Ugai, Tomotaka

Background: Considering the pathophysiological importance of the intestine-passing microbiota, we hypothesized that the mutational and microbial profiles of colorectal cancer might gradually change along detailed anatomical locations and that these trends might differ by the age of diagnosis and the microsatellite instability (MSI) status. Methods: We assessed somatic mutations and selected bacterial species in colorectal carcinomas along the cecum, ascending colon, transverse colon, descending colon, sigmoid colon, and rectum, utilizing targeted sequencing data ( N = 5685) from the Genetics and Epidemiology of Colorectal Cancer Consortium and tissue microbial data ( N = 1389) from the Health Professionals Follow-up Study and Nurses’ Health Study. Results: There were decreasing trends from the cecum to rectum in the prevalence of gene mutations in the IGF/PI3K, receptor tyrosine kinase (RTK)/RAS, TGFB, and WNT signaling pathways (all P trend < 0.0001), as well as high levels of Bacteroides fragilis ( P trend < 0.0001) and high levels of Fusobacterium nucleatum ( P trend = 0.0029) (but not enterotoxigenic Bacteroides fragilis or pks+ Escherichia coli ). Conversely, TP53 pathway mutations showed an increasing cecum-to-rectum trend ( P trend < 0.0001). Similar to later-onset cases diagnosed at ages 50 years and older, in early-onset cases diagnosed under the age of 50 years, we found generally decreasing cecum-to-rectum trends of IGF/PI3K, RTK/RAS, TGFB, and WNT pathway mutations and an increasing cecum-to-rectum trend in TP53 pathway mutations. By contrast, the cecum-to-rectum prevalence-changing trends in IGF/PI3K, RTK/RAS, TGFB, and TP53 pathway mutations were present in non–MSI-high tumors but not in MSI-high tumors (all P interaction < 0.0052). Conclusion: Non–MSI-high colorectal cancers exhibited decreasing cecum-to-rectum trends of IGF/PI3K, RTK/RAS, and TGFB pathway mutations (and an increasing trend of TP53 pathway mutations), in contrast to MSI-high tumors that demonstrated no such trends. Our study highlights the importance of detailed colorectal subsite information combined with tumor molecular and microbial characteristics in colorectal cancer research.

Abstract Background Physical activity is associated with lower colorectal cancer (CRC) risk, but its association with molecular subtypes defined by genetic and epigenetic alterations of the disease is unclear. Such information may enhance the understanding of the mechanisms related to the benefits of physical activity. Methods Pooled observational (cases: n = 5386; controls: n = 6798; studies n = 5) and genome-wide association data (cases: n = 8178; controls: n = 10 472; studies n = 5) were used. We used multivariable logistic regression models and Mendelian randomization to assess the association between physical activity and the risk of CRC subtypes defined by individual tumor markers (and marker combinations), namely microsatellite instability status, CpG island methylator phenotype status, and BRAF and KRAS mutations. We used case-only analysis to test for differences between molecular subtypes. We applied Bonferroni correction to account for multiple tests. Results In the pooled observational analysis, higher levels of physical activity were associated with lower CRC risk (Obs-per 1SD, odds ratio [OR] = 0.94, 95% confidence interval [CI] = 0.90 to 0.97), with an association that was stronger in males (Obs-per 1SD, OR = 0.91, 95% CI = 0.87 to 0.96) than in females (Obs-per 1SD, OR = 0.97, 95% CI = 0.91 to 1.03; Pinteraction = .04). Higher physical activity was associated with a lower risk of CRC across all molecular subtypes, especially in males. There was no difference in the associations by subtypes by pooled observational or Mendelian randomization analyses. The findings did not differ by study design, anatomical site, and early or late age onset of CRC. Conclusions Our findings suggest that physical activity is not differentially associated with the 4 major molecular subtypes involved in colorectal carcinogenesis, indicating that its benefits extend broadly across colorectal cancer pathogenesis.

