Epidemiology Of Colorectal Cancer Study Research Articles

Differences in Tumor-Associated T cell receptor repertoires between Early-Onset and Average-Onset colorectal cancer.

The incidence of colorectal cancer (CRC) among individuals younger than age 50 (early onset CRC; EOCRC) has substantially increased, yet the etiology and molecular mechanisms underlying this alarming rise remain unclear. We compared tumor-associated T cell repertoires between EOCRC and average-onset CRC (AOCRC) to uncover potentially unique immune microenvironment-related features by age of onset. Our discovery cohort included 242 patients who underwent surgical resection at Cleveland Clinic from 2000 to 2020. EOCRC was defined as age < 50 years at diagnosis (N = 126), and AOCRC as age ≥ 60 years (N = 116). T cell receptor (TCR) abundance and clonality were measured by immunosequencing of tumors. Logistic regression models were used to evaluate the associations between TCR repertoire features and age of onset, adjusting for sex, race, tumor location, and stage. Findings were replicated in 152 EOCRC and 1,984 AOCRC cases from the Molecular Epidemiology of Colorectal Cancer Study. EOCRC tumors had significantly higher TCR diversity compared to AOCRC tumors in the discovery cohort (Odds Ratio (OR):0.44, 95% Confidence Interval (CI):0.32-0.61, p < .0001). This association was also observed in the replication cohort (OR : 0.74, 95% CI : 0.62-0.89, p = .0013). No significant differences in TCR abundance were observed between EOCRC and AOCRC in either cohort. Higher TCR diversity, suggesting a more diverse intratumoral T cell response, is more frequently observed in EOCRC than AOCRC. Further studies are warranted to investigate the role of T cell diversity and the adaptive immune response more broadly in the etiology and outcomes of EOCRC.

Germline genetic regulation of the colorectal tumor immune microenvironment

ObjectiveTo evaluate the contribution of germline genetics to regulating the briskness and diversity of T cell responses in CRC, we conducted a genome-wide association study to examine the associations between germline genetic variation and quantitative measures of T cell landscapes in 2,876 colorectal tumors from participants in the Molecular Epidemiology of Colorectal Cancer Study (MECC).MethodsGermline DNA samples were genotyped and imputed using genome-wide arrays. Tumor DNA samples were extracted from paraffin blocks, and T cell receptor clonality and abundance were quantified by immunoSEQ (Adaptive Biotechnologies, Seattle, WA). Tumor infiltrating lymphocytes per high powered field (TILs/hpf) were scored by a gastrointestinal pathologist. Regression models were used to evaluate the associations between each variant and the three T-cell features, adjusting for sex, age, genotyping platform, and global ancestry. Three independent datasets were used for replication.ResultsWe identified a SNP (rs4918567) near RBM20 associated with clonality at a genome-wide significant threshold of 5 × 10− 8, with a consistent direction of association in both discovery and replication datasets. Expression quantitative trait (eQTL) analyses and in silico functional annotation for these loci provided insights into potential functional roles, including a statistically significant eQTL between the T allele at rs4918567 and higher expression of ADRA2A (P = 0.012) in healthy colon mucosa.ConclusionsOur study suggests that germline genetic variation is associated with the quantity and diversity of adaptive immune responses in CRC. Further studies are warranted to replicate these findings in additional samples and to investigate functional genomic mechanisms.

Abstract LB384: Artificial intelligence measures of tumor infiltrating lymphocytes predict colorectal cancer-specific and overall survival

Abstract Tumor infiltrating lymphocytes (TILs) per high-power field (HPF) as measured by expert pathologists is a useful, independent, validated prognostic marker in colorectal cancer (CRC). Artificial intelligence (AI) techniques of deep learning (DL) can predict TILs directly from routinely collected, digitized hematoxylin and eosin (H&amp;E) pathology slides, but AI-predicted TILs have not yet been thoroughly validated as clinically relevant biomarker. Here we test whether AI-predicted TILs per HPF can be validated as an independent prognostic biomarker of CRC-specific survival in patients. We constructed an AI model to predict TILS per HPF using a weakly-supervised, transformer-based deep regression model, trained using 5-fold cross validation in a large dataset (n=1,738) and validated in a second independent dataset (n=223) of CRC cases. The model predicted TILs per HPF as a continuous variable, which was carried forward for survival analyses both as a continuous variable and dichotomized at a previously established a priori threshold of &amp;lt;2 vs. ≥2 TILs per HPF. Using longitudinal data from the population-based Molecular Epidemiology of Colorectal Cancer Study (median follow-up = 95 months), survival analyses were performed using non-parametric and Cox-proportional hazards models, with and without adjustment for age, sex, ethnicity, stage, and molecularly measured microsatellite instability (MSI). Two or more AI-predicted TILs per HPF were significantly associated with 5-year CRC-specific survival (p=0.0000037), 5-year overall survival (p=0.00021), and overall survival (p=0.000067). In a Cox proportional hazards model adjusting for age, sex, ethnicity, stage, and MSI, ≥2 AI-predicted TILs per HPF was significantly associated with improved 5-year CRC-specific survival, with a hazard ratio (HR) = 0.62, (95% confidence interval; 0.43, 0.91), p=0.01. Our new AI-driven deep learning model, which we call HopeSTIL, provides a highly efficient algorithm for analyzing digital pathology images of H&amp;E sections of CRC to quantify TILs per HPF. HopeSTIL is a validated prognostic marker for 5-year CRC-specific survival that does not require a pathologist to manually count and score TILs per HPF. Citation Format: Stephen B. Gruber, Omar S. El Nahhas, Joseph D. Bonner, Joel K. Greenson, Daniel Schmolze, Lawrence Shaktah, Jonathan Salazar, Lorena Reynaga, Sidney Lindsey, Jenny Lu, Allen Mao, Victor Moreno, Stephani L. Schmit, Ya-Yu Tsai, Stanley R. Hamilton, Gad Rennert, Jakob N. Kather. Artificial intelligence measures of tumor infiltrating lymphocytes predict colorectal cancer-specific and overall survival [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB384.

