We hypothesised that breast cancer risk for relatives of women with early-onset breast cancer could be predicted by tumour morphological features.BRCA1 promoter region in peripheral blood DNA. We calculated standardised incidence ratios (SIR) by comparing the number of mothers and sisters with breast cancer with the number expected based on Australian incidence rates specific for age and year of birth.We studied female first-degree relatives of a population-based sample of 452 index cases with a first primary invasive breast cancer diagnosed before the age of 40 years. For the index cases, a standardised tumour morphology review had been conducted for all; estrogen (ER) and progesterone receptor (PR) status was available for 401 (89%), and 77 (17%) had a high-risk mutation in a breast cancer susceptibility gene or methylation of the P <0.02). Excluding index cases with known genetic predisposition or BRCA1 promoter methylation, absence of extensive sclerosis, circumscribed growth, extensive intraductal carcinoma and lobular growth pattern were independent predictors with between a 2.0- and 3.3-fold increased risk for relatives . Relatives of the 128 (34%) index cases with none of these four features were at population risk  while relatives of the 37 (10%) index cases with two or more features were at high risk .Using Cox proportional hazards modelling, absence of extensive sclerosis, extensive intraductal carcinoma, absence of acinar and glandular growth patterns, and the presence of trabecular and lobular growth patterns were independent predictors with between a 1.8- and 3.1-fold increased risk for relatives (all This wide variation in risks for relatives based on tumour characteristics could be of clinical value, help discover new breast cancer susceptibility genes and be an advance on the current clinical practice of using ER and PR as pathology-based predictors of familial and possibly genetic risks. While information about breast tumour morphology is used to make important decisions about treatment, it can also be important in assessing breast cancer risk for an affected woman's relatives. In this paper, we look at the extent to which familial risk of breast cancer depends on tumour morphology, an issue of both clinical and aetiological significance.BRCA1 or BRCA2 [ATM [TP53 [Female relatives of women with breast cancer are at increased risk of the disease, and it is well-established that the magnitude of their risk depends upon the closeness of their relationship to their affected relatives and the number and age at diagnosis of their affected relatives -6. The ror BRCA2 -6 or a mCA2 [ATM  or TP53 TM [TP53 -10, but TM [TP53 .BRCA1 mutation carriers in our population-based sample of breast cancer cases diagnosed before the age of 40 years [BRCA1 mutation carriers [BRCA1 mutation, 16 (31%) were found to have methylation of the BRCA1 promoter region in DNA from peripheral blood [We recently showed that two breast tumour morphology features, trabecular growth pattern and high mitotic index, were sufficient to identify almost all 40 years . We alsocarriers , and of al blood .BRCA1 mutation carriers [BRCA1 promoter methylation [Given the importance of tumour morphology features to inform treatment choices and to identify carriers  or BRCA1hylation , we are This study used data and materials from the Australian Breast Cancer Family Registry (ABCFR), which includes a population-based case-control-family study that has been described in detail elsewhere ,14-16. BMissing family history data were imputed using a previously developed protocol for the following data items: date of birth, age, vital status, date of death, age at death, date at breast cancer diagnosis and age at breast cancer diagnosis ,6. VerifApproval for the study was obtained from the Human Research Ethics Committees of the University of Melbourne and the Cancer Councils of Victoria and New South Wales. All participants provided written informed consent prior to participation in the ABCFR.et al. [et al. [Archival tumour blocks were sought for all index cases under the age of 40 years at diagnosis , and a vet al. . Tumour et al.  as appli [et al. . SclerosEstrogen receptor (ER) and progesterone receptor (PR) status was obtained either from immunohistochemical testing of tumour tissue  or from From the tumour morphology review, the following features were extracted as either absent or present:Number of mitoses >50 (per 10 high powered fields)Nuclear grade - malignant Tubule formation <10 Syncytical pattern ≥75% Circumscribed Pushing margins >50% Lymphocytic Infiltrate I - diffuse Lymphocytic infiltrate II - intense Glandular growth pattern (primary growth pattern)Lobular growth pattern (primary or secondary growth pattern)Trabecular growth pattern (primary or secondary growth pattern)Tubular growth pattern (primary or secondary growth pattern)Acinar growth pattern (secondary growth pattern)Sclerosis - extensive Extensive intraductal carcinoma - present Necrosis - present Apoptosis - moderate or intense Lymphatic invasion - present Note: growth patterns were scored as glandular (primary only), lobular, trabecular, tubular and acinar (secondary only).BRCA1 and BRCA2 mutation testing have been reported elsewhere and included: sequencing, protein truncation testing, two-dimensional gel electrophoresis, screening for large genomic alterations, testing for Ashkenazi founder mutations and testing for duplication of exon 13 of BRCA1 [ATM was tested for the c.7271 T>G mutation [TP53 was performed by Sanger sequencing and specific screening for larger gene rearrangements [Testing for high-risk mutations in breast cancer predisposition genes was conducted for all index cases under the age of 40 years at diagnosis and from whom a blood sample had been obtained. Full details of the of BRCA1 . ATM wasmutation -25 and tngements .BRCA1 was conducted for all index cases under the age of 40 years at diagnosis and for whom a pathology review had been conducted [Testing for methylation of the promoter region of onducted .For the present study, mothers and sisters were included if their index case was under the age of 40 years when diagnosed with a histologically confirmed first primary invasive breast cancer and had a pathology review completed. For each relative included in the analyses, the following data were extracted: relationship to the index case; date of birth; vital status; age at interview or death; breast cancer status; and age at diagnosis, if affected.P-value of 0.05 as a threshold, and to test the null hypothesis that there is no association between a set of predictive features and risks for relatives.Australian population-based female breast cancer incidence rates, specific for age and year of birth (both in five-year groupings), were obtained for 1983 to 2001 and extrapolated to earlier years as described previously . The sta5 samples from the posterior distribution of the parameters given the data, discarded the first 2 × 104 samples as burnin, and accepted every fifth posterior sample, thus reducing autocorrelation within the chain.Multivariate linear regression models were also used for estimating and testing associations between the measured pathology features and risk for relatives. These models were fitted using Bayesian regularised regression with the Least Absolute Shrinkage and Selection Operator (LASSO) penalty function ,28. The Because many of the pathology features were highly correlated, Bayesian LASSO regression and the Cox proportional hazards model were used in tandem to select features to be included in the multivariate model.® version 7.13, MathWorks, Natick, Massachusetts, USA [P-values <0.05 were considered nominally statistically significant.Statistical analyses were performed with Stata version 11, StataCorp LP, College Station, Texas, USA  and MATLtts, USA . All staBRCA1 in peripheral blood DNA . The remaining 375 (83%) constituted the subgroup for further analyses. Table A total of 856 index cases diagnosed with a first primary invasive breast cancer before the age of 40 years was recruited, of whom 452 had a tumour morphology review using archived tissue. Of these, 77 17%) had either a high-risk mutation in a breast cancer susceptibility gene or methylation of the promoter region of 7% had eiThere were 1,041 relatives (452 mothers and 589 sisters) of the index cases, an average of 2.3 relatives per index case, with a total of 11,786 person-years of observation . Data were imputed for relatives with missing data as follows: seven dates of birth; six vital status; four dates and ages at death; and one date and age at breast cancer diagnosis.Overall, 106 breast cancers were observed for the mothers and sisters of the index cases while 36.56 were expected from population incidence rates. The ratio of observed to expected, the SIR, was 2.90 with a 95% CI from 2.40 to 3.51.P-values as a test of the difference between the SIRs for the presence and absence of each pathology feature. The test of the null hypothesis of no association with these variables and risks for relatives was rejected . After fitting the five significant morphology features, there was still significant variation in risks for relatives across the remaining 13 features .Table P <0.04).Breast cancer risks for female first-degree relatives were increased if the index cases' tumours: were circumscribed; had pushing margins >50%; had intense lymphocytic infiltrate II; had number of mitoses >50; had a trabecular growth pattern; had