Abstract Background: Waist circumference (WC) and its allometric counterpart, “a body shape index” (ABSI), are risk factors for colorectal cancer; however, it is uncertain whether associations with these body measurements are limited to specific molecular subtypes of the disease. Methods: Data from 2,772 colorectal cancer cases and 3,521 controls were pooled from four cohort studies within the Genetics and Epidemiology of Colorectal Cancer Consortium. Four molecular markers (BRAF mutation, KRAS mutation, CpG island methylator phenotype, and microsatellite instability) were analyzed individually and in combination (Jass types). Multivariable logistic and multinomial logistic models were used to assess the associations of WC and ABSI with overall colorectal cancer risk and, in case-only analyses, to evaluate heterogeneity by molecular subtype, respectively. Results: Higher WC (ORper 5 cm = 1.06, 95% confidence interval, 1.04–1.09) and ABSI (ORper 1-SD = 1.07, 95% confidence interval, 1.00–1.14) were associated with elevated colorectal cancer risk. There was no evidence of heterogeneity between the molecular subtypes. No difference was observed regarding the influence of WC and ABSI on the four major molecular markers in proximal colon, distal colon, and rectal cancers, as well as in early- and late-onset colorectal cancers. Associations did not differ in the Jass-type analysis. Conclusions: Higher WC and ABSI were associated with elevated colorectal cancer risk; however, they do not differentially influence all four major molecular mutations involved in colorectal carcinogenesis but underscore the importance of maintaining a healthy body weight in colorectal cancer prevention. Impact: The proposed results have potential utility in colorectal cancer prevention.

Abstract Background: The genotoxin colibactin causes a tumor single-base substitution (SBS) mutational signature, SBS88. It is unknown whether epidemiologic factors’ association with colorectal cancer risk and survival differs by SBS88. Methods: Within the Genetic Epidemiology of Colorectal Cancer Consortium and Colon Cancer Family Registry, we measured SBS88 in 4,308 microsatellite stable/microsatellite instability low tumors. Associations of epidemiologic factors with colorectal cancer risk by SBS88 were assessed using multinomial regression (N = 4,308 cases, 14,192 controls; cohort-only cases N = 1,911), and with colorectal cancer–specific survival using Cox proportional hazards regression (N = 3,465 cases). Results: 392 (9%) tumors were SBS88 positive. Among all cases, the highest quartile of fruit intake was associated with lower risk of SBS88-positive colorectal cancer than SBS88-negative colorectal cancer [odds ratio (OR) = 0.53, 95% confidence interval (CI) 0.37-0.76; OR = 0.75, 95% CI 0.66–0.85, respectively, Pheterogeneity = 0.047]. Among cohort studies, associations of body mass index (BMI), alcohol, and fruit intake with colorectal cancer risk differed by SBS88. BMI ≥30 kg/m2 was associated with worse colorectal cancer–specific survival among those SBS88-positive [hazard ratio (HR) = 3.40, 95% CI 1.47–7.84], but not among those SBS88-negative (HR = 0.97, 95% CI 0.78–1.21, Pheterogeneity = 0.066). Conclusions: Most epidemiologic factors did not differ by SBS88 for colorectal cancer risk or survival. Higher BMI may be associated with worse colorectal cancer–specific survival among those SBS88-positive; however, validation is needed in samples with whole-genome or whole-exome sequencing available. Impact: This study highlights the importance of identification of tumor phenotypes related to colorectal cancer and understanding potential heterogeneity for risk and survival.

