Thibodeau, Stephen N.

3523 Background: The association of KRAS or BRAFV600E mutations with prognosis in colon cancers has been inconsistent. Since primary tumor site may influence outcome, we analyzed KRAS and BRAFV600E stratified by tumor site in resected stage III colon cancers from a randomized adjuvant trial of mFOLFOX6 + cetuximab chemotherapy (N= 2,686) where no survival difference by treatment was found. Methods: 2,580 tumors were analyzed for mutations in BRAFV600E (exon 15) or KRAS (codons 12, 13), and for deficient DNA mismatch repair (dMMR). Cox models were used, adjusting for mutation status, age, sex, treatment, T-stage, histologic grade, nodal status, tumor site, and MMR. After study initiation, eligibility was restricted to patients (pts) with KRAS wild-type (WT) tumors. At a median follow-up 4.1 yrs, 83% of pts are alive. Results: KRAS and BRAFV600E mutations were detected in 716 (28%) and 346 (14%) tumors, respectively; dMMR was found in 314 (12%). Proximal (to splenic flexure) tumors (50%) were associated with older age and were more likely to be high grade (33 vs 18%), T-stage3,4 (88 vs 82%), mutated for KRAS (33 vs 23%) or BRAFV600E (23 vs 4%), and dMMR (21 vs 3%)[all p <0.02]. Pts with distal vs proximal tumors showed improved disease-free survival (DFS)[ HR 0.7, 95% CI (0.6-0.9); p<0.01 unadjusted for KRAS or BRAF;]. Mutant KRAS [HR=1.5 (1.2-1.7); p<.0001] or mutant BRAF [HR=1.4 (1.1-1.7); p=0.02] were each independently associated with worse DFS. The KRAS and tumor site interaction was significant (unadjusted pinteraction=0.02; adjusted pinteraction=0.07), with poorer DFS among distal tumors having mutant KRAS [HR 1.7 (1.3-2.2); p<.0001]. No interaction was observed for BRAFV600E and tumor site, despite worse DFS for mutant BRAFV600E among proximal tumors [HR 1.3 (1-1.8); p=0.04]. Conclusions: The adverse prognostic impact of KRAS, but not BRAFV600E, mutations was confined to the distal colon. This finding underscores the importance of tumor site in the interpretation of the prognostic impact of KRAS status, and motivates similar analyses in pts with advanced disease.

Abstract Background: Results from epidemiologic studies examining polyunsaturated fatty acids (PUFA) and colorectal cancer risk are inconsistent. Mendelian randomization may strengthen causal inference from observational studies. Given their shared metabolic pathway, examining the combined effects of aspirin/NSAID use with PUFAs could help elucidate an association between PUFAs and colorectal cancer risk. Methods: Information was leveraged from genome-wide association studies (GWAS) regarding PUFA-associated SNPs to create weighted genetic scores (wGS) representing genetically predicted circulating blood PUFAs for 11,016 non-Hispanic white colorectal cancer cases and 13,732 controls in the Genetics and Epidemiology of Colorectal Cancer Consortium (GECCO). Associations per SD increase in the wGS were estimated using unconditional logistic regression. Interactions between PUFA wGSs and aspirin/NSAID use on colorectal cancer risk were also examined. Results: Modest colorectal cancer risk reductions were observed per SD increase in circulating linoleic acid [ORLA = 0.96; 95% confidence interval (CI) = 0.93–0.98; P = 5.2 × 10−4] and α-linolenic acid (ORALA = 0.95; 95% CI = 0.92–0.97; P = 5.4 × 10−5), whereas modest increased risks were observed for arachidonic (ORAA = 1.06; 95% CI = 1.03–1.08; P = 3.3 × 10−5), eicosapentaenoic (OREPA = 1.04; 95% CI = 1.01–1.07; P = 2.5 × 10−3), and docosapentaenoic acids (ORDPA = 1.03; 95% CI = 1.01–1.06; P = 1.2 × 10−2). Each of these effects was stronger among aspirin/NSAID nonusers in the stratified analyses. Conclusions: Our study suggests that higher circulating shorter-chain PUFAs (i.e., LA and ALA) were associated with reduced colorectal cancer risk, whereas longer-chain PUFAs (i.e., AA, EPA, and DPA) were associated with an increased colorectal cancer risk. Impact: The interaction of PUFAs with aspirin/NSAID use indicates a shared colorectal cancer inflammatory pathway. Future research should continue to improve PUFA genetic instruments to elucidate the independent effects of PUFAs on colorectal cancer.

