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Early detection of ovarian cancer has the potential to impact mortality. A multimodal screening strategy where rising CA125 values over time, analyzed with the risk of ovarian cancer algorithm (ROCA), triggers transvaginal sonography and possible surgery has high sensitivity and specificity, but still fails to detect the 20% of early-stage cases that do not express CA125. Use of multiple biomarkers could detect cases missed by CA125. We have studied the sensitivity and lead time of a multi-marker panel (CA125, HE4, MMP-7, and CA 72-4) compared with CA125 alone. We used PRoBE design principles to select preclinical longitudinal specimens from 75 women (50 screen-positive, 25 screen-negative) who developed invasive epithelial ovarian cancer (3–5 serial specimens each) and 547 corresponding healthy controls (1–10 serial specimens each) from the ovarian cancer screening trial, UKCTOCS, in a blinded fashion. We measured the multi-marker concentrations in ultra-low serum volumes (16 μL) utilizing multiplexed bead-based immunoassays with low detection limits, high inter- and intra-assay precision, negligible cross-reactivity, and good correlation with standard immunoassays. While, at least one of the complementary biomarkers rose with CA125 in 44% (22/50) of screen-positive cases, there was no advantage in lead time over CA125. Therefore, we developed single-marker longitudinal algorithms (ROCA-like) to determine the presence of a change point to distinguish between the cases and controls. Using these algorithms, at 98% specificity, HE4 and CA72-4 identified 16% (4/25) of screen-negative cases, while MMP-7 identified none. Taken together, HE4 and CA72-4 show promise as complementary biomarkers to CA125 for longitudinal screening.

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Supplementary figures, tables and materials & methods Supplementary Figure 1. Schematic presentation of steps in immunoassay development, validation and studies for screening TP53 autoantibody in human serum samples. Supplementary Figure 2. Identifying a common cut-off value for TP53 autoantibody immunoassay. Supplementary Table 1. Characteristics of patients with invasive epithelial ovarian/tubal/peritoneal cancer in MDACC-NROSS set Supplementary Table 2. Characteristics of patients with invasive epithelial ovarian/tubal/peritoneal cancer in AOCS set Supplementary Table 3. Characteristics of patients with invasive epithelial ovarian/tubal/peritoneal cancer by stage and primary site in the UKCTOCS study Supplementary Figure 3. Examples of longitudinal analysis of CA125 and TP53 autoantibody titers in pre-diagnostic serial serum samples from ovarian cancer patients in the UKCTOCS Study. Supplementary Figure 4. ROC curve analysis for TP53 autoantibody and CA125 biomarkers in UKCTOCS trial. Supplementary Figure 5. Comparison of TP53 autoantibody titers between cancer cases with TP53 wild-type and mutant genes in the AOCS biobanking study. Supplementary Figure 6. The list of TP53 mutant protein candidates for preliminary screening of autoantibody against specific TP53 mutant proteins. Supplementary Table 4. Summary of specific TP53 mutant autoantibody titers of individual cancer patients with corresponding TP53 protein mutations in the AOCS biobanking study Supplementary Table 5. Summary of multiplex immunoassay results for screening TP53 mutant-specific autoantibody using samples from the AOCS biobanking study.

PDF file - 206K, Table S1 Numbers of test positive cases and controls using CA19-9 37 U/mL and CA125 30 U/mL cut-offs Figure S1 Scatter plots showing distribution of CA19-9, CA125, CEACAM1 and REG3A levels against time to diagnosis for discovery set. Zero represents the point of clinical diagnosis. Figure S2 Examples of CA19-9 and CA125 levels in individual cases with serial/longitudinal samples. Figure S3 Box and whisker plots showing serum levels of CA19-9 and CA125 for case control validation samples grouped into different time to diagnosis groups. Whisker limits represent the 5th and 95th percentiles, the box limits represent interquartile range, the horizontal line the median and the cross the mean. Case and control groups were compared using the Mann-Whitney test; significant P values (

Purpose: Biomarkers for the early detection of pancreatic cancer are urgently needed. The primary objective of this study was to evaluate whether increased levels of serum CA19-9, CA125, CEACAM1, and REG3A are present before clinical presentation of pancreatic cancer and to assess the performance of combined markers for early detection and prognosis.Experimental Design: This nested case–control study within the UKCTOCS included 118 single and 143 serial serum samples from 154 postmenopausal women who were subsequently diagnosed with pancreatic cancer and 304 matched noncancer controls. Samples were split randomly into independent training and test sets. CA19-9, CA125, CEACAM1, and REG3A were measured using ELISA and/or CLIA. Performance of markers to detect cancers at different times before diagnosis and for prognosis was evaluated.Results: At 95% specificity, CA19-9 (>37 U/mL) had a sensitivity of 68% up to 1 year, and 53% up to 2 years before diagnosis. Combining CA19-9 and CA125 improved sensitivity as CA125 was elevated (>30 U/mL) in approximately 20% of CA19-9–negative cases. CEACAM1 and REG3A were late markers adding little in combined models. Average lead times of 20 to 23 months were estimated for test-positive cases. Prediagnostic levels of CA19-9 and CA125 were associated with poor overall survival (HR, 2.69 and 3.15, respectively).Conclusions: CA19-9 and CA125 have encouraging sensitivity for detecting preclinical pancreatic cancer, and both markers can be used as prognostic tools. This work challenges the prevailing view that CA19-9 is upregulated late in the course of pancreatic cancer development. Clin Cancer Res; 21(3); 622–31. ©2014 AACR.