Abstract Background: Colorectal cancer (CRC) incidence has increased in those 50 years or younger compared to older individuals in recent decades. While the intra-tumoral microbiome is a potent facilitator of tumorigenesis, the role of the microbiome in early onset (eo)CRC is unclear. Identification of the intra-tumoral microbiome may enhance prediction of adverse outcomes to improve survival of CRC overall age groups and eoCRC. Methods: We performed targeted sequencing of tumor and matched normal DNA samples from 3695 colorectal cancer cases within the Genetics and Colorectal Cancer Consortium (GECCO) and the Colon Cancer Family Registry (CCFR). Tumor DNA was extracted from formalin fixed paraffin embedded (FFPE) or fresh frozen tissue. Normal tissue DNA was extracted from blood or normal surrounding colorectal tissue. We designed probes for the bacterial pathogens enterotoxigenic Bacteroides fragilis (ETBF), polyketide synthase positive E. coli (pks+ EC), and Fusobacterium nucleatum (Fn). We used logistic regression to assess tumor-associated bacteria and clinical parameters, tumor molecular characteristics, and tumor stage adjusted for age, sex, and study and a Cox promotional hazard to assess association with CRC-specific survival, overall and in eoCRC, adjusted for age at diagnosis, sex, mutational burden, and microsatellite instability (MSI) and stratified baseline hazards by study. Survival data were available for 2357 and 308 patients, overall and in eoCRC, respectively. Results: The prevalence of different bacteria in tumors differed slightly overall vs. in eoCRC tumors: ETBF (5.2%, 6.3%), pks+ EC (12.9%, 14.8%), and Fn (10.4%, 8.2%). Overall, men had a lower probability of tumors with Fn (odds ratio (OR)=0.8, 95% confidence interval (CI)=0.64-0.99, p=0.04) but higher odds of tumors with pks+ EC (OR=1.38, CI=1.14-1.68, p=0.001), compared to women. The odds of prevalent Fn were higher for proximal vs distal tumors (OR=1.76, CI=1.38-2.26, p&lt;0.001), but there was no association between tumor site and pks+ EC or ETBF. MSI-high tumors were more likely to carry Fn (OR=2.79, CI=2.08-3.75, p&lt;0.001) and less likely to carry pks+ EC (OR=0.63, CI=0.44-0.88, p=0.008) when compared to patients with MSS tumors. In patients with non-hypermutated tumors, we observed that pks+ EC positive tumors were associated with better CRC-specific survival (HR=0.73, CI=0.57-0.93, p=0.01), whereas ETBF positive tumors (HR=1.99, CI=1,18-3.36,p=0.009) and Fn positive tumors (HR=1.33, CI=1.04-1.72, p=0.03) were associated with poorer survival. In eo-CRC patients, Fn positive status was associated with poorer survival (HR=2.4, CI= 1.056 - 5.492, p=0.026). Conclusions: We showed that the intra-tumoral microbiome differs in distribution across patient populations and patient survival over all age groups and, specifically, in eoCRC. Citation Format: Meredith A. Hullar, Keith R. Curtis, Tabitha Harrison, Yi Lin, Robert Steinfelder, Sonja I. Berndt, Daniel D. Buchanan, Andrew T. Chan, David A. Drew, Jane Figueiredo, Amy J. French, Tabitha A. Harrison, Mark A. Jenkins, Johannes Melaku, Victor Moreno, Tomotaka Ugai, Shuji Ogino, Conghui Qu, Chenxu Qu, Steven Thibodeau, Syed Zaidi, Amanda I. Phipps, Ulrike Peters, on behalf of GECCO and CCFR investigators. Evaluation of intra-tumoral pks+ E. coli, enterotoxigenic B. fragilis and Fusobacterium nucleatum, overall and in early onset disease, in colorectal cancer cases [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 3039.

INTRODUCTION: Early-onset colorectal cancer diagnosed before the age of 50 years has been increasing. Likely reflecting the pathogenic role of the intestinal microbiome, which gradually changes across the entire colorectal length, the prevalence of certain tumor molecular characteristics gradually changes along colorectal subsites. Understanding how colorectal tumor molecular features differ by age and tumor location is important in personalized patient management. METHODS: Using 14,004 cases with colorectal cancer including 3,089 early-onset cases, we examined microsatellite instability (MSI), CpG island methylator phenotype (CIMP), and KRAS and BRAF mutations in carcinomas of the cecum, ascending colon, transverse colon, descending colon, sigmoid colon, and rectum and compared early-onset cases with later-onset cases. RESULTS: The proportions of MSI-high, CIMP-high, and BRAF -mutated early-onset tumors were lowest in the rectum (8.8%, 3.4%, and 3.5%, respectively) and highest in the ascending colon (46% MSI-high; 15% CIMP-high) or transverse colon (8.6% BRAF -mutated) (all P trend <0.001 across the rectum to ascending colon). Compared with later-onset tumors, early-onset tumors showed a higher prevalence of MSI-high status and a lower prevalence of CIMP-high status and BRAF mutations in most subsites. KRAS mutation prevalence was higher in the cecum compared with that in the other subsites in both early-onset and later-onset tumors ( P < 0.001). Notably, later-onset MSI-high tumors showed a continuous decrease in KRAS mutation prevalence from the rectum (36%) to ascending colon (9%; P trend <0.001), followed by an increase in the cecum (14%), while early-onset MSI-high cancers showed no such trend. DISCUSSION: Our findings support biogeographical and pathogenic heterogeneity of colorectal carcinomas in different colorectal subsites and age groups.