extensive intraductal carcinoma; had a syncytial pattern; did not have extensive sclerosis; did not have a glandular growth pattern; or were ER or PR negative . When these factors were included in the model, there was at best marginally significant evidence for associations with tubular growth pattern  and tubule formation . Neither ER status nor PR status provided additional (i.e. independent) information on breast cancer risk for relatives (both P >0.3).Table When restricted to the subgroup of 375 index cases, there were 876 relatives (375 mothers and 501 sisters), an average of 2.3 relatives per index case, with a total of 9,954 person-years of observation . Overall, 69 breast cancers were observed and 30.74 were expected based on population incidence rates; the SIR was 2.25 with a 95% CI from 1.77 to 2.84.P <0.04). Neither ER status nor PR status provided independent information on breast cancer risk for relatives (both P >0.4). The test of the null hypothesis of no association with these variables and risks for relatives was rejected . Even after fitting the four significant morphology features there was still significant variation in risks for relatives across the remaining 13 features .Table P-values for the multivariate model. When all features were considered together, the pathology features identified by the Bayesian LASSO regression and the Cox proportional hazards models were: being circumscribed; having extensive intraductal carcinoma; having lobular growth pattern; and not having extensive sclerosis . When these factors were included in the model, no other factors were even marginally associated.Table Figure Figure BRCA1 promoter region methylation in peripheral blood DNA, features of their tumour morphology predict wide variation in the risk of breast cancer for their first-degree female relatives, far more than could be attributed to chance. For example, female relatives of the approximately one-third of early-onset breast cancer cases who had tumours that lacked any morphological feature associated with familial risk were at no increased risk of breast cancer. Female relatives of the approximately one-sixth of early-onset breast cancer cases who had tumours that had two or more of the four features associated with familial risk were at a substantially increased risk of breast cancer. The latter finding suggests there are undiscovered strong familial risk factors for early-onset breast cancer. The former suggests there is a substantial proportion of early-onset breast cancer that does not have a familial, let alone genetic, cause.For women with early-onset breast cancer who neither carry a mutation in a known breast cancer susceptibility gene nor have BRCA1 mutation [BRCA1 in peripheral blood DNA [BRCA1 promoter region.This work has built on previous work by us and others that identified specific morphological features which predict cases with a germline mutation ,31,32. Wlood DNA . Here, wlood DNA . We alsoWhen first considering all index cases, we found that absence of extensive sclerosis, extensive intraductal carcinoma, and trabecular and lobular growth patterns and absence of acinar and glandular growth patterns were all independent predictors of breast cancer for their first-degree female relatives. When considered together, for each of these six features their presence was associated with an approximate doubling of risk for relatives. This means there is a substantial gradient of risk for relatives across a number of these features.BRCA1 mutation status, as well as of methylation of the BRCA1 promoter region [BRCA1 promoter region, absence of extensive sclerosis, circumscribed growth, extensive intraductal carcinoma and lobular growth pattern were each independent predictors of breast cancer risk for the first-degree female relatives of the remaining index cases. Note from Figure BRCA1 germline mutations or methylation of the BRCA1 promoter region), that are associated with increased risks of breast cancer for first-degree relatives.We have previously shown that the presence of trabecular growth pattern was the major predictor of r region ,13. WhenThere are several strengths of our study. First, it is population-based, so inference can readily be made at least to the populations from which these index cases were sampled, if not more generally. Second, because the index cases were not selected on the basis of family history, this study gives insights across the whole spectrum of index cases. Third, we have used data from multiple pathologists conducting a standardised and validated tumour review and any systematic variation across the pathologists would mean that our study has under-estimated the predictive value of these morphology features.BRCA1 and BRCA2 have not yet been identified, although this is likely to be, at most, a few percent [The study also has some weaknesses. It is possible that, despite extensive testing, some index cases carrying mutations in  percent ,6. ThereBRCA1 mutation carriers [While there are several studies of breast cancer in relatives as a function of the ER and PR tumour status of an affected woman , few hacarriers  or, as icarriers ).BRAC1 mutation status has been established for some time [BRCA1 mutation carriers than family history and/or ER and PR tumour status [Being able to quantify a woman's risk of breast cancer is important for genetic counselling, prevention and screening, as well as for aetiological research. Currently, risk estimates associated with having a family history are based on the age at diagnosis of cancer, not on the cancer's morphological features. We have shown here that, at least for first-degree relatives of women with early-onset breast cancer, this is likely leading to a misassignment of familial risks for about one-half of index cases, being too high for one-third and too low for one-sixth, by factors of two-fold or more. Assignment of women with a family history of breast cancer to different screening strategies based on their absolute risk could be substantially improved were the morphology features of the tumours of the relatives taken into account. The role of morphological features to predict ome time . For earr status . Now we It is highly relevant to genetic research and counselling that one-third of early-onset breast cancers might not even have a heritable genetic basis, as evidenced by our observation that, for the group of index cases with none of the four predictive features, their relatives were not at increased risk. Most linkage studies and the recent genome-wide association studies have generally pooled all cases when trying to find predisposing genes. It is also of relevance to know if the type of breast cancers, as defined by the morphology features identified here, 'runs in the family', as this would greatly assist efforts at susceptibility gene discovery.BRCA1 mutations and TP53 mutations have characteristic morphology and acquired genetic and epigenetic mutation profiles [For the affected women, their germline status, as well as changes (both somatic genetic and epigenetic) to tumour DNA, might influence gene expression that manifests in nuances of cell growth and neoplastic tissue organisation that are able to be observed by microscope. For example, breast tumours arising in women with germline profiles -39. For On their own, the ER and PR status of the index cases were both associated with risk for relatives overall, with about a 50% increased risk if the tumour of the index case was negative for either of these immunohistochemical markers. After the exclusions, the associations were of the same magnitude and direction but no longer statistically significant. However, neither of these immunohistochemical markers were independent predictors of risks for relatives once the more strongly predictive morphological features were taken into account. That is, we present here an advance on using ER and PR alone to assess the role of pathology in predicting increased familial and possibly genetic risks.We have shown that relatively standard tumour morphology can help unravel the complex heterogeneity of breast cancers from the perspective of familial and genetic risk, at least in the setting of early-onset disease. There is a very wide variation in risks for relatives based on tumour characteristics and this could not only be of clinical value but might help discover new breast cancer susceptibility genes.ABCFR: Australian Breast Cancer Family Registry; CI: confidence interval; ER: estrogen receptor; HR: hazard ratio; LASSO: least absolute shrinkage and selection operator; PR: progesterone receptor; SIR: standardised incidence ratioThe authors declare that they have no competing interests.GSD led the writing of the manuscript, reviewed the literature, analysed and interpreted data, and also contributed to the study design and data collection. MCS established the ABCFR and its data collection, conceived the study, contributed to the collection of the data (was responsible for the collection of the pathology review data) and contributed to the data analysis and writing of the manuscript. JLH established the ABCFR and its data collection, conceived the study design, data analysis and interpretation and the writing of the manuscript. EM and DFS conducted the LASSO statistical analyses and contributed to the writing of the manuscript. GGG helped establish the ABCFR and its data collection and contributed to the writing of the manuscript. All authors read and approved the final manuscript.


