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Purpose: Apoptosis is essential for chemotherapy responses. In this discovery and validation study, we evaluated the suitability of a mathematical model of apoptosis execution (APOPTO-CELL) as a stand-alone signature and as a constituent of further refined prognostic stratification tools.Experimental Design: Apoptosis competency of primary tumor samples from patients with stage III colorectal cancer (n = 120) was calculated by APOPTO-CELL from measured protein concentrations of Procaspase-3, Procaspase-9, SMAC, and XIAP. An enriched APOPTO-CELL signature (APOPTO-CELL-PC3) was synthesized to capture apoptosome-independent effects of Caspase-3. Furthermore, a machine learning Random Forest approach was applied to APOPTO-CELL-PC3 and available molecular and clinicopathologic data to identify a further enhanced signature. Association of the signature with prognosis was evaluated in an independent colon adenocarcinoma cohort (TCGA COAD, n = 136).Results: We identified 3 prognostic biomarkers (P = 0.04, P = 0.006, and P = 0.0004 for APOPTO-CELL, APOPTO-CELL-PC3, and Random Forest signatures, respectively) with increasing stratification accuracy for patients with stage III colorectal cancer.The APOPTO-CELL-PC3 signature ranked highest among all features. The prognostic value of the signatures was independently validated in stage III TCGA COAD patients (P = 0.01, P = 0.04, and P = 0.02 for APOPTO-CELL, APOPTO-CELL-PC3, and Random Forest signatures, respectively). The signatures provided further stratification for patients with CMS1-3 molecular subtype.Conclusions: The integration of a systems-biology–based biomarker for apoptosis competency with machine learning approaches is an appealing and innovative strategy toward refined patient stratification. The prognostic value of apoptosis competency is independent of other available clinicopathologic and molecular factors, with tangible potential of being introduced in the clinical management of patients with stage III colorectal cancer. Clin Cancer Res; 23(5); 1200–12. ©2016 AACR.

Kaplan-Meier estimates for disease-free and overall survival for n=120 stage III patients of the discovery cohort categorized based on TN staging, tumor location and lymphovascular invasion. Additional exploratory analyses investigated the prognostic value of the APOPTO-CELL-PC3 signature within each sub-group identified by the clinical features.

Unadjusted and multivariate Cox proportional hazards analyses to examine the association of APOPTO-CELL and APOPTO-CELL-PC3 signatures with DFS in n=157 stage III patients of the expansion cohort.

Patient clinicopathological and demographic characteristics in the patients of the discovery, expansion and validation cohorts.

Detailed description of the measurements used as inputs for the apoptosis-based signatures in the discovery, expansion and validation cohorts.

Detailed description of the patients data handling and inclusion criteria for downstream analyses of the discovery, expansion and validation cohorts.

Impact of each predictor included in the Random Forest classifier on the probability of recurrence alone and in combination with the others.

Kaplan-Meier estimates for disease-free and overall survival for n=120 stage III patients of the discovery cohort. Patients were grouped based on the protein expression of Procaspase-9, XIAP, SMAC and Procaspase-3. Median expression was used as cut-off value.

Detailed description of the survival analyses performed in both primary and exploratory studies.

Background: Inherited predisposition may be associated with distinctive breast cancer phenotypes and/or mortality. Past studies have had inconsistent results and little is known about the contributions of screening and treatment. Methods: Within a population-based cohort of 1,260 women diagnosed with invasive breast cancer before age 46, we assessed how family history of breast cancer relates to mortality and tumor characteristics. Analyses were repeated excluding BRCA1/BRCA2 carriers. Medical records were reviewed for treatment history and tumors were centrally reviewed and tested. Cox proportional hazard modeling was used to assess the risk of dying in relation to family history; logistic regression was used to assess the association of family history to tumor characteristics. Results: Compared with women with no family history, women with first-degree family history of breast cancer had a 40% reduction (95% CI: 0.5–0.8) in the risk of dying. Mortality in women with only a second-degree family history was similar to those with no family history. The risk of dying was further reduced in those with a greater number of affected relatives. These relationships did not seem to be attributable to differences in screening, detection method, or treatment. Tumors in women with a first-degree family history had generally more favorable prognostic profiles. Conclusion: Our findings suggest that breast cancer patients with a first-degree family history, compared with their counterparts without such a profile, may have a better prognosis. Impact: These findings support the need for future research directed at replicating these results and identifying factors underlying this possible relationship. Cancer Epidemiol Biomarkers Prev; 20(12); 2560–71. ©2011 AACR.