Heterozygote advantage at HLA class I and II loci and reduced risk of colorectal cancer.

Reduced diversity at Human Leukocyte Antigen (HLA) loci may adversely affect the host's ability to recognize tumor neoantigens and subsequently increase disease burden. We hypothesized that increased heterozygosity at HLA loci is associated with a reduced risk of developing colorectal cancer (CRC). We imputed HLA class I and II four-digit alleles using genotype data from a population-based study of 5,406 cases and 4,635 controls from the Molecular Epidemiology of Colorectal Cancer Study (MECC). Heterozygosity at each HLA locus and the number of heterozygous genotypes at HLA class -I (A, B, and C) and HLA class -II loci (DQB1, DRB1, and DPB1) were quantified. Logistic regression analysis was used to estimate the risk of CRC associated with HLA heterozygosity. Individuals with homozygous genotypes for all loci served as the reference category, and the analyses were adjusted for sex, age, genotyping platform, and ancestry. Further, we investigated associations between HLA diversity and tumor-associated T cell repertoire features, as measured by tumor infiltrating lymphocytes (TILs; N=2,839) and immunosequencing (N=2,357). Individuals with all heterozygous genotypes at all three class I genes had a reduced odds of CRC (OR: 0.74; 95% CI: 0.56-0.97, p= 0.031). A similar association was observed for class II loci, with an OR of 0.75 (95% CI: 0.60-0.95, p= 0.016). For class-I and class-II combined, individuals with all heterozygous genotypes had significantly lower odds of developing CRC (OR: 0.66, 95% CI: 0.49-0.87, p= 0.004) than those with 0 or one heterozygous genotype. HLA class I and/or II diversity was associated with higher T cell receptor (TCR) abundance and lower TCR clonality, but results were not statistically significant. Our findings support a heterozygote advantage for the HLA class-I and -II loci, indicating an important role for HLA genetic variability in the etiology of CRC.

Differences in tumor-associated T cell receptor repertoires between young-onset and average-onset colorectal cancer.

207 Background: The incidence of colorectal cancer (CRC) among individuals younger than age 50 (young-onset CRC; YOCRC) has increased, yet the etiology and molecular mechanisms underlying this alarming rise remain unclear. Some studies suggest that YOCRCs have more aggressive biology. Here, we compare tumor-associated T cell repertoires between patients with YOCRC and average-onset CRC (AOCRC). A robust, focused T cell response is a positive prognostic indicator; therefore, we hypothesized that YOCRCs demonstrate lower T cell abundance and greater T cell diversity than AOCRCs. Methods: The discovery cohort included 242 patients with histologically confirmed Stage I-IV CRC who underwent surgical resection at Cleveland Clinic from 2000 to 2020 and consented to a biorepository. YOCRC was defined as &lt; 50 years at diagnosis (N = 126), and AOCRC as &gt; 60 years (N = 116). DNA was extracted from frozen tumors, and T Cell Receptor (TCR) abundance and clonality were measured using immunoSEQ (Adaptive Biotechnologies). Following quality control, logistic regression models were used to evaluate the associations between TCR repertoire features and age of onset, adjusting for sex, race, tumor location, and stage. Findings were replicated in 152 YOCRC and 1,984 AOCRC cases from the Molecular Epidemiology of Colorectal Cancer Study with adjustment for sex, Jewish ethnicity, tumor location, and stage. Results: YOCRC patients were more likely to have left-sided tumors and be diagnosed at advanced stages in both cohorts. In the discovery cohort, YOCRC tumors had significantly lower TCR clonality (higher T cell diversity) compared to AOCRC tumors in a multivariable model adjusting for sex, race, tumor location and stage (OR: 0.38, 95% Confidence Interval (CI): 0.25-0.56, p &lt; 0.0001). This association was also observed in the replication cohort (OR: 0.74, 95% CI: 0.62-0.89, p = 0.0013). In the replication cohort, further adjustment for microsatellite instability status did not substantially change the association (OR: 0.73, 95% CI: 0.61-0.88 p = 0.0012). When restricting to microsatellite stable tumors, clonality remained statistically significant in both the discovery and replication cohorts. No significant difference in TCR abundance was observed between YOCRC and AOCRC in either cohort. Conclusions: Higher T cell repertoire diversity, indicating a less focused intratumoral T cell response, is more frequently observed in YOCRC. Further studies are warranted to investigate the role of T cell diversity in the etiology and outcomes of YOCRC.

