Diabetes mellitus in relation to colorectal tumor molecular subtypes: A pooled analysis of more than 9000 cases.

Diabetes 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.

Methylene Blue for Vasoplegia During Extracorporeal Membrane Oxygenation Support.

OBJECTIVE: The treatment of refractory vasodilatory shock in patients undergoing extracorporeal membrane oxygenation (ECMO) is an area in which there is minimal literature. Based on previous literature, the authors hypothesized that at least 40% of ECMO patients with vasoplegia would respond positively to methylene blue (MB) administration and that those who responded to MB would have increased survival. DESIGN: Retrospective observational study. SETTING: Single institution, quaternary care hospital. PARTICIPANTS: The study comprised 45 patients who received MB for vasoplegia during ECMO. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Of the 45 patients who received MB, 25 patients (55.6%) experienced a ≥10% increase in mean arterial pressure (MAP) and a reduction in norepinephrine dosing in the one-to-two hour interval after MB administration. There was a trend for improvement in survival to discharge for those who responded to MB (32% v 10%; p = 0.15). In addition, patients who did not have at least a >5% increase in MAP (29 experienced a >5% increase and 16 experienced a ≤5% increase) after MB administration, experienced 100% mortality (p = 0.008). CONCLUSION: This study suggested that approximately 50% of ECMO patients with vasoplegia can be expected to respond to MB with a >10% MAP improvement. The lack of a blood pressure response >5% after MB administration may portend poor survival. Larger prospective studies are needed to verify these preliminary results.

Thirty-day readmissions after atrial fibrillation catheter ablation in patients with heart failure.

BACKGROUND: Randomized clinical trial data have demonstrated catheter ablation (CA) as a viable treatment modality for atrial fibrillation (AF). Patients with heart failure (HF) undergoing AF CA appear to derive improvements in quality of life and mortality compared to those treated with medical therapy (MT). Contemporary national data on 30-day readmissions after CA compared to MT among patients with HF are lacking. METHODS: From the 2016 Nationwide Readmissions Databases, 749 776 (weighted national estimate: 1 421 673) AF HF patients were identified of which 2204 (0.3%) underwent CA and 747 572 (99.7%) received MT. Propensity matching balanced baseline clinical characteristics. Thirty-day readmission rates, causes, predictors, and costs of 30-day readmission were compared. RESULTS: Among both the unmatched and matched cohorts, 30-day readmissions were lower for patients treated with CA compared to MT (16.8% vs 20.1%, P < .001 and 16.8% vs 18.8%, P = .020). CA was associated with reduced risk of readmission compared to MT (odds ratio 0.86, 95% confidence interval [CI]: 0.77-0.97). HF exacerbation and arrhythmias were the most common cause for 30-day readmission after CA. CA costs were higher during index hospitalization but similar to MT during readmission among the matched cohort ($15 858 ± $21 636 vs $16 505 ± $29 171, P = .67). Predictors of readmission were largely nonmodifiable risk factors among both the CA and MT groups. CONCLUSIONS: Nearly one in six patients with HF is readmitted within 30-days after undergoing CA. In propensity matched analyses, CA was associated with decreased rate and risk for readmission compared to MT. CA has higher index hospitalization costs, but lower readmission costs.

Comprehensively evaluating cis-regulatory variation in the human prostate transcriptome by using gene-level allele-specific expression.

The identification of cis-acting regulatory variation in primary tissues has the potential to elucidate the genetic basis of complex traits and further our understanding of transcriptomic diversity across cell types. Expression quantitative trait locus (eQTL) association analysis using RNA sequencing (RNA-seq) data can improve upon the detection of cis-acting regulatory variation by leveraging allele-specific expression (ASE) patterns in association analysis. Here, we present a comprehensive evaluation of cis-acting eQTLs by analyzing RNA-seq gene-expression data and genome-wide high-density genotypes from 471 samples of normal primary prostate tissue. Using statistical models that integrate ASE information, we identified extensive cis-eQTLs across the prostate transcriptome and found that approximately 70% of expressed genes corresponded to a significant eQTL at a gene-level false-discovery rate of 0.05. Overall, cis-eQTLs were heavily concentrated near the transcription start and stop sites of affected genes, and effects were negatively correlated with distance. We identified multiple instances of cis-acting co-regulation by using phased genotype data and discovered 233 SNPs as the most strongly associated eQTLs for more than one gene. We also noted significant enrichment (25/50, p = 2E-5) of previously reported prostate cancer risk SNPs in prostate eQTLs. Our results illustrate the benefit of assessing ASE data in cis-eQTL analyses by showing better reproducibility of prior eQTL findings than of eQTL mapping based on total expression alone. Altogether, our analysis provides extensive functional context of thousands of SNPs in prostate tissue, and these results will be of critical value in guiding studies examining disease of the human prostate.