Organ transplant recipients (OTRs) have a substantially elevated risk of squamous cell skin carcinoma (SCSC), largely attributed to immunosuppressive medications used to prevent graft rejection, although data to support the role of newer drugs in SCSC risk are sparse. We investigated the association between immunosuppressive medications and SCSC risk among cardiac and renal transplant recipients in the SCOT cohort study. Incident cases were ascertained through medical record review after self-report of skin biopsy (n = 170). Controls without SCSC (n = 324) were matched to cases on sex, age, race, transplant year, hospital, donor type, organ transplanted, and time between transplantation and interview. Conditional logistic regression was used to evaluate the association between specific medications and SCSC. Users of the antimetabolite azathioprine were more than twice as likely to develop SCSC (odds ratio [OR] = 2.67, 95% confidence interval [CI] 1.23-5.76). In contrast, the newer antimetabolite preparations (i.e., mycophenolic acid [MPA]) were associated with lower SCSC risk (OR = 0.45, 95% CI 0.29-0.69). This inverse association between MPA and SCSC persisted among OTRs with no history of azathioprine use, even after adjustment for simultaneous use of the calcineurin inhibitor tacrolimus (OR = 0.52, 95% CI 0.32-0.84). Our data suggest that the increased risk of SCSC historically associated with azathioprine is not seen in OTRs prescribed newer regimens, including MPA and tacrolimus.

Although use of non-steroidal anti-inflammatory drugs (NSAIDs) generally decreases colorectal cancer (CRC) risk, inherited genetic variation in inflammatory pathways may alter their potential as preventive agents. We investigated whether variation in prostaglandin synthesis and related pathways influences CRC risk in the Colon Cancer Family Registry by examining associations between 192 single nucleotide polymorphisms (SNPs) and two variable nucleotide tandem repeats (VNTRs) within 17 candidate genes and CRC risk. We further assessed interactions between these polymorphisms and NSAID use on CRC risk. Using a case-unaffected-sibling-control design, this study included 1621 primary invasive CRC cases and 2592 sibling controls among Caucasian men and women aged 18-90. After adjustment for multiple comparisons, two intronic SNPs were associated with rectal cancer risk: rs11571364 in ALOX12 [OR(het/hzv) = 1.87, 95% confidence interval (CI) = 1.19-2.95, P = 0.03] and rs45525634 in PTGER2 (OR(het/hzv) = 0.49, 95% CI = 0.29-0.82, P = 0.03). Additionally, there was an interaction between NSAID use and the intronic SNP rs2920421 in ALOX12 on risk of CRC (P = 0.03); among those with heterozygous genotypes, risk was reduced for current NSAID users compared with never or former users (OR(het) = 0.60, 95% CI = 0.45-0.80), though not among those with homozygous wild-type or variant genotypes. The results of this study suggest that genetic variation in ALOX12 and PTGER2 may affect the risk of rectal cancer. In addition, this study suggests plausible interactions between NSAID use and variants in ALOX12 on CRC risk. These results may aid in the development of genetically targeted cancer prevention strategies with NSAIDs.

Nonsteroidal anti-inflammatory drugs (NSAIDs) target the prostaglandin H synthase enzymes, cyclooxygenase (COX)-1 and COX-2, and reduce colorectal cancer risk. Genetic variation in the genes encoding these enzymes may be associated with changes in colon and rectal cancer risk and in NSAID efficacy.We genotyped candidate polymorphisms and tag SNPs in PTGS1 (COX-1) and PTGS2 (COX-2) in a population-based case–control study (Diet, Activity and Lifestyle Study, DALS) of colon cancer (n = 1,470 cases/1,837 controls) and rectal cancer (n = 583/775), and independently among cases and controls from the Colon Cancer Family Registry (CCFR; colon n = 959/1,535, rectal n = 505/839).In PTGS2, a functional polymorphism (-765G[C; rs20417) was associated with a twofold increased rectal cancer risk (p = 0.05) in the DALS. This association replicated with a significant nearly fivefold increased risk of rectal cancer in the CCFR study (ORCC vs. GG = 4.88; 95 % CI 1.54–15.45; ORGC vs. GG = 1.36; 95 %CI 0.95–1.94). Genotype–NSAID interactions were observed in the DALS for PTGS1 and rectal cancer risk and for PTGS2 and colon cancer risk, but were no longer significant after correcting for multiple comparisons and did not replicate in the CCFR. No significant associations between PTGS1 polymorphisms and colon or rectal cancer risk were observed.