Abstract 5874: Heterozygote advantage at HLA class I and II loci and colorectal cancer risk

Abstract Diversity in Human Leukocyte Antigen (HLA) genes has been associated with risk of several diseases, including Non-Hodgkin’s lymphoma and ulcerative colitis. Reduced diversity at HLA loci may adversely affect the host’s ability to recognize foreign antigens and tumor neoantigens, and subsequently, increase disease burden. To better understand the role of inherited HLA diversity in colorectal cancer (CRC) risk, we utilized data from a population-based study of 10,347 participants (5,574 CRC cases and 4,773 healthy controls) from the Molecular Epidemiology of Colorectal Cancer Study (MECC). Germline DNA samples were genotyped using genome-wide arrays, and HLA Class I and II four-digit resolution alleles were imputed using SNP2HLA and a reference panel of 5,225 individuals from the Type 1 Diabetes Genetics Consortium. Heterozygosity and homozygosity at each HLA locus and the number of homozygous genotypes at class I loci (A, B, C) and class II loci (DQB1, DRB1, DPB1) were quantified. To examine the joint effect of Class I and Class II loci, we combined the total number of homozygotes for all loci and categorized into 3 groups: heterozygotes at all loci, 1 to 4 homozygotes, or 5 or more homozygotes. Logistic regression was used to estimate the risk of CRC associated with HLA locus homozygosity. Individuals with heterozygous genotypes for all loci served as the reference category, and analyses were adjusted for sex, age, genotyping platform, and global ancestry. Individuals with homozygous genotypes at all 3 Class I genes had an increased risk of CRC when compared to those with heterozygous genotypes at all Class I loci (OR: 1.34; 95% CI: 1.02-1.76, P = 0.033; Ptrend = 0.039). A similar association was observed for Class II loci, with an OR of 1.32 (95% CI: 1.05-1.65, P = 0.015; Ptrend = 0.157). For HLA Class I and II combined, individuals with five or more homozygous genotypes at HLA class I or II loci were at higher risk for developing CRC (OR: 1.84, 95% CI: 1.24-2.73, P = 0.0023; Ptrend = 0.015), when compared to those with all heterozygous genotypes. Our findings support a heterozygote advantage at HLA class I and II loci as a protective factor for CRC. This indicates an important role for HLA genetic variability in the etiology of CRC potentially operating through a mechanism of decreased diversity of tumor neoantigens that can be displayed to the adaptive immune system. Citation Format: Ya-Yu Tsai, Chenxu Qu, Joseph D. Bonner, Rebeca Sanz-Pamplona, Sidney Lindsey, Marilena Melas, Kevin J. McDonnell, Gregory E. Idos, Christopher P. Walker, Kevin K. Tsang, Diane M. Da Silva, Ferran Moratalla, Asaf Maoz, Hedy S. Rennert, W. Martin Kast, Joel K. Greenson, Victor Moreno, Gad Rennert, Stephen B. Gruber, Stephanie L. Schmit. Heterozygote advantage at HLA class I and II loci and colorectal cancer risk [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5874.

Abstract 2737: Clinical and epidemiologic predictors of clonal immune responses in colorectal cancer

Abstract The quantity and quality of immune responses in colorectal cancers (CRC) are widely variable and have important clinical, therapeutic, and prognostic implications. We studied clinical and epidemiologic factors that might influence T-cell quantity and clonality within colorectal adenocarcinomas to better understand the drivers of diverse immune responses. Incident cases of CRC from the Molecular Epidemiology of Colorectal Cancer Study (MECC) were interviewed, and 6,006 cases had complete epidemiologic data. Archived tumor blocks were retrieved from 3,865 (64.4%) cases, and all were reviewed by a single expert pathologist who quantified TILS/hpf. Sufficient tissue for macrodissection and measurement of TCR abundance and clonality using the immunoSEQ assay (Adaptive Biotechnologies) was available and completed for 2,750 cases. Logistic regression, negative binomial regression and linear regression models were used to evaluate potential associations between clinical and epidemiologic variables for: TILS/hpf, TCR abundance, and T-cell clonality. The stage distribution was representative of cancer incidence in the population, and the MSI-H phenotype was observed in 14.2% of cases. Clinical, pathologic, and epidemiologic variables including aspirin, alcohol, diet, hormone use, physical activity, smoking, and statins were assessed in relation to immune measures. Among other findings, &amp;gt;5 years of statins (p&amp;lt;0.001) and daily aspirin (p=0.037) were each strongly associated with T-cell clonality. Our study suggests that important parameters of the adaptive immune response may be mediated by modifiable factors. Mechanisms regulating immune responses in CRC may have implications for chemoprevention as well as immunotherapy. Citation Format: Stephen B. Gruber, Joseph D. Bonner, Sidney S. Lindsey, Ya-Yu Tsai, Rebeca Sanz-Pamplona, M. Henar Alonso, Marilena Melas, Hedy S. Rennert, Kevin J. McDonnell, Gregory E. Idos, Christopher P. Walker, W. Martin Kast, Diane Da Silva, Harlan S. Robins, Joel K. Greenson, Victor Moreno, Stephanie L. Schmit, Gad Rennert. Clinical and epidemiologic predictors of clonal immune responses in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2737.