Purpose: The TP53 tumor-suppressor gene is mutated in >95% of high-grade serous ovarian cancers. Detecting an autologous antibody response to TP53 that might improve early detection.Experimental Design: An immunoassay was developed to measure TP53 autoantibody in sera from 378 cases of invasive epithelial ovarian cancer and 944 age-matched healthy controls from the United States, Australia, and the United Kingdom. Serial preclinical samples from cases and controls were also assayed from the UK Collaborative Trial of Ovarian Cancer Screening (UKCTOCS).Results: Using a cutoff value of 78 U/mL to achieve a specificity of 97.4%, TP53 autoantibody was elevated in 30% of 50 cases from MD Anderson, 21.3% of 108 cases from the Australian Ovarian Cancer Study, and 21% of 220 cases from the UKCTOCS. Among 164 cases with rising CA125 detected with the UKCTOCS risk of ovarian cancer algorithm (ROCA), 20.7% had elevated TP53 autoantibody. In cases missed by the ROCA, 16% of cases had elevated TP53 autoantibody. Of the 34 ovarian cancer cases detected with the ROCA, TP53 autoantibody titers were elevated 11.0 months before CA125. In the 9 cases missed by the ROCA, TP53 autoantibody was elevated 22.9 months before cancer diagnosis. Similar sensitivity was obtained using assays with specific mutant and wild-type TP53.Conclusions: TP53 autoantibody levels provide a biomarker with clinically significant lead time over elevation of CA125 or an elevated ROCA value. Quantitative assessment of autoantibodies in combination with CA125 holds promise for earlier detection of invasive epithelial ovarian cancer. Clin Cancer Res; 23(19); 5912–22. ©2017 AACR.

International Cancer Benchmarking Partnership Module 4 reports the first international comparison of ovarian cancer (OC) diagnosis routes and intervals (symptom onset to treatment start), which may inform previously reported variations in survival and stage.Data were collated from 1110 newly diagnosed OC patients aged40 surveyed between 2013 and 2015 across five countries (51-272 per jurisdiction), their primary-care physicians (PCPs) and cancer treatment specialists, supplement by treatment records or clinical databases. Diagnosis routes and time interval differences using quantile regression with reference to Denmark (largest survey response) were calculated.There were no significant jurisdictional differences in the proportion diagnosed with symptoms on the Goff Symptom Index (53%; P = 0.179) or National Institute for Health and Care Excellence NG12 guidelines (62%; P = 0.946). Though the main diagnosis route consistently involved primary-care presentation (63-86%; P = 0.068), onward urgent referral rates varied significantly (29-79%; P 0.001). In most jurisdictions, diagnostic intervals were generally shorter and other intervals, in particular, treatment longer compared to Denmark.This study highlights key intervals in the diagnostic pathway where improvements could be made. It provides the opportunity to consider the systems and approaches across different jurisdictions that might allow for more timely ovarian cancer diagnosis and treatment.

Purpose: The TP53 tumor-suppressor gene is mutated in >95% of high-grade serous ovarian cancers. Detecting an autologous antibody response to TP53 that might improve early detection. Experimental Design: An immunoassay was developed to measure TP53 autoantibody in sera from 378 cases of invasive epithelial ovarian cancer and 944 age-matched healthy controls from the United States, Australia, and the United Kingdom. Serial preclinical samples from cases and controls were also assayed from the UK Collaborative Trial of Ovarian Cancer Screening (UKCTOCS). Results: Using a cutoff value of 78 U/mL to achieve a specificity of 97.4%, TP53 autoantibody was elevated in 30% of 50 cases from MD Anderson, 21.3% of 108 cases from the Australian Ovarian Cancer Study, and 21% of 220 cases from the UKCTOCS. Among 164 cases with rising CA125 detected with the UKCTOCS risk of ovarian cancer algorithm (ROCA), 20.7% had elevated TP53 autoantibody. In cases missed by the ROCA, 16% of cases had elevated TP53 autoantibody. Of the 34 ovarian cancer cases detected with the ROCA, TP53 autoantibody titers were elevated 11.0 months before CA125. In the 9 cases missed by the ROCA, TP53 autoantibody was elevated 22.9 months before cancer diagnosis. Similar sensitivity was obtained using assays with specific mutant and wild-type TP53. Conclusions: TP53 autoantibody levels provide a biomarker with clinically significant lead time over elevation of CA125 or an elevated ROCA value. Quantitative assessment of autoantibodies in combination with CA125 holds promise for earlier detection of invasive epithelial ovarian cancer. Clin Cancer Res; 23(19); 5912–22. ©2017 AACR.