DNA mismatch repair protein deficient non-neoplastic colonic crypts: a novel indicator of Lynch syndrome.

Lynch syndrome is the most common form of hereditary colorectal carcinoma. However, establishing the diagnosis of Lynch syndrome is challenging, and ancillary studies that distinguish between sporadic DNA mismatch repair (MMR) protein deficiency and Lynch syndrome are needed, particularly when germline mutation studies are inconclusive. The aim of this study was to determine if MMR protein-deficient non-neoplastic intestinal crypts can help distinguish between patients with and without Lynch syndrome. We evaluated the expression of MMR proteins in non-neoplastic intestinal mucosa obtained from colorectal surgical resection specimens from patients with Lynch syndrome-associated colorectal carcinoma (n = 52) and patients with colorectal carcinoma without evidence of Lynch syndrome (n = 70), including sporadic MMR protein-deficient colorectal carcinoma (n = 30), MMR protein proficient colorectal carcinoma (n = 30), and "Lynch-like" syndrome (n = 10). MMR protein-deficient non-neoplastic colonic crypts were identified in 19 of 122 (16%) patients. MMR protein-deficient colonic crypts were identified in 18 of 52 (35%) patients with Lynch syndrome compared to only 1 of 70 (1%) patients without Lynch syndrome (p < 0.001). This one patient had "Lynch-like" syndrome and harbored two MSH2-deficient non-neoplastic colonic crypts. MMR protein-deficient non-neoplastic colonic crypts were not identified in patients with sporadic MMR protein-deficient or MMR protein proficient colorectal carcinoma. Our findings suggest that MMR protein-deficient colonic crypts are a novel indicator of Lynch syndrome, and evaluation for MMR protein-deficient crypts may be a helpful addition to Lynch syndrome diagnostics.

FIRE: functional inference of genetic variants that regulate gene expression.

MOTIVATION: Interpreting genetic variation in noncoding regions of the genome is an important challenge for personal genome analysis. One mechanism by which noncoding single nucleotide variants (SNVs) influence downstream phenotypes is through the regulation of gene expression. Methods to predict whether or not individual SNVs are likely to regulate gene expression would aid interpretation of variants of unknown significance identified in whole-genome sequencing studies. RESULTS: We developed FIRE (Functional Inference of Regulators of Expression), a tool to score both noncoding and coding SNVs based on their potential to regulate the expression levels of nearby genes. FIRE consists of 23 random forests trained to recognize SNVs in cis-expression quantitative trait loci (cis-eQTLs) using a set of 92 genomic annotations as predictive features. FIRE scores discriminate cis-eQTL SNVs from non-eQTL SNVs in the training set with a cross-validated area under the receiver operating characteristic curve (AUC) of 0.807, and discriminate cis-eQTL SNVs shared across six populations of different ancestry from non-eQTL SNVs with an AUC of 0.939. FIRE scores are also predictive of cis-eQTL SNVs across a variety of tissue types. AVAILABILITY AND IMPLEMENTATION: FIRE scores for genome-wide SNVs in hg19/GRCh37 are available for download at https://sites.google.com/site/fireregulatoryvariation/. CONTACT: nilah@stanford.edu. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Preoperative three-dimensional echocardiography to assess risk of right ventricular failure after left ventricular assist device surgery.