Abstract 824: Germline genetic regulation of the adaptive immune response in colorectal cancer

Abstract Adaptive immune responses in the tumor microenvironment of colorectal cancer (CRC) play an important role in prognosis. However, the contributions of germline genetic variations to the strength and diversity of T cell responses in CRC are unclear. We conducted a genome-wide association study to examine the relationships between germline genetic variants and measures of the T cell repertoire in colorectal tumors. Germline DNA samples from 5,581 CRC cases recruited into the Molecular Epidemiology of Colorectal Cancer Study (MECC) were genotyped in batches using four different platforms. Genotype data were imputed to the Haplotype Reference Consortium panel separately by genotyping platform. Tumor DNA samples were extracted from paraffin blocks, and tumor infiltrating lymphocytes per high powered field (TILs/hpf) were quantified by a single gastrointestinal pathologist. TCR abundance and clonality within individual CRCs were measured using the immunoSEQ assay (Adaptive Biotechnologies). Appropriate quality control steps and data transformations were applied to fit downstream statistical modeling assumptions. After standard quality control on both imputed genotypes and transformed immune metrics, 2,876 (TILs/hpf) and 2,395 (TCR abundance and clonality) cases with approximately 9 million imputed genetic variants were included in the discovery phase. Logistic or linear regression models were used to evaluate the associations between allelic dosage of each variant and each immune-related outcome, adjusting for sex, age at diagnosis, genotyping platform, and principal components for global ancestry. Three independent datasets were available to replicate our findings using similar quality control measures and regression models: Colonomics (N=96; TILs/hpf, TCR abundance, clonality), the CRC Genetics Study (N=162; TCR abundance, clonality), and the Harvard Cohorts (N=505; TILs/hpf; in progress). The discovery phase identified 5 independent genetic variants associated with TILs/hpf, 15 associated with TCR abundance, and 19 associated with clonality at p&amp;lt;5X10E-06. Replication analyses as well as expression quantitative trait analyses and in silico functional annotation are underway for the loci of interest. Our study suggests that germline genetic variation is associated with the quantity and quality of adaptive immune responses in CRC. Citation Format: Stephanie L. Schmit, Ya-Yu Tsai, Joseph Bonner, Rebeca Sanz-Pamplona, Amit D. Joshi, Sidney S. Lindsey, Marilena Melas, Kevin J. McDonnell, Gregory E. Idos, Christopher P. Walker, W. Martin Kast, Diane Da Silva, Tomotaka Ugai, Hedy S. Rennert, Harlan S. Robins, Joel K. Greenson, Shuji Ogino, Victor Moreno, Gad Rennert, Stephen B. Gruber. Germline genetic regulation of the adaptive immune response in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 824.

Cancer risk and overall survival in APC I1307K carriers.

1592 Background: The germline variant APC I1307K is one of the most commonly identified pathogenic variants on germline genetic testing panels. The purpose of the Molecular Epidemiology of Colorectal Cancer study was to quantify the risk of colorectal cancer among carriers, characterize the clinical, pathologic, and molecular features of colorectal cancers arising in patients with APC I1307K, and to describe the overall and disease-specific survival of carriers with colorectal cancer. Here, the final results of the Molecular Epidemiology of Colorectal Cancer Study are reported with respect to APC I1307K. Methods: We consented 6,006 incident, pathologically confirmed cases of colorectal adenocarcinoma and 5,023 age, sex, and ethnicity matched controls without colorectal cancer between March 31, 1998 and July 1, 2017 within a geographically defined area of Northern Israel. Comprehensive, in-person epidemiologic interviews were conducted for cases and controls, with uniform histopathologic review, detailed molecular analysis, medical record review and clinical follow-up for up to 21 years. Results: The demographic and clinical features of incident colorectal cancer cases matched the population distribution of colorectal cancer in Israel. APC I1307K was identified in 429 (7.1%) of cases and 201 (4.0%) of controls. The estimated relative risk of colorectal cancer among carriers was 1.89 (95% confidence interval, 1.59 - 2.24), p &lt; 0.0001. The prevalence and odds ratios differed by ethnic group. Homozygous carriers were at especially high risk, with an odds ratio of 3.90 (95% confidence interval 1.11–13.71). APC I1307K carriers were significantly less likely to have microsatellite instable tumors (p = 0.04). Overall survival of APC I1307K carriers was not significantly different than survival of non-carriers, after adjustment for age, stage, sex, ethnicity, and microsatellite instability. Conclusions: APC I1307K is an actionable germline mutation that confers meaningful lifetime risk of colorectal cancer in heterozygous and homozygous carriers. APC I1307K is not an independent prognostic factor for overall survival or disease specific survival and is not associated with the MSI phenotype. Cumulative lifetime risk estimates inform genetic counseling and provide data for policies regarding the timing and frequency of screening and other preventive strategies.

Hormonal and reproductive factors and reduction in the risk of colorectal cancer.