// Stefan Kiechl 1 , Daniel Schramek 2,3 , Martin Widschwendter 4 , Evangelia-Ourania Fourkala 4 , Alexey Zaikin 4,5 , Allison Jones 5 , Bernadette Jaeger 4 , Brigitte Rack 4 , Wolfgang Janni 6 , Christoph Scholz 7 , Johann Willeit 1 , Siegfried Weger 8 , Agnes Mayr 8 , Andrew Teschendorff 9 , Adam Rosenthal 10 , Lindsay Fraser 5 , Susan Philpott 5 , Louis Dubeau 11 , Mohammed Keshtgar 12 , Rebecca Roylance 10 , Ian J. Jacobs 4,13 , Usha Menon 4 , Georg Schett 14 and Josef M. Penninger 2 1 Department of Neurology, Medical University of Innsbruck, Innsbruck, Austria 2 IMBA, Institute of Molecular Biotechnology of the Austrian Academy of Sciences, Vienna, Austria 3 The Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, Ontario, Canada 4 Department of Women’s Cancer, EGA Institute of Women’s Health, University College London, London, United Kingdom 5 Department of Mathematics, University College London, London, United Kingdom 6 Department of Gynecology and Obstetrics, University Duesseldorf, Duesseldorf, Germany 7 Heinrich-Heine-University Dusseldorf, Dusseldorf, Germany 8 Department of Internal Medicine, Bruneck Hospital, Bruneck, Italy 9 Statistical Genomics Group, Paul O’Gorman Building, UCL Cancer Institute, University College London, London, United Kingdom 10 Barts Cancer Institute CR UK Centre of Excellence, Queen Mary University of London, Charterhouse Square, London, United Kingdom 11 Department of Pathology, USC/Norris Comprehensive Cancer Center, Keck School of Medicine, University of Southern California, Los Angeles, California, USA 12 Department of Surgery, Royal Free and University College London Medical School, London, United Kingdom 13 UNSW Australia, Sydney, New South Wales, Australia 14 Department of Internal Medicine 3, University of Erlangen-Nuremberg, Erlangen, Germany Correspondence to: Josef M. Penninger, email: // Keywords : breast cancer, RANKL/RANK, Gerotarget Received : October 10, 2016 Accepted : December 05, 2016 Published : December 18, 2016 Abstract Breast cancer is the most common female cancer, affecting approximately one in eight women during their lifetime in North America and Europe. Receptor Activator of NF-kB Ligand (RANKL), its receptor RANK and the natural antagonist osteoprotegerin (OPG) are essential regulators of bone resorption. We have initially shown that RANKL/RANK are essential for hormone-driven mammary epithelial proliferation in pregnancy and RANKL/RANK have been implicated in mammary stem cell biology. Using genetic mouse-models, we and others identified the RANKL/RANK system as a key regulator of sex hormone, BRCA1 -mutation, and oncogene-driven breast cancer and we proposed that RANKL/RANK might be involved in the initiation of breast tumors. We now report that in postmenopausal women without known genetic predisposition, high RANKL and progesterone serum levels stratify a subpopulation of women at high risk of developing breast cancer 12-24 months before diagnosis (5.33-fold risk, 95%CI 1.5-25.4; P= 0.02). In women with established breast cancer, we demonstrate that RANKL/OPG ratios change dependent on the presence of circulating tumor cells (CTCs). Finally, we show in a prospective human breast cancer cohort that alterations in RANKL/OPG ratios are significantly associated with breast cancer manifestation. These data indicate that the RANKL/RANK/OPG system is deregulated in post-menopausal women at high risk for breast cancer and in women with circulating tumor cells. Thus, serum levels of RANKL/OPG are potentially indicative of predisposition and progression of breast cancer in humans. Advancement of our findings towards clinical application awaits prior validation in independent patient cohorts.

Early detection of ovarian cancer has the potential to impact mortality. A multimodal screening strategy where rising CA125 values over time, analyzed with the risk of ovarian cancer algorithm (ROCA), triggers transvaginal sonography and possible surgery has high sensitivity and specificity, but still fails to detect the 20% of early-stage cases that do not express CA125. Use of multiple biomarkers could detect cases missed by CA125. We have studied the sensitivity and lead time of a multi-marker panel (CA125, HE4, MMP-7, and CA 72-4) compared with CA125 alone. We used PRoBE design principles to select preclinical longitudinal specimens from 75 women (50 screen-positive, 25 screen-negative) who developed invasive epithelial ovarian cancer (3–5 serial specimens each) and 547 corresponding healthy controls (1–10 serial specimens each) from the ovarian cancer screening trial, UKCTOCS, in a blinded fashion. We measured the multi-marker concentrations in ultra-low serum volumes (16 μL) utilizing multiplexed bead-based immunoassays with low detection limits, high inter- and intra-assay precision, negligible cross-reactivity, and good correlation with standard immunoassays. While, at least one of the complementary biomarkers rose with CA125 in 44% (22/50) of screen-positive cases, there was no advantage in lead time over CA125. Therefore, we developed single-marker longitudinal algorithms (ROCA-like) to determine the presence of a change point to distinguish between the cases and controls. Using these algorithms, at 98% specificity, HE4 and CA72-4 identified 16% (4/25) of screen-negative cases, while MMP-7 identified none. Taken together, HE4 and CA72-4 show promise as complementary biomarkers to CA125 for longitudinal screening.

Highlights • Ninety-two & 99% of colorectal cancers were registered after one & six years. • Hospital Episode Statistics tend to capture events unregistered after one year. • Self-reporting by women aged ≥50 was less reliable and had low respondents. • Electronic health records in the UK are suitable for studying colorectal cancer in women., Background Electronic health records are frequently used for cancer epidemiology. We report on their quality for ascertaining colorectal cancer (CRC) in UK women. Methods Population-based, retrospective cohort study nested within the UK Collaborative Trial of Ovarian Cancer Screening (UKCTOCS). Postmenopausal women aged 50–74 who were diagnosed with CRC during 2001–11 following randomisation to the UKCTOCS were identified and their diagnosis confirmed with their treating clinician. The sensitivity and positive predictive value (PPV) of cancer and death registries, hospital episode statistics, and self-reporting were calculated by pairwise comparisons to the treating clinician’s confirmation, while specificity and negative predictive value were estimated relative to expected cases. Results Notification of CRC events were received for 1,085 women as of 24 May 2011. Responses were received from 61% (660/1,085) of clinicians contacted. Nineteen women were excluded (18 no diagnosis date, one diagnosed after cut-off). Of the 641 eligible, 514 had CRC, 24 had a benign polyp, and 103 had neither diagnosis. The sensitivity of cancer registrations at one- and six-years post-diagnosis was 92 (95% CI 90–94) and 99% (97–100), respectively, with a PPV of 95% (95% CI 92/93–97). The sensitivity & PPV of cancer registrations (at one-year post-diagnosis) & hospital episode statistics combined were 98 (96–99) and 92% (89–94), respectively. Conclusions Cancer and death registrations in the UK are a reliable resource for CRC ascertainment in women. Hospital episode statistics can supplement delays in cancer registration. Self-reporting seems less reliable.