Breast cancer incidence differs by ethnicity in New Zealand (NZ) with Māori (the indigenous people) women having the highest rates followed by Pakeha (people primarily of British/European descent), Pacific and Asian women, who experience the lowest rates. The reasons for these differences are unclear. Breast density, an important risk factor for breast cancer, has not previously been studied here. We used an automated system, Volpara™, to measure breast density volume from the medio-lateral oblique view of digital mammograms, by age (≤50 years and >50 years) and ethnicity (Pakeha/Māori/Pacific/Asian) using routine data from the national screening programme: age; x-ray system and mammography details for 3,091 Pakeha, 716 Māori, 170 Pacific and 662 Asian  women. Linear regression of the natural logarithm of absolute and percent density values was used, back-transformed and expressed as the ratio of the geometric means. Covariates were age, x-ray system and, for absolute density, the natural log of the volume of non-dense tissue (a proxy for body mass index). Median age for Pakeha women was 55 years; Māori 53 years; and Pacific and Asian women, 52 years. Compared to Pakeha women (reference), Māori had higher absolute volumetric density  which remained following adjustment  and was stronger for older compared to younger Māori women. Asian women had the greatest risk of high percentage breast density  while Pacific women in both the ≤50 and >50 year age groups  had the lowest percentage breast density compared to Pakeha. As well as expected age differences, we found differential patterns of breast density by ethnicity consistent with ethnic differences seen in breast cancer risk. Breast density may be a contributing factor to NZ’s well-known, but poorly explained, inequalities in breast cancer incidence. Breast cancer remains the most important cancer for New Zealand women, who experience high breast cancer incidence and mortality rates compared to many other developed countries Breast density, a measure of the relative amounts of dense and fatty breast tissue, is an important risk factor for breast cancer; women with dense breasts have been found to be four to six times more likely to develop breast cancer than those with fatty breasts Quantitative methods to measure the area of the mammogram that is dense include visual assessment and computer-assisted thresholding methods A national breast screening service has been offered in New Zealand since 1999. Currently, digital mammography is installed in four of the eight centres of the service, Breast Screen Aotearoa (BSA) No studies of breast density have previously been conducted in New Zealand, thus, there is an opportunity to obtain some baseline information about this important risk factor within the context of an ethnically diverse population. Since mammographic density distributions both contribute to breast cancer risk profiles The study was approved by the Northern X Regional Ethics Committee (ref: NTX/11/EXP/112) of New Zealand. All analyses were carried out on routinely collected data, made available to the researchers in a de-identified form, which had already been obtained at the time of the mammogram by BSWN. The Northern X Regional Ethics Committee therefore accepted that written consent was not required by individual participants and granted approval for the study.The aim of the study was to describe the distribution of breast density in a population of New Zealand women. New Zealand has a population of approximately 4.4 million The participants were all women who underwent routine mammography screening during the period 3/12/2010–30/6/2011 at the Waitakere and Takapuna fixed sites and the mobile units operating in Northland as part of the BSWN programme. BSWN has an eligible screening population of approximately 100,000 women and screens around 35,000 women per year Only data routinely collected by the BSA programme was available for the study which included: age at examination; self-identifed ethnicity; x-ray system (GE or Siemens) and mammography screen details using the medio-lateral oblique (MLO) view (left breast & right breast). The volume of the mammogram that appeared dense was estimated using the Volpara™ system version 1,4,1. This is a computer algorithm which models the x-ray physics in order to produce estimates of the total breast volume, the absolute dense volume, the volume of non-dense tissue and the percentage of the total breast volume that is dense (density percentage). Volpara™ works by using the physics parameters to estimate the x-ray attenuation between any pixel and the x-ray source. From that attenuation, it is possible to estimate the amounts of fat and fibroglandular (dense) tissue present Both GE and Siemens x-ray systems are in use in BSWN. Initial work indicated that the two x-ray systems gave slightly different results of Volpara™ density for the same women. Thus, a standardisation procedure was used for the Siemens data, using results from Volpara™ system version 1,4,2, looking at how the results changed between versions 1,4,1 and 1,4,2 and applying a correction factor to the absolute dense volume and the total breast volume estimates. Breast thickness was determined from the distance between the compression plates during mammography.Absolute dense volume was divided into quintiles for analysis. Percent volumetric density was analysed in pre-defined categories 0 to 4.5%, ≥4.5 to 7.5%, ≥7.5 to 15.5%, and ≥15.5%, which map to the BI-RADS breast composition categories based on their 4 groupings of: 1-fatty (VDG 1); 2-scattered (VDG 2); 3-heterogenous (VDG 3); and 4-extremely dense (VDG 4) The MLO views were selected for measurement using the combined average for the left and right breast screen image estimates. Descriptive analyses used medians and inter-quartile ranges because of the positively skewed nature of the outcome variables (data available on request). For multivariable analyses, the natural logarithm of the absolute and percentage density values were used. Linear regression of these logged variables was used, and the output was back-transformed and expressed as the ratio of the geometric means. Covariates included in the models were age (as a continuous variable), x-ray system and for absolute density, the natural log of the volume of non- dense tissue.Statistics were performed using STATA version 11.2.Mammographic density measures were available for a total of 4,300 women screened during the pilot study period. These included assessment mammograms for 61 of the women which were subsequently excluded from the final analyses, leaving a total of 4,239. Of these, 3,091 (73.0%) were Pakeha, 716 (7.4%) were Māori, 170 (4.0%) were Pacific and 662 (15.6%) were Asian women.The median age was highest in Pakeha women at 55 years while Asian women had the lowest median age at 52 years . The datFurther exploration showed ethnic differences in the distribution of absolute mammographic density both overall and by age categories . OverallDifferent patterns by ethnicity were also observed in the density percentage categories  with theThe crude associations between ethnicity and absolute volumetric breast density are further shown in A different pattern of associations was seen with volumetric percent density, shown in These findings have highlighted differential patterns of breast density between ethnic groups in New Zealand. In particular, compared to Pakeha women, Māori women had higher levels of absolute volumetric density and Asian women had lower levels. For percentage breast density, Asian women had higher and Pacific women had lower breast density.The main limitation of the study is our lack of additional data in order to assess the potential effect of confounding on our results. While imperfect, the estimate of non-dense volume as a proxy for BMI is not unreasonable since body weight