases and 1190 matched controls. The data included plasma concentrations of 14 one-carbon metabolites, 17 related single nucleotide polymorphisms, and other lifestyle-related information for all subjects, and clinical and molecular information for the tumors. In order to account for interactions and dependencies between variables, we applied multivariate Bayesian network learning in combination with more traditional univariate multinomial logistic regression to investigate independent relations to subtype specific CRC risk. Results: Preliminary findings show that vitamin B6 (Pyridoxal 5-phosphate, PLP) and vitamin B2 (riboflavin) had the strongest independent relations to overall CRC risk in the multivariate analyses. Folate, glycine, vitamin B6, and the RFC1 80G>A polymorphism displayed signs of independent relations to CRC subtypes. In univariate multinomial logistic regression models, plasma folate expressed the strongest etiologic heterogeneity, and was more clearly related to the risk of CIMP low/high tumors, tumors with MSI, or tumors without KRAS mutation, respectively. Conclusions: This study investigated a large set of prediagnostic circulating metabolites and single nucleotide polymorphisms related to one-carbon metabolism using multivariate statistical learning. We found that some components showed signs of subtype specific relations, of which the association for folate was clearest. Our results suggest that the role of one-carbon metabolism in colorectal carcinogenesis may differ between specific molecular pathways. Citation Format: Robin Myte, Björn Gylling, Jenny Häggström, Jörn Schneede, Per Magne Ueland, Göran Hallmans, Ingegerd Johansson, Richard Palmqvist, Bethany Van Guelpen. One-carbon metabolism and colorectal cancer risk according to molecular subtypes: a Bayesian network learning approach. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4294.