Background Toll-like receptors (TLRs) and the transcription factor nuclear factor-κB (NFκB) are important in inflammation and cancer. Methods We examined the association between breast cancer risk and 233 tagging single nucleotide polymorphisms within 31 candidate genes involved in TLR or NFκB pathways. This population-based study in the Seattle area included 845 invasive breast cancer cases, diagnosed between 1997 and 1999, and 807 controls aged 65–79. Results Variant alleles in four genes were associated with breast cancer risk based on gene-level tests: MAP3K1, MMP9, TANK, and TLR9. These results were similar when the risk of breast cancer was examined within ductal and luminal subtypes. Subsequent exploratory pathway analyses using the GRASS algorithm found no associations for genes in TLR or NFκB pathways. Using publicly available CGEMS GWAS data to validate significant findings (N = 1,145 cases, N = 1,142 controls), rs889312 near MAP3K1 was confirmed to be associated with breast cancer risk (P = 0.04, OR 1.15, 95% CI 1.01–1.30). Further, two SNPs in TANK that were significant in our data, rs17705608 (P = 0.05) and rs7309 (P = 0.04), had similar risk estimates in the CGEMS data (rs17705608 OR 0.83, 95% CI 0.72–0.96; CGEMS OR 0.90, 95% CI 0.80–1.01 and rs7309 OR 0.83, 95% CI 0.73–0.95; CGEMS OR 0.91, 95% CI 0.81–1.02). Conclusions Our findings suggest plausible associations between breast cancer risk and genes in TLR or NFκB pathways. Given the few suggestive associations in our data and the compelling biologic rationale for an association between genetic variation in these pathways and breast cancer risk, further studies are warranted that examine these effects.

Proinflammatory cytokines are associated with age-related diseases including arthritis and heart disease. IL6 and TNF also play key roles in estrogen modulation in older women. We explored whether variation in IL6 and TNF genes influenced the risk of breast cancer in samples that differed by age group: T alleles at IL6 rs2069861 among subjects in the oldest age group (OR 1.8, 95% CI 1.1–2.9), but no overall increased risk of breast cancer associated with any IL6 or IL6R variants in the combined data. There were significantly elevated risks of breast cancer among women 45–64 years old associated with a UTR 5′ flanking SNP LTA rs2009658 C>G allele (OR 1.5, 95% CI 1.1–1.9) and a nonsynonomous coding SNP TNFRSF1A rs767455 T>C allele (OR 1.3, 95% CI 1.1–1.6); these two variants were also elevated in the combined data (OR 1.3, 95% CI 1.1–1.5 and OR 1.2, 95% CI 1.1–1.4, respectively). This study supports a modest association between a variant in IL6 and breast cancer among older women and TNF-related variants and breast cancer among middle-aged women. Further evaluation of these genes in other studies is warranted.