Abstract 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 &lt;0.001) followed by an increase in the cecum (14%), while early-onset MSI-high cancer showed no such trend. In survival analyses, there was a trend for better CRC-specific survival from the cecum to sigmoid colon (Ptrend&lt;0.001) (but not to rectum), which did not significantly differ by age of diagnosis (Pinteraction=0.61). Multivariable-adjusted hazard ratios (HRs) for sigmoid vs. cecum were 0.86 (95% CI, 0.64-1.14) for early-onset cases and 0.81 (95% CI, 0.70-0.93) for later-onset cases. Notably, the prognostic association of tumor location differed by MSI status (Pinteraction=0.001). Non-MSI-high tumors exhibited the cecum-to-sigmoid trend for better CRC-specific survival [Ptrend &lt;0.001; multivariable-adjusted HR for the sigmoid colon (vs. cecum), 0.80; 95% CI, 0.70-0.92], whereas MSI-high tumors demonstrated an opposite cecum-to-sigmoid trend for worse survival (Ptrend = 0.020; the corresponding HR, 2.13; 95% CI, 1.15-3.92). Conclusions: Our findings support biogeographical and pathogenic heterogeneity of CRC in different age groups and colorectal subsites. Both detailed colorectal location and tumor molecular features need to be accounted for prognostication of early-onset and later-onset CRCs to advance precision medicine. 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.

Abstract Background Obesity is an established risk factor for colorectal cancer (CRC), but the evidence for the association is inconsistent across molecular subtypes of the disease. Methods We pooled data on body mass index (BMI), tumor microsatellite instability status, CpG island methylator phenotype status, BRAF and KRAS mutations, and Jass classification types for 11 872 CRC cases and 11 013 controls from 11 observational studies. We used multinomial logistic regression to estimate odds ratios (OR) and 95% confidence intervals (CI) adjusted for covariables. Results Higher BMI was associated with increased CRC risk (OR per 5 kg/m2 = 1.18, 95% CI = 1.15 to 1.22). The positive association was stronger for men than women but similar across tumor subtypes defined by individual molecular markers. In analyses by Jass type, higher BMI was associated with elevated CRC risk for types 1-4 cases but not for type 5 CRC cases (considered familial-like/Lynch syndrome microsatellite instability-H, CpG island methylator phenotype-low or negative, BRAF-wild type, KRAS-wild type, OR = 1.04, 95% CI = 0.90 to 1.20). This pattern of associations for BMI and Jass types was consistent by sex and design of contributing studies (cohort or case-control). Conclusions In contrast to previous reports with fewer study participants, we found limited evidence of heterogeneity for the association between BMI and CRC risk according to molecular subtype, suggesting that obesity influences nearly all major pathways involved in colorectal carcinogenesis. The null association observed for the Jass type 5 suggests that BMI is not a risk factor for the development of CRC for individuals with Lynch syndrome.

AbstractDiabetes is an established risk factor for colorectal cancer. However, colorectal cancer is a heterogeneous disease and it is not well understood whether diabetes is more strongly associated with some tumor molecular subtypes than others. A better understanding of the association between diabetes and colorectal cancer according to molecular subtypes could provide important insights into the biology of this association. We used data on lifestyle and clinical characteristics from the Colorectal Cancer Family Registry (CCFR) and the Genetics and Epidemiology of Colorectal Cancer Consortium (GECCO), including 9756 colorectal cancer cases (with tumor marker data) and 9985 controls, to evaluate associations between reported diabetes and risk of colorectal cancer according to molecular subtypes. Tumor markers included BRAF and KRAS mutations, microsatellite instability and CpG island methylator phenotype. In the multinomial logistic regression model, comparing colorectal cancer cases to cancer‐free controls, diabetes was positively associated with colorectal cancer regardless of subtype. The highest OR estimate was found for BRAF‐mutated colorectal cancer, n = 1086 (ORfully adj: 1.67, 95% confidence intervals [CI]: 1.36‐2.05), with an attenuated association observed between diabetes and colorectal cancer without BRAF‐mutations, n = 7959 (ORfully adj: 1.33, 95% CI: 1.19‐1.48). In the case only analysis, BRAF‐mutation was differentially associated with diabetes (Pdifference = .03). For the other markers, associations with diabetes were similar across tumor subtypes. In conclusion, our study confirms the established association between diabetes and colorectal cancer risk, and suggests that it particularly increases the risk of BRAF‐mutated tumors.