Purpose: Biomarkers for the early detection of pancreatic cancer are urgently needed. The primary objective of this study was to evaluate whether increased levels of serum CA19-9, CA125, CEACAM1, and REG3A are present before clinical presentation of pancreatic cancer and to assess the performance of combined markers for early detection and prognosis. Experimental Design: This nested case–control study within the UKCTOCS included 118 single and 143 serial serum samples from 154 postmenopausal women who were subsequently diagnosed with pancreatic cancer and 304 matched noncancer controls. Samples were split randomly into independent training and test sets. CA19-9, CA125, CEACAM1, and REG3A were measured using ELISA and/or CLIA. Performance of markers to detect cancers at different times before diagnosis and for prognosis was evaluated. Results: At 95% specificity, CA19-9 (>37 U/mL) had a sensitivity of 68% up to 1 year, and 53% up to 2 years before diagnosis. Combining CA19-9 and CA125 improved sensitivity as CA125 was elevated (>30 U/mL) in approximately 20% of CA19-9–negative cases. CEACAM1 and REG3A were late markers adding little in combined models. Average lead times of 20 to 23 months were estimated for test-positive cases. Prediagnostic levels of CA19-9 and CA125 were associated with poor overall survival (HR, 2.69 and 3.15, respectively). Conclusions: CA19-9 and CA125 have encouraging sensitivity for detecting preclinical pancreatic cancer, and both markers can be used as prognostic tools. This work challenges the prevailing view that CA19-9 is upregulated late in the course of pancreatic cancer development. Clin Cancer Res; 21(3); 622–31. ©2014 AACR.

Supplemental Digital Content is available in the text., Background— Adiponectin, a circulating adipocyte-derived protein, has insulin-sensitizing, anti-inflammatory, antiatherogenic, and cardiomyocyte-protective properties in animal models. However, the systemic effects of adiponectin in humans are unknown. Our aims were to define the metabolic profile associated with higher blood adiponectin concentration and investigate whether variation in adiponectin concentration affects the systemic metabolic profile. Methods and Results— We applied multivariable regression in ≤5909 adults and Mendelian randomization (using cis-acting genetic variants in the vicinity of the adiponectin gene as instrumental variables) for analyzing the causal effect of adiponectin in the metabolic profile of ≤37 545 adults. Participants were largely European from 6 longitudinal studies and 1 genome-wide association consortium. In the multivariable regression analyses, higher circulating adiponectin was associated with higher high-density lipoprotein lipids and lower very-low-density lipoprotein lipids, glucose levels, branched-chain amino acids, and inflammatory markers. However, these findings were not supported by Mendelian randomization analyses for most metabolites. Findings were consistent between sexes and after excluding high-risk groups (defined by age and occurrence of previous cardiovascular event) and 1 study with admixed population. Conclusions— Our findings indicate that blood adiponectin concentration is more likely to be an epiphenomenon in the context of metabolic disease than a key determinant.

Background: It has been suggested that lower UK cancer survival may be due to incomplete case ascertainment by cancer registries. Methods: We assessed concordance between self-reported breast, bowel and lung cancer and cancer registration (CR) for 1995–2007 in England and Wales in the UK Collaborative Trial of Ovarian Cancer Screening. Results: Concordance of breast cancer CR was higher (94.7%:95% CI: 94.1–95.3%) than for bowel (85.1%:95% CI: 82.1–87.8%) and lung (85.4%:95% CI: 76.3–92.0%). CR concordance was lower in breast cancer (94.5% vs 98.8%) survivors compared with deceased but the difference was small. No difference was found for bowel (85.3% vs 94.6%) or lung (87.1% vs 90.5%) cancer. Conclusion: Concordance of CR and self-reported cancer is high. Incomplete registration is unlikely to be a major cause of lower UK survival rates.

Background: Autoantibodies have been detected in sera before diagnosis of cancer leading to interest in their potential as screening/early detection biomarkers. As we have found autoantibodies to MUC1 glycopeptides to be elevated in early-stage breast cancer patients, in this study we analysed these autoantibodies in large population cohorts of sera taken before cancer diagnosis. Methods: Serum samples from women who subsequently developed breast cancer, and aged-matched controls, were identified from UK Collaborative Trial of Ovarian Cancer Screening (UKCTOCS) and Guernsey serum banks to formed discovery and validation sets. These were screened on a microarray platform of 60mer MUC1 glycopeptides and recombinant MUC1 containing 16 tandem repeats. Additional case–control sets comprised of women who subsequently developed ovarian, pancreatic and lung cancer were also screened on the arrays. Results: In the discovery (273 cases, 273 controls) and the two validation sets (UKCTOCS 426 cases, 426 controls; Guernsey 303 cases and 606 controls), no differences were found in autoantibody reactivity to MUC1 tandem repeat peptide or glycoforms between cases and controls. Furthermore, no differences were observed between ovarian, pancreatic and lung cancer cases and controls. Conclusion: This robust, validated study shows autoantibodies to MUC1 peptide or glycopeptides cannot be used for breast, ovarian, lung or pancreatic cancer screening. This has significant implications for research on the use of MUC1 in cancer detection.