Abstract Germline mutations in exons 10, 11, and 16 of the RET protooncogene are associated with the heritable cancer syndromes multiple endocrine neoplasia (MEN) type 2A, familial medullary thyroid carcinoma (FMTC), and MEN type 2B. Nonradioactive mutation analysis with nondenaturing Phastgels® and the Phast System™ was performed on DNA amplified by the polymerase chain reaction from exons 10, 11, and 16 of the RET protooncogene from patients with MEN 2A, MEN 2B, or FMTC. The analysis requires ∼45–90 min for electrophoresis and 35 min for staining. This assay detected 20 of 21 different mutations that represented ∼90% of all known mutations associated with these lesions. A rare silent polymorphism within exon 10 was also detected. This form of mutation analysis provides simple, rapid, and highly sensitive nonradioactive detection of mutations known to be associated with MEN 2A, FMTC, and MEN 2B.

Purpose The Mayo Clinic Biobank was established to provide a large group of patients from which comparison groups (ie, controls) could be selected for case–control studies, to create a prospective cohort with sufficient power for common outcomes and to support electronic health record (EHR) studies. Participants A total of 56 862 participants enrolled (21% response rate) into the Mayo Clinic Biobank from Rochester, Minnesota (77%, n=43 836), Jacksonville, Florida (18%, n=10 368) and La Crosse, Wisconsin (5%, n=2658). Participants were all Mayo Clinic patients, 18 years of age or older and US residents. Findings to date Overall, 43% of participants were 65 years of age or older and female participants were more frequent (59%) than males at all sites. Most participants resided in the Upper Midwest regions of the USA (Minnesota, Iowa, Illinois or Wisconsin), Florida or Georgia. Self-reported race among Biobank participants was 90% white. Here we provide examples of the types of studies that have successfully utilised the resource, including (1) investigations of the population itself, (2) provision of controls for case–control studies, (3) genotype-driven research, (4) EHR-based research and (5) prospective recruitment to other studies. Over 270 projects have been approved to date to access Biobank data and/or samples; over 200 000 sample aliquots have been approved for distribution. Future plans The data and samples in the Mayo Clinic Biobank can be used for various types of epidemiological and clinical studies, especially in the setting of case–control studies for which the Biobank samples serve as control samples. We are planning cohort studies with additional follow-up and acquisition of genetic information on a large scale.

AbstractWe determined that the molecular defect of 2 patients with hemoglobin (Hb) M-Kankakee [Hb M-Iwate, 87 (F8) His → Tyr] resides in the 1-globin gene. The proportion of Hb M observed is higher than that predicted for an 1-globin variant. Our evidence suggests that the greater-than-expected proportion of Hb M-Kankakee results from preferential association of the electronegative β-globin chains with the M-globin chains that are more electropositive than normal -globin chains.

We determined that the molecular defect of 2 patients with hemoglobin (Hb) M-Kankakee [Hb M-Iwate, 87 (F8) His → Tyr] resides in the 1-globin gene. The proportion of Hb M observed is higher than that predicted for an 1-globin variant. Our evidence suggests that the greater-than-expected proportion of Hb M-Kankakee results from preferential association of the electronegative β-globin chains with the M-globin chains that are more electropositive than normal -globin chains.

Genome-wide association studies have identified over 150 risk loci that increase prostate cancer risk. However, few causal variants and their regulatory mechanisms have been characterized. In this study, we utilized our previously developed single-nucleotide polymorphisms sequencing (SNPs-seq) technology to test allele-dependent protein binding at 903 SNP sites covering 28 genomic regions. All selected SNPs have shown significant cis-association with at least one nearby gene. After preparing nuclear extract using LNCaP cell line, we first mixed the extract with dsDNA oligo pool for protein–DNA binding incubation. We then performed sequencing analysis on protein-bound oligos. SNPs-seq analysis showed protein-binding differences (>1.5-fold) between reference and variant alleles in 380 (42%) of 903 SNPs with androgen treatment and 403 (45%) of 903 SNPs without treatment. From these significant SNPs, we performed a database search and further narrowed down to 74 promising SNPs. To validate this initial finding, we performed electrophoretic mobility shift assay in two SNPs (rs12246440 and rs7077275) at CTBP2 locus and one SNP (rs113082846) at NCOA4 locus. This analysis showed that all three SNPs demonstrated allele-dependent protein-binding differences that were consistent with the SNPs-seq. Finally, clinical association analysis of the two candidate genes showed that CTBP2 was upregulated, while NCOA4 was downregulated in prostate cancer (p < 0.02). Lower expression of CTBP2 was associated with poor recurrence-free survival in prostate cancer. Utilizing our experimental data along with bioinformatic tools provides a strategy for identifying candidate functional elements at prostate cancer susceptibility loci to help guide subsequent laboratory studies.