BACKGROUND: Right ventricular failure (RVF) is associated with significant morbidity after left ventricular assist device (LVAD) surgery. Hemodynamic, clinical, and 2-dimensional echocardiographic variables poorly discriminate patients at risk of RVF. We examined the utility of 3-dimensional echocardiography (3DE) right ventricular (RV) volumetric assessment to identify patients at risk for RVF. METHODS AND RESULTS: RVF was defined as the need for inotropic infusion for >14 days after LVAD surgery or the need for biventricular assist device support. Preoperative RV volumes and ejection fraction (EF) were measured, blinded to clinical data, from transthoracic 3DE full volume data sets in 26 patients. Baseline variables and 3DE RV indices were compared between patients with and without RVF. Twenty-four patients received continuous-flow LVADs, and 2 required biventricular support devices. Ten patients required prolonged inotropes after LVAD placement. Baseline characteristics associated with RVF included higher right atrial pressure, higher right atrial pressure to pulmonary capillary wedge pressure ratio, and lower cardiac index and RV stroke work index (RVSWI). Echocardiographic indices associated with RVF included 3DE indexed RV end-diastolic and end-systolic volumes (RVEDVI and RVESVI) and RV ejection fraction (RVEF). The relationship between 3DE quantification of RV volumes and the development of RVF was independent from RVSWI: RVEDVI: odds ratio (OR) 1.16, 95% confidence interval (CI) 1.00-1.33 (P = .04); RVESVI: OR 1.14, 95% CI 1.01-1.28 (P = .03). CONCLUSIONS: Quantitative 3DE is a promising method for pre-LVAD RV assessment. RV volumes assessed by 3DE are predictive of RVF in LVAD recipients independently from hemodynamic correlates of RV function.

Sequential expression of miR-182 and miR-503 cooperatively targets FBXW7, contributing to the malignant transformation of colon adenoma to adenocarcinoma.

Genetic changes in colon cancer are known to parallel the tissue abnormalities associated with the disease, namely adenoma and adenocarcinoma. The role of microRNA dysregulation in dysplastic progression, however, is not well understood. Here, we show that miR-182 and miR-503 undergo sequential up-regulation and drive the progression of colon adenoma to adenocarcinoma by cooperatively down-regulating the tumour suppressor FBXW7. We identified that increased expression of miR-182 is a feature of adenomas. A subsequent increase in miR-503 expression works cooperatively with miR-182 to induce transformation of an adenoma to adenocarcinoma. We show that introducing miR-503 into AAC1 cells, which are derived from a benign adenoma, confers tumourigenic potential. We also demonstrated that blocking both miR-182 and miR-503 in HCT116 colon cancer cells resulted in increased FBXW7 expression and significantly reduced tumour size in xenograft models. We confirmed relevance of these results in patients by examining the expression levels of miR-182 and miR-503 in over 200 colon cancer patients with 12 year survival outcome data. Decreased patient survival was correlated with elevated expression of both miRNAs, suggesting that elevated levels of both miR-182 and miR-503 define a novel prognostic biomarker for colon cancer patients. In conclusion, we show that a sequential expression of miR-182 and miR-503 in benign adenoma cooperatively regulates the tumour suppressor FBXW7, contributing to the malignant transformation of colon adenoma to adenocarcinoma and miR-182 and miR-503 may prove to be novel therapeutic targets. Array data are available at: http://www.oncomir.umn.edu/

CpG island methylator phenotype underlies sporadic microsatellite instability and is tightly associated with BRAF mutation in colorectal cancer.

Aberrant DNA methylation of CpG islands has been widely observed in human colorectal tumors and is associated with gene silencing when it occurs in promoter areas. A subset of colorectal tumors has an exceptionally high frequency of methylation of some CpG islands, leading to the suggestion of a distinct trait referred to as 'CpG island methylator phenotype', or 'CIMP'. However, the existence of CIMP has been challenged. To resolve this continuing controversy, we conducted a systematic, stepwise screen of 195 CpG island methylation markers using MethyLight technology, involving 295 primary human colorectal tumors and 16,785 separate quantitative analyses. We found that CIMP-positive (CIMP+) tumors convincingly represent a distinct subset, encompassing almost all cases of tumors with BRAF mutation (odds ratio = 203). Sporadic cases of mismatch repair deficiency occur almost exclusively as a consequence of CIMP-associated methylation of MLH1 . We propose a robust new marker panel to classify CIMP+ tumors.