Purpose Ovarian cancer (OC) is the most lethal gynaecological cancer. Early detection is required to improve patient survival. Risk estimation models were constructed for Type I (Model I) and Type II (Model II) OC from analysis of Protein Z, Fibronectin, C-reactive protein and CA125 levels in prospectively collected samples from the United Kingdom Collaborative Trial of Ovarian Cancer Screening (UKCTOCS). Results Model I identifies cancers earlier than CA125 alone, with a potential lead time of 3-4 years. Model II detects a number of high grade serous cancers at an earlier stage (Stage I/II) than CA125 alone, with a potential lead time of 2-3 years and assigns high risk to patients that the ROCA Algorithm classified as normal. Materials and Methods This nested case control study included 418 individual serum samples serially collected from 49 OC cases and 31 controls up to six years pre-diagnosis. Discriminatory logit models were built combining the ELISA results for candidate proteins with CA125 levels. Conclusions These models have encouraging sensitivities for detecting pre-clinical ovarian cancer, demonstrating improved sensitivity compared to CA125 alone. In addition we demonstrate how the models improve on ROCA for some cases and outline their potential future use as clinical tools.

Objectives This paper describes the methods used in the International Cancer Benchmarking Partnership Module 4 Survey (ICBPM4) which examines time intervals and routes to cancer diagnosis in 10 jurisdictions. We present the study design with defining and measuring time intervals, identifying patients with cancer, questionnaire development, data management and analyses.Design and setting Recruitment of participants to the ICBPM4 survey is based on cancer registries in each jurisdiction. Questionnaires draw on previous instruments and have been through a process of cognitive testing and piloting in three jurisdictions followed by standardised translation and adaptation. Data analysis focuses on comparing differences in time intervals and routes to diagnosis in the jurisdictions.Participants Our target is 200 patients with symptomatic breast, lung, colorectal and ovarian cancer in each jurisdiction. Patients are approached directly or via their primary care physician (PCP). Patients’ PCPs and cancer treatment specialists (CTSs) are surveyed, and ‘data rules’ are applied to combine and reconcile conflicting information. Where CTS information is unavailable, audit information is sought from treatment records and databases.Main outcomes Reliability testing of the patient questionnaire showed that agreement was complete (κ=1) in four items and substantial (κ=0.8, 95% CI 0.333 to 1) in one item. The identification of eligible patients is sufficient to meet the targets for breast, lung and colorectal cancer. Initial patient and PCP survey response rates from the UK and Sweden are comparable with similar published surveys. Data collection was completed in early 2016 for all cancer types.Conclusion An international questionnaire-based survey of patients with cancer, PCPs and CTSs has been developed and launched in 10 jurisdictions. ICBPM4 will help to further understand international differences in cancer survival by comparing time intervals and routes to cancer diagnosis.

Background: Recent reports from cancer screening trials in high-risk populations suggest that autoantibodies can be detected before clinical diagnosis. However, there is minimal data on the role of autoantibody signatures in cancer screening in the general population. Methods: Informative p53 peptides were identified in sera from patients with colorectal cancer using an autoantibody microarray with 15-mer overlapping peptides covering the complete p53 sequence. The selected peptides were evaluated in a blinded case–control study using stored serum from the multimodal arm of the United Kingdom Collaborative Trial of Ovarian Cancer Screening where women gave annual blood samples. Cases were postmenopausal women who developed colorectal cancer following recruitment, with 2 or more serum samples preceding diagnosis. Controls were age-matched women with no history of cancer. Results: The 50 640 women randomised to the multimodal group were followed up for a median of 6.8 (inter-quartile range 5.9–8.4) years. Colorectal cancer notification was received in 101 women with serial samples of whom 97 (297 samples) had given consent for secondary studies. They were matched 1 : 1 with 97 controls (296 serial samples). The four most informative peptides identified 25.8% of colorectal cancer patients with a specificity of 95%. The median lead time was 1.4 (range 0.12–3.8) years before clinical diagnosis. Conclusion: Our findings suggest that in the general population, autoantibody signatures are detectable during preclinical disease and may be of value in cancer screening. In colorectal cancer screening in particular, where the current need is to improve compliance, it suggests that p53 autoantibodies may contribute towards risk stratification.

Background: In research studies, accurate information of cancer diagnosis is crucial. In women with breast cancer (BC), we compare cancer registration (CR) in England/Wales and self-reporting with independent confirmation. Methods: In the UK Collaborative Trial of Ovarian Cancer Screening, notification of BC diagnosed between randomisation and 31 December 2009 was obtained through (1) CR (17 October 2011) and (2) self-reporting using postal-questionnaire. Breast cancer was confirmed using a detailed questionnaire (BC questionnaire BCQ) completed by the treating clinician (gold standard). Apparent sensitivity and positive-predictive value of CR/self-reporting vs BCQ were calculated. Results: Of 1065 women with possible BC notification, diagnosis was confirmed in 932 (87.5%). A total of 3.1% (28 out of 918) of BC CR and 12.4% (128 out of 1032) of women with self-reported BC only had in-situ carcinoma on BCQ. Another 4.6% (43 out of 932) of BCQ-confirmed cancer did not have a BC registration, and 3.6% (34 out of 932) did not self-report BC. Apparent sensitivity of CR and self-reporting vs BCQ were 95.4 and 96.4%, respectively. Positive-predictive value of self-reporting (87.1%) was significantly lower than that of CR (96.8%). Women aged