Abstract Background: Different molecules within coffee may be pro-carcinogenic, while others may have anti-oxidant and anti-neoplastic effects. Hence, it can be hypothesized that coffee may have differing risk associations with specific colorectal cancer (CRC) subtypes. The aim of this study was to evaluate associations between coffee consumption and incident CRC overall, by anatomic subsite, and by molecularly defined tumor subtypes, in the population-based Iowa Women's Health Study (IWHS). Methods: IWHS participants, ages 55-69 years, provided dietary intake estimates using self-reported food frequency questionnaires at baseline (1986). From 35,216 women included the final analysis, 1298 developed CRC (673 with proximal CRC, 597 with distal CRC) through December 31, 2002 (end of the IWHS molecular epidemiology project study period). Archived, paraffin-embedded tissue specimens for 732 representative CRC cases were collected and analyzed to determine if coffee consumption (none or &lt;1 cup/month, &lt;1 cup/week, 1 cup/day, 2-3 cups/day and ≥4 cups/day) was associated with microsatellite instability (MSI-L/MSS or MSI-H), CpG island methylator phenotype (CIMP-negative or CIMP-positive), BRAF mutation status (BRAF-wildtype or BRAF-mutated), KRAS mutation status (KRAS-wildtype and KRAS-mutated), p53 protein expression level (p53 absent, low or high) and estrogen receptor β protein expression level (ERβ absent, low or high). Multivariable Cox regression models were fit to estimate relative risks (RR) and 95% confidence intervals (CI) with adjustment for age, smoking, alcohol use, diabetes, body mass index and waist-hip ratio, physical activity, estrogen use, dietary intake of fat, calcium, vitamin E, sucrose, red meat and methionine. P-values were not adjusted for multiple testing. Results: There was no significant difference in the risk of overall (RR, 1.05; 95% CI, 0.85-1.29; Ptrend=0.50), proximal (RR, 0.91; 95% CI, 0.68-1.21; Ptrend=0.57) or distal CRC (RR, 1.18; 95% CI, 0.86-1.64; Ptrend=0.19) in women who consumed ≥4 cups of coffee/day (caffeinated or decaffeinated), as compared to non-consumers. By CRC molecular subtypes for these same two exposure groups, women with ≥4 cups of coffee/day were more likely to develop ERβ-low CRC (RR, 2.98; 95% CI, 1.04-8.51; Ptrend=0.06), though no significant associations were observed for caffeinated or decaffeinated coffee specifically. In contrast, women who consumed ≥4 cups of decaffeinated coffee/day were less likely to develop MSI-L/MSS (RR, 0.57; 95% CI, 0.36-0.92; Ptrend=0.04) and BRAF-WT (RR, 0.69; 95% CI, 0.44-1.06; Ptrend=0.06) CRCs than non-consumers. Women who consumed ≥4 cups of caffeinated coffee/day were also less likely to develop p53-low CRCs (RR, 0.53; 95% CI, 0.24-1.15; Ptrend=0.02). Null associations were observed between decaffeinated coffee consumption and risk of CIMP-, KRAS- and p53- defined CRC subtypes, as well as between caffeinated coffee consumption and risk of MSI-, BRAF-, CIMP- or KRAS-defined CRC subtypes. Conclusion: In this prospective cohort study of older women, though coffee consumption was not associated with risk of overall or site-specific CRC, unique associations were observed with molecularly defined CRC subtypes, supporting a need for further investigation of the potential pro- and anti-carcinogenic effects of coffee constituents on biologic targets involved in colorectal carcinogenesis. Citation Format: Siddharth Singh, Robert Vierkant, Lori Tillmans, Alice Wang, Daniel Weisenberger, Peter Laird, Maki Inoue-Choi, Andrea Mariani, Charles Lynch, Kristin Anderson, Amy French, Robert Haile, Lisa Harnack, John Potter, Susan Slager, Thomas Smyrk, Stephen Thibodeau, James Cerhan, Paul Limburg. Coffee consumption and colorectal cancer risk by molecularly defined subtypes in a prospective cohort study of older women. [abstract]. In: Proceedings of the Twelfth Annual AACR International Conference on Frontiers in Cancer Prevention Research; 2013 Oct 27-30; National Harbor, MD. Philadelphia (PA): AACR; Can Prev Res 2013;6(11 Suppl): Abstract nr A44.

AbstractThe familial recurrence risk is the probability a person will have disease, given a reported family history. When family histories are obtained as simple counts of disease among family members, as often obtained in cancer registries or surveys, we propose methods to estimate recurrence risks based on truncated binomial distributions. By this approach, we are able to obtain unbiased estimates of risk for a person with at least k‐affected relatives, where k can be specified to determine how risk varies with k. We also derive robust variances of the recurrence risk estimate, to account for correlations within families, such as those induced by shared genes or shared environment, without explicitly modeling the factors that cause familial correlations. Furthermore, we illustrate how mixture models can be used to account for a sample composed of low‐ and high‐risk families. Using simulations, we illustrate the properties of the proposed methods. Application of our methods to a family history survey of prostate cancer shows that the recurrence risk for prostate cancer increased from 16%, when there was at least one affected relative, to 52%, when there was at least five affected relatives.