Associations between colorectal cancer molecular markers and pathways with clinicopathologic features in older women.

BACKGROUND & AIMS: Colorectal tumors have a large degree of molecular heterogeneity. Three integrated pathways of carcinogenesis (ie, traditional, alternate, and serrated) have been proposed, based on specific combinations of microsatellite instability (MSI), CpG island methylator phenotype (CIMP), and mutations in BRAF and KRAS. We used resources from the population-based Iowa Women's Health Study (n = 41,836) to associate markers of colorectal tumors, integrated pathways, and clinical and pathology characteristics, including survival times. METHODS: We assessed archived specimens from 732 incident colorectal tumors and characterized them as microsatellite stable (MSS), MSI high or MSI low, CIMP high or CIMP low, CIMP negative, and positive or negative for BRAF and/or KRAS mutations. Informative marker data were collected from 563 tumors (77%), which were assigned to the following integrated pathways: traditional (MSS, CIMP negative, BRAF mutation negative, and KRAS mutation negative; n = 170), alternate (MSS, CIMP low, BRAF mutation negative, and KRAS mutation positive; n = 58), serrated (any MSI, CIMP high, BRAF mutation positive, and KRAS mutation negative; n = 142), or unassigned (n = 193). Multivariable-adjusted Cox proportional hazards regression models were used to assess the associations of interest. RESULTS: Patients' mean age (P = .03) and tumors' anatomic subsite (P = .0001) and grade (P = .0001) were significantly associated with integrated pathway assignment. Colorectal cancer (CRC) mortality was not associated with the traditional, alternate, or serrated pathways, but was associated with a subset of pathway-unassigned tumors (MSS or MSI low, CIMP negative, BRAF mutation negative, and KRAS mutation positive) (n = 96 cases; relative risk = 1.76; 95% confidence interval, 1.07-2.89, compared with the traditional pathway). CONCLUSIONS: We identified clinical and pathology features associated with molecularly defined CRC subtypes. However, additional studies are needed to determine how these features might influence prognosis.

Genotoxic colibactin mutational signature in colorectal cancer is associated with clinicopathological features, specific genomic alterations and better survival.

Background and Aims: The microbiome has long been suspected of a role in colorectal cancer (CRC) tumorigenesis. The mutational signature SBS88 mechanistically links CRC development with the strain of Escherichia coli harboring the pks island that produces the genotoxin colibactin, but the genomic, pathological and survival characteristics associated with SBS88-positive tumors are unknown. Methods: SBS88-positive CRCs were identified from targeted sequencing data from 5,292 CRCs from 17 studies and tested for their association with clinico-pathological features, oncogenic pathways, genomic characteristics and survival. Results: In total, 7.5% (398/5,292) of the CRCs were SBS88-positive, of which 98.7% (392/398) were microsatellite stable/microsatellite instability low (MSS/MSI-L), compared with 80% (3916/4894) of SBS88 negative tumors (p=1.5x10-28). Analysis of MSS/MSI-L CRCs demonstrated that SBS88 positive CRCs were associated with the distal colon (OR=1.84, 95% CI=1.40-2.42, p=1x10-5) and rectum (OR=1.90, 95% CI=1.44-2.51, p=6x10-6) tumor sites compared with the proximal colon. The top seven recurrent somatic mutations associated with SBS88-positive CRCs demonstrated mutational contexts associated with colibactin-induced DNA damage, the strongest of which was the APC:c.835-8A>G mutation (OR=65.5, 95%CI=39.0-110.0, p=3x10-80). Large copy number alterations (CNAs) including CNA loss on 14q and gains on 13q, 16q and 20p were significantly enriched in SBS88-positive CRCs. SBS88-positive CRCs were associated with better CRC-specific survival (p=0.007; hazard ratio of 0.69, 95% CI=0.52-0.90) when stratified by age, sex, study, and by stage. Conclusion: SBS88-positivity, a biomarker of colibactin-induced DNA damage, can identify a novel subtype of CRC characterized by recurrent somatic mutations, copy number alterations and better survival. These findings provide new insights for treatment and prevention strategies for this subtype of CRC.