Postmenopausal women with elevated serum sex steroids have an increased risk of breast cancer. Most of this risk is believed to be exerted through binding of the sex steroids to their receptors. For the first time, we investigate the association of estrogen receptor (ER) and androgen receptor (AR) serum bioactivity (SB) in addition to hormone levels in samples from women with breast cancer collected before diagnosis. Two hundred postmenopausal women participating in the UK Collaborative Trial of Ovarian Cancer Screening who developed ER-positive breast cancer 0.6–5 years after sample donation were identified and matched to 400 controls. ER and AR bioassays were used to measure ERα, ERβ, and AR SB. Androgen and estrogen levels were measured with immunoassays. Subjects were classified according to quintiles of the respective marker among controls and the associations between SB and hormones with breast cancer risk were determined by logistic regression analysis. ERα and ERβ SB were significantly higher before diagnosis compared with controls, while estrogens showed no difference. Women had a twofold increased breast cancer risk if ERα SB (odds ratio (OR), 2.114; 95% confidence interval (CI), 1.050–4.425; P=0.040) was in the top quintile >2 years before diagnosis or estrone (OR, 2.205; 95% CI, 1.104–4.586; P=0.029) was in the top quintile P=0.0003) and testosterone (OR, 2.145; 95% CI, 1.256–3.712; P=0.006) were significantly higher compared with controls and showed a strong association with an almost threefold increased breast cancer risk independent of time to diagnosis. This study provides further evidence on the association of androgens and estrogens with breast cancer. In addition, it reports that high ER but not AR SB is associated with increased breast risk >2 years before diagnosis.

Objective: To evaluate the malignant potential of ultrasound-detected ovarian inclusion cysts in the development of ovarian cancer (OC) in postmenopausal women. Design: Prospective cohort study. Setting: UK Collaborative Trial of Ovarian Cancer Screening (UKCTOCS). Population: Postmenopausal women. Methods: In UKCTOCS, women in the ultrasound group have annual scans. Women with inclusion cysts (single/multiple anechoic £10-mm ovarian cysts) and normal ovaries (both uniform hypoechogenicity) on their first scan were identified and followed up through cancer registry/questionnaires. Main outcome measures: Relative risk (RR) of developing OC, invasive epithelial ovarian cancer (iEOC), breast cancer (BC) and endometrial cancer (EC) in women with inclusion cysts relative to those with normal ovaries. The incidence was compared with UK age-adjusted expected rates (Office for National Statistics, 2005). Results: Postmenopausal women (n = 48 230) attended the year 1(11 June 2001–6 December 2006) screen; 1234 (2.5%) had inclusion cysts alone and 22 914 had normal scans. By 1 November 2009 (median follow-up, 6.13 years; interquartile range,4.96–6.98 years), four, three (one Type II), seven and 22 women with inclusion cysts and 32, 29 (20 Type II), 90 and 397 women with normal ovaries were diagnosed with OC, iEOC, EC and BC,respectively. The RR values for the respective cancers (OC [RR,2.32; confidence interval [CI], 0.86–6.28], iEOC [RR, 1.92; CI,0.62–5.92], EC [RR, 1.44; CI, 0.68–3.05], BC [RR, 1.12; CI,0.73–1.73]) were not increased. There was no difference between the observed versus expected incidence rates for these cancers in women with inclusion cysts. Conclusions: Postmenopausal women with ultrasound-detected inclusion cysts do not seem to be at increased risk of ovarian or breast/endometrial (hormone-dependent) cancers.

We previously demonstrated that methylation of neugogenic differentiation 1 (NEUROD1) gene, a polycomb group target (PCGT) gene is a predictor of response to neoadjuvant chemotherapy in breast cancer. Here, we address the question whether NEUROD1 methylation provides clinical information independent from its expression level, and whether PCGT methylation is homogeneous in breast cancer. We examined: (1) NEUROD1 methylation and mRNA expression in 9 breast cancer cell lines and 63 tumour specimens, (2) DNA methylation in a training set of 55 PCGT genes taken from the centre (TUC) and periphery (TUP) of 15 breast cancer specimens, and compared this with 22 non neoplastic controls, and finally, (3) validated statistically significant genes in an independent set of 20 cases versus 18 controls. 8/9 cell lines demonstrated NEUROD1 methylation, whereas, there was only one cell-line that showed NEUROD1 expression. There was no association between methylation and expression in breast tumour specimens, with only 14% exhibiting NEUROD1 expression. Of the 55 PCGT genes analysed, 24% (13/55) were shown to be cancer specific (p0.05) with a receiver-operating-characteristic (ROC) area-under-the-curve (AUC) of0.7 (range 0.71-0.95). DNA methylation accurately predicted the presence of cancer in both TUC and TUP. DNA methylation of PCGT genes predicts the presence of breast cancer and is not subject to tumour heterogeneity. Further work will reveal if methylation of PCGT genes will serve as a robust means for the clinical detection and assessment of breast cancer.

Introduction Associations of endogenous sex hormone levels and all as well as estrogen-receptor (ER)-positive breast cancers are well described. However, studies investigating their association with ER-negative tumours are limited and none use accurate assays such as mass spectrometry. Methods Within the UK Collaborative Trial of Ovarian Cancer Screening (UKCTOCS), a nested case-control study was undertaken of postmenopausal-women who developed ER-negative (n = 92) or ER-positive (n = 205) breast cancer after sample donation and 297 (1:1) age-matched controls. Androgens (testosterone and androstenedione) were measured using mass spectrometry and estradiol by extraction radioimmunoassay (RIA). Bioavailable estradiol and testosterone were calculated using the total hormone level and the sex hormone-binding globulin concentration. Subjects were classified according to the quartile range among controls. Logistic regression was used to estimate odds-ratio (OR) and 95% confidence-intervals (CI) of the associations between two factors and breast cancer risk. A separate analysis was done by stratifying the women based on whether they provided their samples less than or more than 2 years before diagnosis. Results Estradiol and free estradiol were significantly higher prior to diagnosis of ER-negative breast cancer compared with controls while androgens and SHBG did not show any difference. Estradiol, free estradiol, free testosterone and SHBG were significantly higher before ER-positive breast cancer diagnosis compared with controls. Women had a twofold increased ER-negative breast cancer risk if estradiol and free estradiol were in the top quartile but not androgens (testosterone and androstenedione) or SHBG. These associations remained significant only when samples closer (median 1.1 y before) to diagnosis were analyzed rather than farther from diagnosis (median 2.9 y before). Women had a 2.34 (95% CI: 1.21–4.61, p = 0.001), 2.21 (95% CI: 1.14–4.38, p = 0.001), 2 (95% CI: 1.05–3.89, p = 0.005) fold increased ER-positive breast cancer risk if estradiol, free estradiol and free testosterone respectively were in the top quartile. These associations remained significant regardless of whether the samples were collected less than or more than 2 years prior to diagnosis. Conclusion In postmenopausal women increased estrogens but not androgens are associated with ER-negative breast cancer. Previously reported associations of estradiol and free testosterone with ER-positive breast cancer are confirmed. The use of mass spectrometry and sensitive RIA add validity to these findings.