is associated with the volume of non–dense tissue in the mammogram Our finding of a higher absolute dense volume in Māori compared to Pakeha women remained following adjustment for age, x-ray system and non-dense volume. To date, international literature on differences in breast density by ethnicity is scarce and mixed. Most of the previous work documenting ethnic differences in mammographic density has been conducted in the US With regard to percent density, we found a high percent density in Asian and low percent density in Pacific women which is again, most probably due to body fat, although of note, for Māori women, who have an on average higher BMI than Pakeha women A further key area of future exploration will be whether the demonstrated differences in breast density can explain ethnic differences in breast cancer risk in New Zealand. Although we were unable to explore this in the current study, our results are generally consistent with documented differences in breast cancer incidence rates seen in this country In summary, in these novel analyses of breast density distribution in New Zealand women, we found not only expected age differences in breast density but also differences by ethnicity, with Māori being more likely and Asian women less likely, to have high volumes of dense tissue than Pakeha women. Further work in this area is warranted, in particular, examining the role of breast density and its potential contribution to New Zealand’s well-known, but poorly explained ethnic disparities in breast cancer incidence and outcomes.


The relationship between physical activity (PA) throughout life and the risk of postmenopausal breast cancer overall and by estrogen receptor (ER) and progesterone receptor (PR) status, has been reported, but without consistent results. The present study aimed to investigate PA from young age to adulthood in participants of the Norwegian Women and Cancer (NOWAC) Study, in order to determine whether changes in PA level affect the risk of postmenopausal breast cancer.1767 invasive breast cancer cases were identified among 80,202 postmenopausal participants of the NOWAC Study during 8.2 years of median follow-up. PA levels at age 14 years, 30 years and at cohort enrollment were obtained via a self-administered questionnaire. Multivariate Cox proportional hazard regression models were used to estimate relative risks and 95% confidence intervals of the risk of postmenopausal breast cancer overall and by ER/PR status.P for trend = 0.04) compared to women with a moderate physical activity level at age 30. Women with a low physical activity level at all three periods of life had a 20% significantly reduced risk of postmenopausal breast cancer, as well as a reduced risk of ER+/PR + and ER+/PR- breast tumors, compared with women who maintained a moderate physical activity level. However, when analyses were corrected for multiple tests, the result was no longer statistically significant. The findings were consistent over strata of age, body mass index and use of hormone replacement therapy.Risk of postmenopausal breast cancer overall and by ER/PR status was not associated with physical activity level at enrollment. Women with a low PA level at age 30 had an increased risk of ER+/PR + breast tumors (The study results from this large Norwegian cohort do not support an association between physical activity at different periods of life and the risk of postmenopausal breast cancer. Breast cancer is the most frequent cancer type among women worldwide in terms of both incidence and mortality , and comIn 2005, Margolis et al. analyzed the relationship between risk of premenopausal breast cancer and PA at different periods of life in the Women’s Lifestyle and Health Study. The Women’s Lifestyle and Health Study is a prospective study of women in Norway and Sweden , and incA total of 80,202 postmenopausal women from the NOWAC study were followed for an average of 8.2 years, constituting 648,731 person-years. Mean age at enrollment for the study sample was 48 years. During follow-up, 1,767 women were diagnosed with breast cancer (mean age at diagnosis: 59.7 years). Information on ER status was available for 79% of breast cancer cases; of these 83% were ER + and 17% were ER-. PR status was available for 79% of breast cancer cases, of which 63% were PR + and 37% were PR-. Seventy-four percent of the women reported moderate, high or very high PA levels at enrollment. This proportion was even higher for the same PA levels at age 14 years (85.7%) and age 30 years (86.0%) . The same comparison using women with a very low PA level at age 14 rendered an RR of 0.77 .In adjusted multivariable models, there was no association between PA level at enrollment or at age 30 and risk of postmenopausal breast cancer (Table P for trend = 0.04) of increased risk of ER+/PR + breast tumors was observed among women with a very low PA level compared to those with a moderate PA level (Table P for trend = 0.02) and ER+/PR- (P for trend = 0.02) breast tumors. Analysis of women with unknown ER/PR status showed no significant associations with PA level (data not shown). We found no indication that increasing PA beyond the moderate PA level was inversely associated with ER/PR status.No statistically significant associations were observed between PA level at enrollment and ER/PR status of breast tumors Table . At age A significant increased risk of postmenopausal breast cancer  was found among women who changed their PA level from active to inactive between age 14 and age 30. Women who changed their PA level in the opposite direction; i.e. from inactive to active, had a significant reduced risk of postmenopausal breast cancer . In contrast, women who were inactive at age 14 and age 30 had a significantly reduced risk of postmenopausal breast cancer . Women who were inactive from age 14 to enrollment had a non-significant reduced risk , compared to women who were active at all three periods of life . All multivariate models were adjusted for BMI, but results were almost identical to those from models without adjustment for BMI. Homogeneity tests confirmed that there were no statistical differences between the magnitudes of the effect estimates. We assessed the potential effect modification of BMI and hormone replacement therapy use by conducting stratified analyses. These analyses revealed no statistically significant interaction of BMI or hormone replacement therapy use on the relationship between PA level at enrollment or changes in PA level and risk of postmenopausal breast cancer overall, or by ER/PR status (data not shown). The analyses stratified by different periods of enrollment did not change the estimates (data not shown). Finally, the Holms procedure for multiple testing showed that no estimates reached statistical significance, neither for different periods of life, or for different ER/PR status of breast cancer tumors (data not shown).In this Norwegian cohort study, there was no consistent, significant relationship between PA level and postmenopausal breast cancer overall, or by ER/PR status. We found an increased risk of postmenopausal breast cancer overall, and of ER+/PR + breast tumors among women who went from active to inactive between age 14 and age 30. Furthermore, among women who changed from inactive to active at the same ages, the opposite was true; they had a decreased risk of postmenopausal breast cancer overall, and of ER+/PR + breast tumors compared to women who remained physically active throughout all three periods of life. Furthermore, we observed that women with a very low PA level at age 30 had an increased risk of ER+/PR + breast tumors compared to women with a moderate PA level at the same age.We also observed a statistically significant 20% reduction in the risk of postmenopausal breast cancer among women with PA levels that were low at age 14, and remained