Females differ from males in incidence and clinical characteristics of colorectal cancer. Understanding the differences can lead to development of preventive approaches. To identify reproductive factors currently associated with the risk of colorectal cancer. Consecutively diagnosed female colorectal cancer cases and randomly chosen colorectal cancer-free controls matched on age/ethnicity/primary care clinic within the molecular epidemiology of colorectal cancer study, a population-based case-control study in Northern Israel, were included. A total of 2867 female cases and 2333 controls participated in this analysis. Participants were interviewed on reproductive history: ages at menarche, menopause, first birth, terminations of pregnancies, miscarriages, births, use of oral contraceptives. Among 5200 women, spontaneous miscarriages (odds ratio = 0.71, 0.61-0.83 for ever/never in Jews; odds ratio = 0.76, 0.53-1.08 in Arabs) and number of miscarriages, but not termination of pregnancies, as well as use, and duration of use, of oral contraceptives (Jews: odds ratio = 0.49, 0.39-0.62 for ever/never; Arabs: odds ratio = 0.14, 0.04-0.47) were strongly inversely associated with colorectal cancer risk. Up to 5 pregnancies were associated with increased risk while ages at menarche, at menopause and at first birth were not associated with colorectal cancer risk. Miscarriages but not terminations of pregnancy or full-term pregnancies, and use of oral contraceptives, were strongly associated with reduced odds of developing colorectal cancer suggesting unique hormonal influences on colorectal cancer.

Abstract 2332: Tumor infiltrating lymphocytes, immunoSeq, and CMS classification in the molecular epidemiology of colorectal cancer study

Abstract Background: Tumor infiltrating lymphocytes (TILs) are prognostic and predictive biomarkers in colorectal cancer and are associated with improved prognosis and response to immunotherapy. While TILs are routinely assessed by pathologists, a standardized technique (immunoSEQ, Adaptive Biotechnologies) that leverages targeted next-generation sequencing can also be used to quantify and characterize the T-cell receptor (TCR) repertoire of individual colorectal cancers. In a large, population-based study of incident colorectal cancer, the host immune responses were measured by an expert pathologist and ImmunoSEQ to understand the relationships between TILs, TCRs/cell and specific subgroups of colorectal cancer. Methods: Incident cases of adenocarcinoma of the colon or rectum from the Molecular Epidemiology of Colorectal Cancer (MECC) study included 1,000 cancers that were uniformly evaluated for TILs and other histopathologic features by one pathologist. FFPE-derived DNA from microdissected tumor tissue was extracted and sequenced using ImmunoSEQ analysis for the same 1,000 individuals. A resulting quantitative metric from this assay includes TCRs/cell, a measure of rearranged T cell quantity relative to all nucleated cells in a tumor sample. Gene expression in snap-frozen tissue available from 342/1,000 MECC colorectal cancers was measured with Affymetrix Human Genome U133 Arrays (U133A and U133 Plus2.0) as previously described. CMS classification was performed using the R package 3.5.1, CMS classifier, randomForest 4.6-14. Multivariate analysis assessed CMS by age, gender, TILs/HPF, TCRs/cell, MSI status, BRAF and KRAS mutational status. Results: TILs/HPF and TCRs/cell were significantly correlated among all 1000 cases (r=0.5, p&amp;lt;0.001). Among the 342 cases with available expression profiles, CMS1 constituted 12.0% of all CRC, with CMS2 (41.8%), CMS3 (8.5%), and CMS4 (13.7%) and unclassified (24%) representing the remaining distribution. There were statistically significant differences in the molecular and histopathologic features of colorectal cancers by CMS subgroups. MSI-H tumors were most frequently observed within CMS1 cancers (56.6% of CMS1 were MSI-H), with lower representation among CMS2 (1.5%), CMS3 (10%), CMS4 (3.5%), and unclassified CRC (9.5%) (p&amp;lt;0.0001). In addition, BRAF positive tumors were more frequently observed within the CMS1 group (12.2%, p =0.0065) and KRAS positive tumors within the CMS3 group (31%, p&amp;lt;0.0001). Consistent with prior reports, TILs/HPF were significantly higher in the CMS1 group (mean=7.7, p&amp;lt;0.0001). Similar statistically significant trends were observed across classes for TCRs/cell (mean=0.16, p=0.04). Conclusions: Subtypes of CRC have distinct histopathologic and molecular features that can be distinguished by expression profiles and immunoSEQ. Citation Format: Marilena Melas, Charalampos Lazaris, Stephanie L. Schmit, Asaf Maoz, Rebeca Sanz Pamplona, Chenxu Qu, Joel K. Greenson, Rork Kuick, Flavio Lejbkowicz, Hedy S. Rennert, Christopher P. Walker, Chase M. Bowen, Diane M. Da Silva, W. Martin Kast, Gregory E. Idos, Kevin J. McDonnell, Victor Moreno, Gad Rennert, Stephen B. Gruber. Tumor infiltrating lymphocytes, immunoSeq, and CMS classification in the molecular epidemiology of colorectal cancer study [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 2332.

Red meat and processed meat intake and risk of colorectal cancer: a population-based case-control study.