Epidemiologic Factors in Relation to Colorectal Cancer Risk and Survival by Genotoxic Colibactin Mutational Signature.

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.

Body size and risk of colorectal cancer molecular defined subtypes and pathways: Mendelian randomization analyses.

BACKGROUND: Obesity has been positively associated with most molecular subtypes of colorectal cancer (CRC); however, the magnitude and the causality of these associations is uncertain. METHODS: We used Mendelian randomization (MR) to examine potential causal relationships between body size traits (body mass index [BMI], waist circumference, and body fat percentage) with risks of Jass classification types and individual subtypes of CRC (microsatellite instability [MSI] status, CpG island methylator phenotype [CIMP] status, BRAF and KRAS mutations). Summary data on tumour markers were obtained from two genetic consortia (CCFR, GECCO). FINDINGS: A 1-standard deviation (SD:5.1 kg/m2) increment in BMI levels was found to increase risks of Jass type 1MSI-high,CIMP-high,BRAF-mutated,KRAS-wildtype (odds ratio [OR]: 2.14, 95% confidence interval [CI]: 1.46, 3.13; p-value = 9 × 10-5) and Jass type 2non-MSI-high,CIMP-high,BRAF-mutated,KRAS-wildtype CRC (OR: 2.20, 95% CI: 1.26, 3.86; p-value = 0.005). The magnitude of these associations was stronger compared with Jass type 4non-MSI-high,CIMP-low/negative,BRAF-wildtype,KRAS-wildtype CRC (p-differences: 0.03 and 0.04, respectively). A 1-SD (SD:13.4 cm) increment in waist circumference increased risk of Jass type 3non-MSI-high,CIMP-low/negative,BRAF-wildtype,KRAS-mutated (OR 1.73, 95% CI: 1.34, 2.25; p-value = 9 × 10-5) that was stronger compared with Jass type 4 CRC (p-difference: 0.03). A higher body fat percentage (SD:8.5%) increased risk of Jass type 1 CRC (OR: 2.59, 95% CI: 1.49, 4.48; p-value = 0.001), which was greater than Jass type 4 CRC (p-difference: 0.03). INTERPRETATION: Body size was more strongly linked to the serrated (Jass types 1 and 2) and alternate (Jass type 3) pathways of colorectal carcinogenesis in comparison to the traditional pathway (Jass type 4). FUNDING: Cancer Research UK, National Institute for Health Research, Medical Research Council, National Institutes of Health, National Cancer Institute, American Institute for Cancer Research, Brigham and Women's Hospital, Prevent Cancer Foundation, Victorian Cancer Agency, Swedish Research Council, Swedish Cancer Society, Region Västerbotten, Knut and Alice Wallenberg Foundation, Lion's Cancer Research Foundation, Insamlingsstiftelsen, Umeå University. Full funding details are provided in acknowledgements.

School attendance in childhood cancer survivors and their siblings.

OBJECTIVE: To investigate school absenteeism among childhood cancer survivors and their siblings and examine factors related to absenteeism in survivors. STUDY DESIGN: A cross-sectional study was conducted among consecutive cancer survivors attending a large pediatric cancer survivor clinic. Absenteeism rates were obtained for survivors and their closest in age sibling from school report cards. Absenteeism was compared with a population control group of 167752 students using 1-sample t tests. The Child Vulnerability Scale, Pediatric Quality of Life Inventory, and Behavior Assessment System for Children were administered to survivors. Univariate and multiple regression analyses assessed variables associated with days absent. RESULTS: One hundred thirty-one survivors (median age at assessment: 13.4 years, range 8.0-19.2; median age at diagnosis: 9.4 years, range 4.3-17.3) and 77 siblings (median age at assessment: 13 years, age range 7-18) participated. Survivors and siblings missed significantly more school days than the population control group (mean ± SD: 9.6 ± 9.2 and 9.9 ± 9.8 vs 5.0 ± 5.6 days, respectively, P < .0001). Among matched survivor-sibling pairs (N = 77), there was no difference in absenteeism (9.6 ± 9.2 vs 9.9 ± 9.8 days, P = .85). Absenteeism in survivors was significantly associated with a low Pediatric Quality of Life Inventory Physical Health Summary Score (P = .01). Parents' perception of their child's vulnerability and emotional and social functioning were not associated with absenteeism. CONCLUSIONS: Childhood cancer survivors and siblings miss more school than the general population. The only predictor of absenteeism in survivors is poor physical quality of health. More research should be devoted to school attendance and other outcomes in siblings of childhood cancer survivors.