Introduction: Inflammation is known to affect the risk of colorectal cancer (CRC). Although use of non-steroidal anti-inflammatory drugs (NSAIDs) is effective in decreasing CRC risk, inherited variation in genes that affect NSAID metabolism may alter their potential as a preventive agent. As NSAIDs function by inhibiting cyclooxygenase (COX) enzymes, which control the conversion of arachidonic acid to prostaglandins and thromboxane, we investigated whether variation in prostaglandin synthesis and related pathways affects the risk of CRC in the Colon Cancer Family Registry. Methods: We examined associations between 221 tagging and candidate single nucleotide polymorphisms (SNPs) within 18 candidate genes (ALOX12, ALOX5, ALOX5AP, CRP, EGFR, GIPC1, HPGD, PARP12, PKN1, PLA2G1B, PTGER1, PTGER2, PTGER3, PTGER4, PTGES, PTGIS, PTGS2, and TBXAS1) and the risk of CRC. We further assessed whether the association between SNPs and CRC risk varied by use of NSAIDs. Using a case-unaffected-sibling-control design, this study included 4,213 Caucasian men and women age 20-74 with no known familial adenomatous polyposis and no personal history of ulcerative colitis or Crohn's disease. Cases (N=1,621) were recruited from six registry centers and were diagnosed with primary invasive CRC from 1998-2002. Controls (N=2,592) were siblings not diagnosed with CRC at the time of ascertainment. We computed odds ratios and 95% confidence intervals using conditional logistic regression and corrected for multiple comparisons using the Conneely and Boehnke method. All models were adjusted for age and sex; NSAID interactions were further adjusted for BMI, smoking, and physical activity. Results: After adjustment for multiple comparisons, two intronic SNPs were associated with a decreased risk of rectal cancer: rs45525634 in PTGER2 (ORhet/hzv 0.49, 95% CI 0.29-0.82) and rs6091005 in PTGIS (ORhet 0.73, 95% CI 0.52-1.03; ORhzv 0.39, 95% CI 0.22-0.68). Additionally, an intronic SNP, rs11571364, in ALOX12 was associated with an increased risk of rectal cancer (ORhet/hzv 1.87, 95% CI 1.19-2.95). The risk of CRC associated with NSAID use differed by genotype at three intronic SNPs: rs10142849 (p = 0.02) in PTGER2 as well as rs11239516 (p = 0.04) and rs2115819 (p = 0.05) in ALOX5. These SNPs in ALOX5 shared low LD among Caucasian controls (r2 = 0.28). There was also evidence for interactions between SNP genotypes and NSAID use for colon cancer for two intronic SNPs, rs11239516 (p = 0.04) in ALOX5 and rs7806848 (p = 0.02) in TBXAS1, and for rectal cancer for rs16870224 (p = 0.05) located in the 3' UTR of PTGER4, the coding SNP rs35416389 (R641Q, p = 0.02) in PKN1, and two intronic SNPs rs17222919 (p = 0.03) in ALOX5 and rs6664305 (p = 0.01) in PTGER3. Conclusion: The results of this study suggest that genetic variation in ALOX12, PTGER2, and PTGIS may affect the risk of rectal cancer. In addition, this study suggests that current use of NSAIDs may modify associations between CRC risk and SNPs in ALOX5, PKN1, PTGER2, PTGER3, PTGER4, and TBXAS1. These findings may aid in the development of genetically targeted cancer prevention strategies with NSAIDs. Citation Format: Alexa J. Resler, Laura Heath, Karen W. Makar, John Whitton, Nina Habermann, Hansong Wang, Polly A. Newcomb, Loic LeMarchand, John D. Potter, Cornelia M. Ulrich. Genetic variation in prostaglandin synthesis and related pathways, NSAID use, and risk of colorectal cancer in the Colon Cancer Family Registry. [abstract]. In: Proceedings of the AACR Special Conference on Post-GWAS Horizons in Molecular Epidemiology: Digging Deeper into the Environment; 2012 Nov 11-14; Hollywood, FL. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2012;21(11 Suppl):Abstract nr 37.

Syndecans are transmembrane proteins on the cell surface that act as receptors for ligands from a number of protein families. These proteoglycans are involved in a large number of important normal cell functions, including signaling, proliferation, and cell-matrix adhesion; they are also involved in aberrant cell functions such as inflammation, angiogenesis, and carcinogenesis. In a recent pathway directed re-analysis of GWAS data (Menashe, Cancer Research, 2010), variants in the syndecan-1 (SDC1) gene on chromosome 2p were found to be associated with excess risk of breast cancer. We sought to extend this finding by investigating the association between tagging SNPs in the SDC1 gene and risk of breast cancer in our case-control study (The PACE study) in older women. This population-based study includes 845 women who were 65-79 years old at diagnosis of incident breast cancer and 807 age-matched unaffected controls from the Seattle area. The analysis was restricted to Caucasian women. A total of 11 SNPs were chosen to represent 10 bins using Snagger, with double coverage of one large bin. Five SNPs failed genotyping, leaving six SNPs for analysis. At the gene level, a P-min permutation test was not significant for SDC1 (p=0.10). Three SNPs, rs10495704, rs2002090, and rs3771240, were in high linkage disequilibrium in our controls (r2 0.81-0.87) and all three were separately associated with marginally increased risks of breast cancer (OR 1.2, 95% CI 1.0-1.4). One of these SNPs, rs10495704, is in the 5’UTR and thus could be associated with protein processing. No excess risk of breast cancer was associated with the other three SDC1 variants: rs4432408, rs2348478, or rs2015110. A similar pattern of marginally increased risk for the three polymorphisms in high LD (rs10495704, rs2002090, and rs3771240) was found in two subsequent analyses restricted to women with ductal histology and luminal type tumors. We are currently assessing other genes related to inflammation that may be related to breast cancer risk in this study population. The data for SDC1 in the PACE study cannot rule out a modest association with breast cancer risk. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5593. doi:10.1158/1538-7445.AM2011-5593