To examine the association between red meat subtypes intake and risk of colorectal cancer (CRC) among Jewish and Arabs populations in a unique Mediterranean environment. The Molecular Epidemiology of Colorectal Cancer study (n=10 026) is a prospective population-based case-control study in northern Israel. Participants were interviewed in-person about their dietary intake and lifestyle using a questionnaire that included a food-frequency questionnaire. Red meat consumption in Israel was found to be especially low in the Jewish population (1.29±1.45 servings/week), but higher in Arabs (3.0±1.98 servings/week) (P<0.001). Beef was the most commonly consumed red meat by Jews (1.15/1.29 servings/week, 89%) and proportionally less so by Arabs (2.00/3.00, 67%). Processed meat consumption (mostly pork free) was lower among Arabs (0.9±1.56 servings/week) compared with Jews (1.97±2.97 servings/week) (P<0.001). The adjusted odds of CRC per one serving/week of red meat were 1.05 (95% confidence interval: 1.01-1.08) in Jews and 0.94 (0.88-1.01) in Arabs. Compared with no consumption, beef consumption was associated with odds ratio (OR)=0.96 (0.86-1.07) in Jews and 0.94 (0.61-1.45) in Arabs, lamb consumption with OR=1.28 (1.10-1.5) and 1.01 (0.75-1.37), pork consumption with OR=1.44 (1.24-1.67) and 1.07 (0.73-1.56), and processed meat consumption with OR=1.22 (1.10-1.35) and 1.04 (0.82-1.33) in Jews and Arabs, respectively. Overall red meat consumption was associated weakly with CRC risk, significant only for lamb and pork, but not for beef, irrespective of tumor location. Processed meat was associated with mild CRC risk.

Highlights of This Issue

Bonn and colleagues investigated the effect of physical activity after a prostate cancer diagnosis in a large cohort. Statistically significantly lower overall mortality rates were found among men engaged in walking/bicycling ≥20 min/day, performing household work ≥1 h/day, or exercising ≥1 h/week, compared to less active men. Higher levels of physical activity were associated with reduced rates of overall and prostate cancer–specific mortality, further strengthening previous results indicating beneficial effects of physical activity on survival among men with prostate cancer.Fecal immunochemical testing (FIT) is one of the primary screening tests for colorectal cancer. Clarke and colleagues performed a meta-analysis to determine if uptake of FIT screening differs between men and women. Nineteen studies met the authors' inclusion criteria. Male FIT uptake was significantly lower than female uptake and this male/female difference persisted regardless of study design, screening organization, number of samples, age range, and setting. Further investigation is required to identify the factors that influence male nonparticipation in FIT screening.Milne and colleagues investigated associations between pediatric acute lymphoblastic leukemia (ALL) risk and folate pathway gene polymorphisms and their modification by maternal folic acid supplements. Seven folate pathway gene polymorphisms were genotyped in 392 pediatric ALL cases and pre-pregnancy maternal folic acid supplement use information was collected. The authors report some folate-pathway gene polymorphisms in the child or parent may influence ALL risk. Folate pathway polymorphisms may be related to risk of childhood ALL, but larger studies are needed for conclusive results.Schmit and colleagues searched for miRNA-related polymorphisms contributing to colorectal cancer etiology using the Axiom miRNA Target Site Genotyping Array containing 237,858 markers. The study included 596 cases from the Molecular Epidemiology of Colorectal Cancer study. The authors identified twenty-three markers with P values less than 5.0E–04 and replicated a previously published locus within the 3′-untranslated region of the insulin receptor gene INSR. This study demonstrates the potential for a miRNA-targeted genome-wide association study to identify candidate susceptibility loci and prioritize them for functional characterization.

MicroRNA polymorphisms and risk of colorectal cancer.

miRNAs act as post-transcriptional regulators of gene expression. Genetic variation in miRNA-encoding sequences or their corresponding binding sites may affect the fidelity of the miRNA-mRNA interaction and subsequently alter the risk of cancer development. This study expanded the search for miRNA-related polymorphisms contributing to the etiology of colorectal cancer across the genome using a novel platform, the Axiom miRNA Target Site Genotyping Array (237,858 markers). After quality control, the study included 596 cases and 429 controls from the Molecular Epidemiology of Colorectal Cancer study, a population-based case-control study of colorectal cancer in northern Israel. The association between each marker and colorectal cancer status was examined assuming a log-additive genetic model using logistic regression adjusted for sex, age, and two principal components. Twenty-three markers had P values less than 5.0E-04, and the most statistically significant association involved rs2985 (chr6:34845648; intronic of UHRF1BP1; OR = 0.66; P = 3.7E-05). Furthermore, this study replicated a previously published risk locus, rs1051690, in the 3'-untranslated region of the insulin receptor gene INSR (OR = 1.38; P = 0.03), with strong evidence of differences in INSR gene expression by genotype. This study is the first to examine associations between genetic variation in miRNA target sites and colorectal cancer using a genome-wide approach. Functional studies to identify allele-specific effects on miRNA binding are needed to confirm the regulatory capacity of genetic variation to influence risk of colorectal cancer. This study demonstrates the potential for an miRNA-targeted genome-wide association study to identify candidate susceptibility loci and prioritize them for functional characterization.

Polymorphisms in alcohol metabolism genes ADH1B and ALDH2, alcohol consumption and colorectal cancer.