low throughout adulthood, as compared to women who were either moderately active at age 14, or had a moderately active lifestyle throughout adulthood . This unexpected association was confirmed in the analysis of ER/PR status, which revealed a decreased risk of ER+/PR + and ER+/PR- breast tumors, though this was not the case for any of the other combinations of ER/PR status. Taken together these findings are inconsistent, and the risk of false-positive findings only strengthens the uncertainty. Analysis for multiple testing confirmed that none of the estimates reached statistical significance, indicating that the inverse relationship between PA level and postmenopausal breast cancer was not strong in our study, and this must be considered when reviewing the results.The strengths of our study include the prospective cohort study design, long follow-up period and the completeness of follow-up data, which minimizes the potential for selection bias. The ascertainment of postmenopausal breast cancer and of the ER/PR status of breast tumors was performed through the linkage to the national cancer registry, which is virtually complete. Our study sample was taken from a source population that is considered representative of Norwegian general female population 34 to 70 years of age . The avaOur study also has some limitations. The lack of a clear risk pattern may be partly due to the fact that PA level was self-reported, and collected all at once for three different periods of life. Therefore this information was based on the participant’s capability to recall PA levels correctly; incorrect recall could have led to misclassification that might have influenced the true exposure status early in life. Moreover, the changes in PA level at age 14, age 30 and at enrollment may not be entirely representative of the 15–20 years in between. Indeed, elderly women have been shown to overestimate PA levels earlier in life ,28. ThisER and PR status was missing for 21% of the women in the present analysis. Therefore analyses on ER/PR status of breast tumors had less power than analyses on postmenopausal breast cancer overall. Our choice of age 53 as a cut-off for age at menopause ended up excluding women that were likely menopausal. Also, we used PA levels reported at enrollment for all three time periods for women who became postmenopausal during follow-up (and thus were included only from that point in time). Finally, lack of controlling for unknown factors, and the fact that information on potential confounding factors was only collected at enrollment, may have introduced residual confounding.The original aim of the NOWAC study was to investigate the association between breast cancer and oral contraceptive use, with no specific emphasis on PA other than as a possible confounding factor. Although the correlation between our PA measure and the reference method  was mode2, premenopausal women, and for ER-/PR- breast tumors [Our results did not corroborate findings from other studies that suggested PA protects against postmenopausal breast cancer ,18,33-38t tumors . Howevert tumors ,6,40-45.t tumors . Others t tumors . Converst tumors .In principle, our findings indicate the same trend, suggesting that low PA level at age 14 has a modest protective effect on the risk of postmenopausal breast cancer overall, and on ER+/PR + breast tumors. These results are in contrast with those of The Women’s Health Initiative, which investigated PA at age 18 years and risk of postmenopausal breast cancer and found no association either overall or by hormone receptor status. However, they found an inverse effect of PA at age 35 , whereasThe inverse relationship of PA on hormone receptor-positive breast tumors has been reported ,38,53,54Some studies have confirmed that the association between PA and breast cancer is not modified by BMI ,47,62-64Our study did not detect a consistent inverse effect of PA on the risk of postmenopausal breast cancer in this large cohort of Norwegian women. For future research, information on PA during follow-up is essential. Despite the results of the present study, PA remains an important modifiable lifestyle factor with the potential to reduce the risk of postmenopausal breast cancer.The NOWAC Study has been previously described in detail ,31. In sA total of 122,857 women who participated in the NOWAC Study were initially eligible for the present analysis. We excluded all women with prevalent cancer at enrollment (n = 4620), those who died within the first year of follow-up (n = 265) and those with missing information on PA level at enrollment . Women who reported their age at menopause at the time of enrollment or during follow-up (available from second questionnaire for some of the women) were categorized as postmenopausal. All other women, including those with missing information on age at menopause, were categorized as postmenopausal once they reached 53 years of age during follow-up, i.e., 53 years of age was used as a proxy for age at menopause. This cut-off was based on the definition used in the Million Women Study, and later in the NOWAC study -68. Base“By physical activity we mean activity both at work and outside work, at home, as well as training/exercise and other physical activity, such as walking, etc. Please mark the number that best describes your level of physical activity; 1 being very low and 10 being very high”. The PA scale used for this study has recently been validated for the assessment at enrollment [P <0.001) Spearman’s rank correlation coefficients were found (range: 0.36-0.46) between the PA level at enrollment and concurrent outcomes from criterion measures of a combined sensor monitoring heart rate and movement. The scale ranged from 1 (very low) to 10 (very high), and corresponded to mean values of 0.8 and 3.4 hours/day of moderate/vigorous PA, respectively, with a linear increase (P for trend <0.001), and appeared valid to rank PA level in a Norwegian population of women [PA level at age 14, age 30 and at enrollment (ages 34–70) was assessed by self-report on an ordinal scale of 1 to 10. PA was defined in the questionnaire as follows: rollment , and refof women . The PA of women ,69. We fInformation collected at enrollment included age, duration of education, weight, height, BMI, smoking history  and age at smoking initiation, alcohol consumption (grams/day), age at menarche, use of oral contraceptives, age at first birth, parity, age at menopause and use of hormone replacement therapy. Self-reported illnesses included cardiovascular diseases  and diabetes mellitus. Information on history of breast cancer in the participant’s mother (yes/no) was also collected. Information on prevalent cancer was obtained through linkage to the Cancer Registry of Norway.Person-years were calculated from start of follow-up for women who were postmenopausal at study enrollment, and from age at menopause (either reported or 53 years) for all other women, until date of diagnosis, emigration, death, or end of the study period (December 31st 2009), whichever occurred first. We obtained information on date of death or emigration from the Norwegian National Population Register, and on cancer diagnosis and ER and PR status of breast tumors through linkage to the Cancer Registry of Norway. Cancer diagnoses were coded according to the 10th revision of the International Statistical Classification of Diseases, Injuries and Causes of Death. The main endpoint in this study was incidence of invasive breast cancer (C50), and ER and PR status of breast tumors was classified as follows: ER+/PR+, ER+/PR-, ER-/PR+, ER-/PR- and unknown.Characteristics of the study participants were examined by PA level and breast cancer incidence. Cox proportional hazard regression models were used to estimate the hazard ratio as a measure of relative risk (RR) with corresponding 95% confidence intervals (CI). In specific analyses of ER and PR status, breast tumors without a specific ER or PR status were considered censored observations. There were too few ER-/PR + breast tumors to allow for a meaningful analysis, therefore no corresponding results are presented. Tests for trend were estimated using the collapsed five-level scale and entered as a continuous term in the Cox proportional hazards regression models.2), smoking status , smoking habits, which included smoking duration (years), quantity  and age at smoking initiation combined as one variable , alcohol consumption , age at menarche (years), use of oral contraceptives (ever/never), age at first birth , parity , use of hormone replacement therapy (current/never), self-reported cardiovascular disease (yes/no) and diabetes mellitus (yes/no), and history of breast cancer in the participant’s mother (yes/no).The proportional hazard assumption was checked using Schoenfeld residuals and Kaplan-Meier log (-log) survival plots, which suggested no evidence of deviation from proportionality. In the multivariable Cox regression models, confounders adjusted for in the different models were height (cm), BMI (kg/m2), use of hormone replacement therapy (ever/never) and age  for change in PA between age 30 and enrollment. Use of hormone replacement therapy may modify the association between BMI and risk of postmenopausal breast cancer [Duration of education and total energy intake were not included in the final models, as they were not appreciably related to breast cancer after adjustment for the other confounders in the NOWAC cohort  or the pt cancer ,72, and Homogeneity of the effect estimates in the different models was evaluated using the Wald test to compare different regression estimates. Information regarding BMI at 18 years of age was available for 95% of the participants, thus we carried out a sensitivity analysis; BMI at age 18 has no effect on out risk estimates, therefore this variable was not included in the main models. Finally, we conducted Holm’s multiple-test procedure to protect against false-positive conclusions when analyzing subgroups of breast cancer. Analyses were conducted using STATA version 12.0, special edition , with all statistical tests two-sided and conducted at the 0.05 significance level.The authors declare that they have no competing interests.KBB carried out and interpreted the statistical analysis and drafted the manuscript. TB contributed to the statistical analysis and interpretation of the data and critical revision of the manuscript. EL is the principle investigator and designed the NOWAC Study and contributed with critical revision of the manuscript. EW contributed with the design of the present study and critical revision of the manuscript. All authors read and approved the final manuscript.


In vitro studies have demonstrated the role of the BCL-2 family of genes in endometrial carcinogenesis. The role of genetic variants in BCL-2 genes and their interactions with non-genetic factors in the development of endometrial cancer has not been investigated in epidemiological studies.BCL2, BAX, BAD and BAK1 genes.We examined the relationship between BCL-2 gene family variants and endometrial cancer risk among 1,028 patients and 1,922 age-matched community controls from Shanghai, China. We also investigated possible interactions between genetic variants and established risk factors . Individuals were genotyped for 86 tagging single nucleotide polymorphisms (SNPs) in the BCL2 gene . For SNPs rs17759659 and rs7243091 , the associations were independent. The odds ratio was 1.27 (95% CI: 1.04–1.53) for women with AG genotype for the SNP rs17759659 and 1.82 (95% CI: 1.21–2.73) for women with the GG genotype for the SNP rs7243091. No interaction between these two SNPs and established non-genetic risk factors of endometrial cancer was noticed.Significant associations with endometrial cancer risk were found for 9 SNPs in the BCL2 gene may be associated with the risk of endometrial cancer in Chinese women.Genetic polymorphisms in the  BCL2, BCL-Xl) or pro-apoptotic  proteins BCL2 expression in the normal endometrium during the menstrual cycle BCL2 gene may be the most hormone-dependent member of the BCL-2 family of genes Apoptosis is a selective process for deleting cells, which is an essential physiological process required for tissue size regulation and morphogenesis BAX, BAK and BAD genes are believed to oppose cell carcinogenesis, while the BCL2 gene can promote cancer cell growth by blocking apoptosis BCL2 gene has been observed in various human cancer tissues, including breast, colon, thyroid and endometrial carcinomas in vitro studies of human tissue samples BCL2 slows down cell growth and very high expression can promote cell death, while lower BCL2 expression can be a sign of inhibition of apoptosis in human breast and endometrial carcinoma tissues BCL2 expression differs by degree of tumor aggressiveness and differentiation BCL2 expression has been shown to be very low or absent in higher grade carcinomas compared with lower grade carcinomas BCL2 expression may be suppressed during cancer progression BCL2 expression could be a valuable predictor of cancer progression and prognosis BCL-2-family proteins play a crucial role in carcinogenesis. The pro-apoptotic BCL2 gene, with susceptibility to myeloid leukemia, squamous cell carcinoma of the head and neck, esophageal cancer and prostate cancer BAD, BAX, BCL2 or BAK1 are associated with endometrial cancer risk. Furthermore, we examined whether variants in these genes modify the effect of established non-genetic risk factors for endometrial cancer by using data from the Shanghai Endometrial Cancer Genetics Study (SECGS).Only a few observational studies have investigated the relation of genetic variants in the BCL-2-gene family, particularly the The study protocols were approved by the Institutional Review Boards of participating institutes, i.e., the Vanderbilt University School of Medicine, Vanderbilt University, Nashville, Tennessee, United States; and the Shanghai Cancer Institute, Shanghai, China; Shanghai, China. All participants provided written, informed consent.The SECGS uses resources from two studies and includes subjects who participated in the Shanghai Endometrial Cancer Study (SECS) and control subjects who participated in the Shanghai Breast Cancer Study . Both the SECS and SBCS are population-based, case-control studies conducted in Shanghai, China between 1996 and 2003, which used nearly identical study protocols. Details of the study designs for these two studies have been described in detail elsewhere 2) and waist-to-hip circumference ratio (WHR) were calculated by using measured anthropometrics as described previously Study participants were interviewed to obtain detailed information on demographics, lifestyle habits, dietary intakes and supplement use, menstrual and reproductive history, hormone use, disease history, weight history and family history of any cancer. Anthropometric measurements, including weight, height and circumferences of the waist and hips, were taken by the interviewers. Menopause was defined as the cessation of the menstrual period for at least 12 months before the reference date (diagnosis date for cases and interview date for controls), excluding lapses caused by pregnancy, breastfeeding or estrogen hormone use. Body mass index  by using the Tagger program 2r≥0.9. SNPs with a known or potential function were all included. Genotyping was conducted at the Vanderbilt Microarray Shared Resource. As a quality control (QC) procedure, we included 39 blinded duplicate samples and 12 HapMap DNA samples in the genotyping. The average consistency rate for these samples was 99.6%. The laboratory staff members were blinded to the case-control status and identity of all samples. A total of 86 SNPs in BCl-2-family genes  were