Background Area-level socioeconomic indicators have been shown to be associated with range of health outcomes. However, only few studies have been able to examine the role of area-level indicators alongside individual-level indicators in large national samples. The aim of this project is to assess the role of Index of Multiple Deprivation (IMD) as well as range of individual factors on all-cause, CVD and cancer mortality in sample of women resident in …

Objectives Several studies suggest that overall and central-obesity are associated with increased breast cancer (BC) risk in postmenopausal-women. However, there are no studies investigating changes of central obesity and BC. We report on the association of BC risk with self-reported skirt size (SS; waist-circumference proxy) changes between 20s and postmenopausal-age. Design Prospective cohort-study. Setting UK Collaborative Trial of Ovarian Cancer Screening (UKCTOCS) involving the nine trial centres in England. Participants Postmenopausal-women aged >50 with no known history of BC prior to or on the day of completion of the study-entry questionnaire. Interventions At recruitment and at study entry, women were asked to complete a questionnaire. Women were followed-up via ‘flagging’ at the NHS Information Centre in England and the Hospital Episode Statistics. Main outcome-measure Time to initial BC diagnosis. Results Between 2 January 2005 and 1 July 2010, 92 834 UKCTOCS participants (median age 64.0) completed the study-entry questionnaire. During median follow-up of 3.19 years (25th–75th centile: 2.46–3.78), 1090 women developed BC. Model adjusted analysis for potential confounders showed body mass index (BMI) at recruitment to UKCTOCS (HR for a 5 unit change=1.076, 95% CI 1.012 to 1.136), current SS at study entry (HR=1.051; 95% CI 1.014 to 1.089) and change in SS per 10 years (CSS) (HR=1.330; 95% CI 1.121 to 1.579) were associated with increased BC risk but not SS at 25 (HR=1.006; 95% CI 0.958 to 1.056). CSS was the most predictive singe adiposity measure and further analysis including both CSS and BMI in the model revealed CSS remained significant (HR=1.266; 95% CI 1.041 to 1.538) but not BMI (HR=1.037; 95% CI 0.970 to 1.109). Conclusions CSS is associated with BC risk independent of BMI. A unit increase in UK SS (eg, 12–14) every 10-years between 25 and postmenopausal-age is associated with postmenopausal BC risk by 33%. Validation of these results could provide women with a simple and easy to understand message. Trial registration number ISRCTN22488978.

Purpose Ovarian cancer is a devastating disease and biomarkers for its early diagnosis are urgently required. Serum may be a valuable source of biomarkers that may be revealed by proteomic profiling. Herein, complementary serum protein profiling strategies were employed for discovery of biomarkers that could discriminate cases of malignant and benign ovarian cancer. Experimental design Identically collected and processed serum samples from 22 cases of invasive epithelial ovarian cancer, 45 benign ovarian neoplasms, and 64 healthy volunteers were subjected to immunodepletion and protein equalization coupled to 2D‐DIGE/MS and multidimensional fractionation coupled to SELDI‐TOF profiling with MS/MS for protein identification. Selected candidates were verified by ELISA in samples from malignant (n = 70) and benign (n = 89) cases and combined marker panels tested against serum CA125. Results Both profiling platforms were complementary in identifying biomarker candidates, four of which (A1AT, SLPI, APOA4, VDBP) significantly discriminated malignant from benign cases. However, no combination of markers was as good as CA125 for diagnostic accuracy. SLPI was further tested as an early marker using prediagnosis serum samples. While it rose in cases toward diagnosis, it did not discriminate prediagnosis cases from controls. Conclusions and clinical relevance The candidate biomarkers warrant further validation in independent sample sets.

Objective Hopelessness is an important construct in psychosocial epidemiology, but there is great pressure on the length of questionnaire measures in large-scale population and clinical studies. We examined the validity and test–retest reliability of two brief measures of hopelessness, an existing negatively worded two-item measure of hopelessness (Brief-H-Neg) and a positively worded version of the same instrument (Brief-H-Pos). Design Cohort study. Setting Control arm of the UK Collaborative Trial of Ovarian Cancer Screening. Participants A non-clinical research-based sample of 5000 postmenopausal women selected from 56 512 participants. Primary and secondary outcome measures Spearman's rank correlation of brief measures of hopelessness with the Beck Hopelessness Scale (BHS). Spearman's rank correlation with the Centre for Epidemiological Studies Depression Scale (CES-D) and change in mean score on repeat testing. Methods Two short hopelessness measures, a negatively worded brief measure of hopelessness (Brief-H-Neg) and a positively worded brief measure of hopelessness (Brief-H-Pos), were administered by postal questionnaire to 5000 women together with the 20-item BHS and 20-item CES-D. The Brief-H-Neg and Brief-H-Pos were readministered to 500 women after a 2-week interval. Results 2413 postmenopausal women (mean age 68.9 years) completed the questionnaire. The Brief-H-Neg and Brief-H-Pos correlated 0.93 and 0.87 with the BHS after correction for attenuation and their association with the CES-D mirrored that seen with the BHS (Spearman's rank correlation 0.88 and 0.68, respectively). There was no change in mean scores on the two measures with repeat testing in the 433 women who completed them and test–retest reliability was good (intraclass correlations Brief-H-Neg 0.67 and Brief-H-Pos 0.72). Conclusions These findings provide support for the validity of the Brief-H-Neg and Brief-H-Pos. These brief measures are likely to be useful in large population studies assessing hopelessness. Trial registration number NCT00058032.