BackgroundColorectal cancer (CRC) is a leading cause of cancer death worldwide. Epidemiological risk factors for CRC included alcohol intake, which is mainly metabolized to acetaldehyde by alcohol dehydrogenase and further oxidized to acetate by aldehyde dehydrogenase; consequently, the role of genes in the alcohol metabolism pathways is of particular interest. The aim of this study is to analyze the association between SNPs in ADH1B and ALDH2 genes and CRC risk, and also the main effect of alcohol consumption on CRC risk in the study population.Methodology/Principal FindingsSNPs from ADH1B and ALDH2 genes, included in alcohol metabolism pathway, were genotyped in 1694 CRC cases and 1851 matched controls from the Molecular Epidemiology of Colorectal Cancer study. Information on clinicopathological characteristics, lifestyle and dietary habits were also obtained. Logistic regression and association analysis were conducted. A positive association between alcohol consumption and CRC risk was observed in male participants from the Molecular Epidemiology of Colorectal Cancer study (MECC) study (OR = 1.47; 95%CI = 1.18-1.81). Moreover, the SNPs rs1229984 in ADH1B gene was found to be associated with CRC risk: under the recessive model, the OR was 1.75 for A/A genotype (95%CI = 1.21-2.52; p-value = 0.0025). A path analysis based on structural equation modeling showed a direct effect of ADH1B gene polymorphisms on colorectal carcinogenesis and also an indirect effect mediated through alcohol consumption.Conclusions/SignificanceGenetic polymorphisms in the alcohol metabolism pathways have a potential role in colorectal carcinogenesis, probably due to the differences in the ethanol metabolism and acetaldehyde oxidation of these enzyme variants.

P-0206 - Kras Mutations in Metastatic Colorectal Cancer and the Effect of Bevacizumab

Background: Laboratory research of metastatic colorectal cancer subdivided the disease into a number of molecular sub-types mainly based on the status of KRAS gene. Recently different effect of certain monoclonal antibody (cetuximab) was shown with G13D KRAS mutation against previous data of drug resistance in KRAS mutant tumors. The question of different biological behavior of a tumor and treatment effect of bevacizumab was evaluated according to a KRAS sub-types status. Methods: Included were 397 colorectal cancer patients from the Israeli population-based Molecular Epidemiology of Colorectal Cancer study treated for their first metastatic event with irinotecan-based or 5-fluorouracil protocols, with or without bevacizumab, and no surgery. Patients were diagnosed between 1998 and 2010 and were followed up until October 2011; Mutations in KRAS codons 12, and 13 were identified by direct sequencing and SNP assay-by-design. Date of metastases and death and treatment details were extracted from the oncological follow-up records supported by computerized pharmacy records. Results: A total of 267 cases were treated with irinotecan-based chemotherapy, 96 (36%) of whom with combination with bevacizumab. A total of 130 cases were treated with 5-FU treatment, majority (98%) without bevacizumab. Mutation in codon 12 or codon 13 in KRAS was identified in 162 cases (41%), with 16.4%, 10.1% and 7.1% harboring mutations in the codon G12d, codon G12V and codon G13D, respectively. Combination 1st line treatment with IRINOTECAN-based chemotherapy and bevacizumab showed improved OS in cases with KRAS wild type tumors (HR = 0.69; p = 0.048), and KRAS codon 12 or codon 13 mutated tumors (HR = 0.68, p = 0.07). This effect was not different across types of KRAS mutation: G13D (HR = 0.46, p = 0.25), G12V (HR = 0.40; p = 0.049), G12D (HR = 0.76; p = 0.46), KRAS mutations (p for interaction =0.54) Assessment of prognostic effect in 5-FU only treated patients shows that relative to cases without any mutation in the codon 12 or codon 13, G13D (HR = 2.43) and G12D (HR = 1.75) mutations experienced a worse OS, while G12V (HR = 1.28) and all other codon 12 mutations combined (HR = 1.17) did not (p for difference = 0.075). Conclusion: The treatment effect of bevacizumab was not found to have a significant difference between KRAS sub-types of metastatic colorectal cancer. The data supports a prognostic effect of G13D and G12D KRAS mutations.

Genetic polymorphisms in fatty acid metabolism genes and colorectal cancer

Colorectal cancer (CRC) is a leading cause of cancer death worldwide. Epidemiological risk factors for CRC included dietary fat intake; consequently, the role of genes in the fatty acid biosynthesis and metabolism pathways is of particular interest. Moreover, hyperlipidaemia has been associated with different type of cancer and serum lipid levels could be affected by genetic factors, including polymorphisms in the lipid metabolism pathway. The aim of this study is to assess the association between single-nucleotide polymorphisms (SNPs) in fatty acid metabolism genes, serum lipid levels, body mass index (BMI) and dietary fat intake and CRC risk; 30 SNPs from 8 candidate genes included in fatty acid biosynthesis and metabolism pathways were genotyped in 1780 CRC cases and 1864 matched controls from the Molecular Epidemiology of Colorectal Cancer study. Information on clinicopathological characteristics, lifestyle and dietary habits were also obtained. Logistic regression and association analysis were conducted. Several LIPC (lipase, hepatic) polymorphisms were found to be associated with CRC risk, although no particular haplotype was related to CRC. The SNP rs12299484 showed an association with CRC risk after Bonferroni correction. We replicate the association between the T allele of the LIPC SNP rs1800588 and higher serum high-density lipoprotein levels. Weak associations between selected polymorphism in the LIPC and PPARG genes and BMI were observed. A path analysis based on structural equation modelling showed a direct effect of LIPC gene polymorphisms on colorectal carcinogenesis as well as an indirect effect mediated through serum lipid levels. Genetic polymorphisms in the hepatic lipase gene have a potential role in colorectal carcinogenesis, perhaps though the regulation of serum lipid levels.