included in the study, with an average call rate of 99.8%.Haplotype-tagging SNPs (tagSNPs) in BCL-2-family genes were selected from the Han Chinese data of the International HapMap Project  were based on control data. LD between polymorphisms in the BCL2, BAD, BAX, and BAK1 genes was assessed using HaploView, version 4.2 software AA) for the major allele, heterozygous (Aa) and homozygous (aa) for the minor allele] under additive, dominant and recessive genetic models. Age (continuous) and education  were adjusted for in all analyses. Additional adjustment for menopausal status, family history of endometrial cancer in first-degree relatives and BMI did not alter the gene-disease associations.We used SAS software  for the statistical analyses. Demographic, lifestyle and clinical factors were compared between cases and controls by using the χWe derived risk scores for established risk factors for endometrial cancer based on previous literature P value<0.05 was considered statistically significant. P-values presented in this paper were not corrected for multiple tests. None of the associations for the 86 SNPs tested would reach statistical significance (5.8*10−4), if the Bonferroni correction were applied.In addition, we examined the effect of two independent SNPs on associations between scored established risk factors and endometrial cancer in the stratified analysis by genotype. Tests for trend were performed by entering the categorical variables as continuous parameters in the models. Multiplicative interactions between scored risk factors and genotype subgroups were assessed by comparing the difference of the log likelihoods between models with the main effects and models with both the main effects and the interaction terms. We used the Hosmer and Lemeshow goodness-of-fit test to check the logistic regression models. All statistical tests were two-tailed, and a vs. 50.8 P<0.01). Associations for most factors presented in the P trend <0.01) compared with women who were in the lowest tertile. Risk estimates increased 3.76-fold among women with a risk score ≥7 (median range) compared with women whose risk score was <7.Associations of non-genetic risk scores with endometrial cancer risk are presented in BCL2 gene  had a statistically significant association with endometrial cancer risk  between seven of these SNPs (data not shown). SNP rs17759659 was in LD (r2 of 0.825) with SNPs rs10460159, rs11663788 and rs6810, which are in predicted miRNA binding sites. Overall, only 2 SNPs, rs17759659 and rs7243091, (r2<0.3) were found to be independent of each other and were both associated with risk of endometrial cancer, even after adjusting for each other in the model. The minor alleles (G) of both rs17759659 and rs7243091 were associated with increased risk of endometrial cancer . None of the SNPs in the BAD, BAX or BAK1 genes were associated with risk.Nine of the 72 SNPs in the  P<0.05, Table 3,Table 4). We found no evidence that these two SNPs modify the association of non-genetic risk factors with endometrial cancer (P for interaction  = 0.87 for rs7243091 and 0.62 for rs17759659).We further evaluated whether SNPs rs7243091 or rs17759659 have a modifying effect on associations between the score of established risk factors and endometrial cancer risk  were found to be independent of each other and were both associated with risk of endometrial cancer. The 9 disease-associated SNPs in the intronic region of BCL2 do not appear to alter amino acids, so their relationship to the underlying biology of endometrial cancer remains unclear. We also found that a risk score created based on established endometrial cancer-risk factors To the best of our knowledge, this is the first study to evaluate genetic variants in the BCL-2 family of genes together with the quantified effect of established risk factors on endometrial cancer risk. Common genetic variants in the BCL-2-family genes BCL2 gene and endometrial cancer risk are consistent with findings from previous human tumor tissue studies, which have clearly demonstrated a role for members of the BCL-2 family of genes, particularly the BCL2 and BAX genes, in endometrial carcinoma BCL2 in endometrial carcinoma BCL2 gene expression is related to the degree of aggressiveness and differentiation in endometrial carcinoma et al. observed low levels of BCL2 expression in grade I endometrial carcinomas and very low levels or no BCL2 expression in grade II and III endometrial carcinomas, respectively BCL2 g ene is a valuable predictor of disease progression Despite the limited observational data on the relationship between BCL-2-gene family polymorphisms and endometrial cancer, our findings on the associations between genetic variants in the BCL2 may be the most hormone-dependent member of the BCL-2 family of genes and that BCL2 expression patterns in the normal endometrium may vary depending on the menstrual cycle phase or hormonal environment BCL2 expression decreased or disappeared after administration of levonogestrel, a synthetic progestogen used in some hormonal contraceptives BCL2 was over expressed in the anti-progestin-treated endometrium BCL2 expression in the human endometrium. However, in our study, we found no modifiable effect of two independent SNPs within the BCL2 gene on associations between non-genetic factors, including hormonal factors, and endometrial cancer risk. These results may be due to the size of our sample not being large enough to detect such interactions or the studied SNPs may not be causally linked to BCL2 gene function. Further studies with a larger sample size, comprehensive evaluation of the genetic variants and, preferably, a direct measurement of estrogen levels are needed to understand the role of BCL2 in endometrial carcinogenesis.It has also been suggested that −4), if the Bonferroni correction were applied, probably due to the small sample size. Second, our study is the first systematic study of genetic associations between BCL-2-family genes and endometrial cancer risk, and the findings need to be replicated in an independent cohort of endometrial cancer patients. Last, the lack of a direct measurement of estrogen hormone levels prevented us from investigating the potential role of BCL-2-family genes in the context of hormone exposure.Our study has several strengths, including use of a population-based sample, a relatively large sample size, a genetically homogeneous population, high response rates at recruitment (82.8% for cases and 74.4% [the SECS] and 90.3% [the SBCS] for controls), and histopathology-confirmed case status, all of which help to limit selection and misclassification biases. Detailed information on reproductive and lifestyle factors, medical history, and measured anthropometrics were collected by trained interviewers. No epidemiologic study has yet simultaneously evaluated a large number of polymorphisms in several apoptosis-related genes in the BCL-2 family with endometrial cancer risk. In addition, we also evaluated the potential interactions between established risk factors and genetic markers. Limitations of this study should also be considered when interpreting the results. First, this study evaluated 86 SNPs, and none of the associations would reach statistical significance (5.8*10BCL2 gene were associated with endometrial cancer risk among Chinese women. However, we found little evidence that these polymorphisms modify the association of established risk factors with endometrial cancer. Further studies are needed to confirm our findings and to elucidate the role of gene-environment interactions in endometrial cancer development.In summary, we found that two independent SNPs in the