Postmenopausal hormone therapy and colorectal cancer risk by molecularly defined subtypes among older women.

BACKGROUND: Postmenopausal hormone (PMH) therapy may reduce colorectal cancer (CRC) risk, but existing data are inconclusive. OBJECTIVES: To evaluate associations between PMH therapy and incident CRC, overall and by molecularly defined subtypes, in the population-based Iowa Women's Health Study of older women. METHODS: Exposure data were collected from Iowa Women's Health Study participants (55-69 years) at baseline (1986). Archived, paraffin-embedded tissue specimens for 553 CRC cases were collected and analysed to determine microsatellite instability (MSI-L/MSS or MSI-H), CpG island methylator phenotype (CIMP-negative or CIMP-positive) and BRAF mutation (BRAF-wildtype or BRAF-mutated) status. Multivariable Cox regression models were fit to estimate RRs and 95% CIs. RESULTS: PMH therapy (ever vs never use) was inversely associated with incident CRC overall (RR=0.82; 95% CI 0.72 to 0.93), with a significantly lower risk for MSI-L/MSS tumours (RR=0.75; 95% CI 0.60 to 0.94), and borderline significantly lower risks for CIMP-negative (RR=0.79; 95% CI 0.63 to 1.01) and BRAF-wildtype (RR=0.83; 95% CI 0.66 to 1.04) tumours. For PMH therapy >5 years, the subtype-specific risk estimates for MSI-L/MSS, CIMP-negative and BRAF-wildtype tumours were: RR=0.60, 95% CI 0.40 to 0.91; RR=0.68, 95% CI 0.45 to 1.03; and RR=0.70, 95% CI 0.47 to 1.05, respectively. PMH therapy was not significantly associated with the MSI-H, CIMP-positive or BRAF-mutated CRC subtypes. CONCLUSIONS: In this prospective cohort study, PMH therapy was inversely associated with distinct molecularly defined CRC subtypes, which may be related to differential effects from oestrogen and/or progestin on heterogeneous pathways of colorectal carcinogenesis.

A High-Throughput Workflow for FFPE Tissue Proteomics.

Laser capture microdissection (LCM) has become an indispensable tool for mass spectrometry-based proteomic analysis of specific regions obtained from formalin-fixed paraffin-embedded (FFPE) tissue samples in both clinical and research settings. Low protein yields from LCM samples along with laborious sample processing steps present challenges for proteomic analysis without sacrificing protein and peptide recovery. Automation of sample preparation workflows is still under development, especially for samples such as laser-capture microdissected tissues. Here, we present a simplified and rapid workflow using adaptive focused acoustics (AFA) technology for sample processing for high-throughput FFPE-based proteomics. We evaluated three different workflows: standard extraction method followed by overnight trypsin digestion, AFA-assisted extraction and overnight trypsin digestion, and AFA-assisted extraction simultaneously performed with trypsin digestion. The use of AFA-based ultrasonication enables automated sample processing for high-throughput proteomic analysis of LCM-FFPE tissues in 96-well and 384-well formats. Further, accelerated trypsin digestion combined with AFA dramatically reduced the overall processing times. LC-MS/MS analysis revealed a slightly higher number of protein and peptide identifications in AFA accelerated workflows compared to standard and AFA overnight workflows. Further, we did not observe any difference in the proportion of peptides identified with missed cleavages or deamidated peptides across the three different workflows. Overall, our results demonstrate that the workflow described in this study enables rapid and high-throughput sample processing with greatly reduced sample handling, which is amenable to automation.

Body mass index and molecular subtypes of colorectal cancer.

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.