Use of Bisphosphonates and Reduced Risk of Colorectal Cancer

Bisphosphonates are commonly used for the treatment of osteoporosis and bone metastases caused by breast cancer and were recently reported to be associated with a reduced risk of breast cancer, possibly acting through the mevalonate pathway, but their association with risk of other cancers is unknown. The Molecular Epidemiology of Colorectal Cancer study is a population-based, case-control study in northern Israel of patients with colorectal cancer and age-, sex-, clinic-, and ethnic group-matched controls. Long-term use of bisphosphonates before diagnosis was assessed in a subset of 933 pairs of postmenopausal female patients and controls, enrolled in Clalit Health Services, using computerized pharmacy records. The use of bisphosphonates for more than 1 year before diagnosis, but not for less than 1 year, was associated with a significantly reduced relative risk (RR) of colorectal cancer (RR, 0.50; 95% CI, 0.35 to 0.71). This association remained statistically significant after adjustment in a model for vegetable consumption, sports activity, family history of colorectal cancer, body mass index, and use of low-dose aspirin, statins, vitamin D, and postmenopausal hormones (RR, 0.41; 95% CI, 0.25 to 0.67). Concomitant use of bisphosphonates and statins did not further reduce the risk. The use of oral bisphosphonates for more than 1 year was associated with a 59% relative reduction in the risk of colorectal cancer, similar to the recently reported association of this drug class with reduction in breast cancer risk.

Clinical implications of UGT1A1*28 genotype testing in colorectal cancer patients

Metastatic colorectal cancer is frequently treated with irinotecan, a topoisomerase-I inhibitor. The UGT1A1 gene encodes for an enzyme that metabolizes irinotecan, and its genetic variants were shown to be associated with increased drug toxicity. We evaluated clinical outcomes associated with the UGT1A1*28 variant. The study included 329 colorectal cancer patients from the Israeli population-based Molecular Epidemiology of Colorectal Cancer study who were treated with a chemotherapy regimen that included irinotecan. Patients with metastases or disease recurrence were followed up for a median period of 2 years after occurrence of the event. Study end points were appearance of grade 3-4 hematological and gastroenterological toxicity, hospitalization due to toxic events (mostly neutropenia, fever, diarrhea, or vomiting), length of hospitalization, and overall survival. UGT1A1*28 was genotyped from peripheral blood DNA by fragment analysis and reported as number of TATA sequence repeats in the promoter of the gene. The 7/7 variant of UGT1A1*28 was detected in 11.9% of the 329 participants. Grade 3-4 hematological toxicity was significantly higher in 7/7 carriers compared with 6/7 and 6/6 carriers (48.0%,10.2%, and 7.7% respectively; P < .001), as was the risk of toxicity-related hospitalization (45.8%, 25.3%, and 14.4% respectively; P = .001). Both short-term death within 2 months of treatment start (12.8%, 5.2%, and 2.9%, respectively) and median overall survival (1.6, 2.0, and 2.4 years, respectively; P = .01) were significantly worse in the 7/7 carriers. The age/stage-adjusted hazard ratio for patients with the 7/7 genotype compared with 6/6 was 1.7 (95% confidence interval, 1.1-2.3). The UGT1A1*28 7/7 genotype is strongly associated with severe hematological toxicity and higher hospitalization rate and predicts lower survival of colorectal cancer in users of irinotecan.

Risk of colorectal cancer in self‐reported inflammatory bowel disease and modification of risk by statin and NSAID use

Statins and nonsteroidal anti-inflammatory drugs (NSAIDs) are associated with reduced risk of colorectal cancer (CRC) in some studies. The objective of this study was to quantify the relative risk of inflammatory bowel disease (IBD) as a risk factor for CRC and to estimate whether this risk may be modified by long-term use of NSAIDs or statins. The Molecular Epidemiology of Colorectal Cancer study is a population-based, case-control study of incident colorectal cancer in northern Israel and controls matched by age, sex, clinic, and ethnicity. Personal histories of IBD and medication use were measured by structured, in-person interview. The relative risk of IBD and effect modification by statins and NSAIDs were quantified by conditional and unconditional logistic regression. Among 1921 matched pairs of CRC cases and controls, a self-reported history of IBD was associated with a 1.9-fold increased risk of CRC (95% confidence interval [CI], 1.12-3.26). Long-term statin use was associated with a reduced risk of both IBD-associated CRC (odds ratio [OR] = 0.07; 95% CI, 0.01-0.78) and non-IBD CRC (OR = 0.49; 95% CI, 0.39-0.62). Stratified analysis suggested that statins may be more protective among those with IBD (ratio of OR = 0.14; 95% CI, 0.01-1.31; P = .51), although not statistically significant. NSAID use in patients with a history of IBD was suggestive of reduced risk of CRC but did not reach statistical significance (OR = 0.47; 95% CI, 0.12-1.86). The risk of CRC was elevated 1.9-fold in patients with IBD. Long-term statin use was associated with reduced risk of CRC in patients with IBD.