Penetrance for breast cancer, ovarian cancer, or both in carriers of BRCA1/BRCA2 mutations is disproportionately high. Sex hormone dysregulation and altered end-organ hormone sensitivity might explain this organ-specific penetrance. We sought to identify differences in hormone regulation between carriers of BRCA1/2 and women who are negative for BRCA1/2 mutations.We assessed endometrial thickness for each menstrual cycle day (as an index of hormone regulation) in 393 scans from 228 women in the UK Familial Ovarian Cancer Screening Study (UK FOCSS) known to carry either mutation and 1573 scans from 754 women known to be negative for the mutations. To quantify differences in endometrial thickness we focused on days 10-14 and days 21-26, and calculated the area under the curve. We then compared serum oestradiol and progesterone titres during these days of the menstrual cycle in the same groups. Follicular and luteal oestradiol and progesterone serum titres were grouped into quartiles and odds ratios were calculated with logistic regression.Follicular phase endometrial thickness of carriers of the mutations adjusted for age and day of the menstrual cycle was higher (odds ratio [OR] 1·11, 95% CI 1·03-1·20; p=0·0063) and luteal phase endometrial thickness lower (0·90, 0·83-0·98; p=0·027) than for women negative for the mutations. Median luteal phase titres of progesterone were 121% higher (p=0·00037) in carriers than in women negative for the mutations, and for oestradiol were 33% higher (p=0·007)-ie, 59% of carriers had concentrations of serum progesterone that would have been in the top quartile of concentrations in the control group (OR 8·0, 95% CI 2·1-52·57; p=0·008).Carriers of BRCA1/BRCA2 mutations are exposed to higher titres of oestradiol and progesterone-known risk-factors for breast cancer. Higher titres of oestradiol in carriers are compatible with this hormone having a role in ovarian carcinogenesis in such women. Our findings could not be explained by differential contraceptive pill use.

Over the last two decades, survival rates from women’s cancers (breast, ovarian, endometrial and cervical cancer) have all but modestly improved despite huge efforts from both research and clinical communities. In parallel with this, the field of epigenetics has grown from its infancy into a promising scientific discipline. In particular, DNA methylation analysis has been adopted by oncologists in an attempt to better understand and manage cancer. Now that the epigenetic technological base has caught up, the potential of methylation markers in cancer research is finally being realized. In this review, we present the current status of epigenetic research into women’s cancers with a main focus on DNA methylation analysis. We provide an overview of technological development, current markers of risk prediction, early detection, diagnosis, prognosis and response to treatment, and highlight the progression of epigenetic therapies. Finally, we comment on the potential impact of epigenetic analyses on the future of women’s health.

DNA methylation of polycomb group target (PCGT) genes is an early step in carcinogenesis and could potentially be assayed to determine cancer risk prediction. To assess whether methylation changes in PCGT genes in normal tissue is able to predict the presence of cancer, we studied HOXA gene methylation in normal endometrium from premenopausal ovarian cancer patients and age-matched healthy controls without ovarian cancer. DNA methylation of HOXA9 and HOXA11 genes in normal endometrium was associated with ovarian cancer in an initial test set and this was subsequently confirmed in independent validation sample sets. The overall risk of ovarian cancer was increased 12.3-fold by high HOXA9 methylation for all stages, and 14.8-fold for early stage ovarian cancers, independent of age, phase of the menstrual cycle and histology of the cancer. The results of this proof of principle study demonstrate the potential to detect ovarian cancer via analysis of normal endometrial cells and provide insight into the possible contribution of this novel approach in ovarian cancer risk prediction and prevention.

Introduction: Identification of serum biomarkers enabling earlier diagnosis of pancreatic ductal adenocarcinoma (PDAC) could improve outcome. Aims: Serum protein profiles in patients with pre-clinical disease and at diagnosis were investigated. Patients & methods: Serum from cases up to 4 years prior to diagnosis of PDAC and controls enrolled on the UK Collaborative Trial of Ovarian Cancer Screening (UKCTOCS, n=174) were studied, alongside samples from patients diagnosed with PDAC, chronic pancreatitis, benign biliary disease and from healthy subjects (n=199). Isobaric tags for relative and absolute quantification (iTRAQ) enabled comparisons of pooled serum from a test set (n=150). Validation was undertaken using multiple reaction monitoring (MRM) and/or western blotting in all 373 human samples and in a KPC mouse model. Results: iTRAQ identified thrombospondin-1 (TSP-1) as down-regulated pre-clinically and in diagnosed patients. MRM confirmed significant down-regulation of TSP-1 up to 24 months prior to diagnosis. A combination of TSP-1 and CA19-9 (AUC= 0.86), significantly outperformed both markers alone (0.69 & 0.77 respectively). TSP-1 was also decreased in PDAC patients compared to healthy controls (P