CYP11A1 gene encodes the key enzyme that catalyzes the initial and rate-limiting step in steroid hormone synthesis. In this study, the associations between single nucleotide polymorphisms (SNPs) in CYP11A1 and breast cancer susceptibility were examined. Six SNPs in CYP11A1 were genotyped using the MassARRAY IPLEX platform in 530 breast cancer patients and 546 healthy controls. Association analyses based on a χ2 test and binary logistic regression were performed to determine the odds ratio (OR) and 95% confidence interval (95% CI) for each SNP. Two loci (rs2959008 and rs2279357) showed evidence of associations with breast cancer risk. The variant genotype C/T-C/C of rs2959008 was significantly associated with a decreased risk  compared with the wild-type TT. However, the homozygous TT variant of rs2279357 exhibited increased susceptibility to breast cancer . The locus rs2959003 also showed an appreciable effect, but no associations were observed for three other SNPs. Our results suggest that polymorphisms of CYP11A1 are related to breast cancer susceptibility in Han Chinese women of South China.Exposure to endogenous sex hormones has been reported as a risk factor for breast cancer. The  BRCA1/2, TP53 and PTEN), moderate penetrance genes , and low penetrance genes   repeat polymorphisms. Here, we investigated the associations between single nucleotide polymorphisms (SNPs) in CYP11A1 and breast cancer among Han Chinese women in Guangdong province, South China. Six SNPs in CYP11A1  were genotyped to perform a case-control study in women from Guangdong province.Previous reports indicate that prolonged exposure to endogenous sex hormones, especially estrogens and progestogens, can increase the risk of breast cancer ,15. Highynthesis . CYP11A1t cancer –19, polyt cancer , prostatt cancer ,23, and t-test indicated a significant difference in age distribution between the two groups , so all statistical analyses were subsequently adjusted by age.All subjects included in this study were Han Chinese women. The mean ages of the patients and controls were 48.48 ± 10.18 and 44.59 ± 11.26 years, respectively. An independent-sample P > 0.05) with minor allele frequency (MAF) ≥ 0.22 (All SNPs conformed to Hardy-Weinberg equilibrium (HWE) among both cases and controls  ≥ 0.22 . The SNP) ≥ 0.22 . D′ valuCYP11A1 were considered in the analysis for all subjects. The results of individual haplotype analysis and breast cancer are shown in OR, 0.64; 95% CI, 0.48–0.86; P = 0.0033).Six main haplotypes of OR, 0.75; 95% CI, 0.58–0.96; P = 0.023) compared with the wild-type TT. Individuals carrying the C allele showed protection from breast cancer . In contrast to rs2959008, the homozygous TT variant and allele T of rs2279357 were associated with elevated susceptibility to breast cancer . A correlation between the rs2959003 polymorphism and breast cancer was also observed under the log-additive model. Three other SNPs  did not show significant differences between the groups in the present study  .A further association analysis was conducted to identify the interactions of two susceptibility-associated SNPs, rs2959008 and rs2279357, and their impact on the risk of breast cancer. The results indicated that the united genotype TT-TT of rs2959008 and rs2279357 showed increased breast cancer risk (= 0.003) . There wCYP11A1, with the first four in introns and the latter two upstream of the gene. The results showed that rs2959008 and rs2279357 of the CYP11A1 gene were strongly associated with breast cancer risk, as well as the interaction of the two SNPs.Breast cancer risk is partially determined by endogenous hormone levels . Given tCYP11A1 gene polymorphisms. The best studied region is the pentanucleotide [TTTTA]n repeat (D15S520) located 528 bp upstream of the translational initiation site, which showed six common polymorphisms with four, six, seven, eight, nine, or ten [TTTTA] repeats. In the Chinese population, the main variants are the four-, six-, and eight-repeat alleles, which are correlated with risks of breast cancer [4 and [TTTTA]6 alleles were also shown to be associated with prostate cancer in other ethnic groups [n] polymorphism was different from common biallelic variations in the present study.Numerous studies have examined t cancer ,17 and pt cancer . The [TTc groups ,23. Howeet al. systematically evaluated genetic variations spanning 67 kb of CYP11A and suggested that common variations in CYP11A may be related to breast cancer susceptibility in a western Multiethnic Cohort Study, but no particular SNP or haplotype was identified as the most likely causal variant [CYP11A1 were also confirmed to be associated with hypertension and blood pressure (BP) in the Japanese population, with the presence of more of these alleles indicating higher risk [CYP11A1 was associated with increased risk of breast cancer among Chinese women in Shanghai, but three of our SNPs  were not included [Setiawan  variant . Mammogr variant . In Swed variant . Another variant . Along wher risk . These rincluded . Here, wThe subjects participated in our study were recruited between April 2009 and October 2011 from Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong Province, China. All participants were permanent residents of Guangdong from the Han Chinese population. Clinical information for each subject was collected from medical records.A total of 621 female patients with breast cancer were recruited after confirmed by pathological diagnosis through breast clinics. 550 (88.6%) of them were approached with their written informed consent and blood samples were collected. The age ranged from 22 to 80 years old. Other 71 patients refused to provide blood samples. Blood samples from 14 patients were unusable for DNA extraction due to transportation and storage at −70 °C After genotyping, 6 samples were excluded as all six SNPs could not be determined. Therefore, 530 case subjects were included in the final statistical analysis.550 healthy control women were randomly selected from outpatients ranging in age from 16 to 84 years old during the same period with no history of cancer or other breast-related diseases as determined by molybdenum target mammography and color Doppler ultrasonography and blood samples were collected. However, 4 samples failed in genotyping and 546 healthy controls were included in the final statistical analysis.The study protocol was approved by the Clinical Research Ethics Committee of Nanfang Hospital and written informed consent was obtained from all participants.Peripheral blood samples were collected from the participants and stored at −70 °C until DNA extraction. Genomic DNA was extracted from peripheral blood samples using an E.Z.N.A.™ blood DNA kit  according to the manufacturer’s protocol.For each SNP, a pair of amplification primers and an extension primer was designed using Assay Design 3.1 software . Genotypes were generated using the SEQUENOM MassARRAY matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry platform  according to the manufacturer’s instructions. This is one of the most powerful tools for detecting insertions, deletions, substitutions, and other polymorphisms in amplified DNA, and allows rapid, efficient, and high-throughput detection without any interactions. The overall call rates ranged from 99.63% to 100%.OR) and 95% confidence interval (95% CI) were evaluated by binary logistic regression analyses. There was a significant difference in age distribution between case and control groups as described previously, so the age was adjusted by setting as a covariate in association analyses for each SNP. Statistical analyses were implemented using SPSS 13.0 software  and the Web-based tool SNPstats [P < 0.05 was regarded as statistically significant.Hardy–Weinberg equilibrium (HWE) and linkage disequilibrium (LD) for the six SNPs were calculated using Haploview 4.2 . The genotype and allele distributions as well as the interactions of SNPs between case and control subjects were compared. Multiple inheritance models  were chosen to evaluate the associations between each SNP and breast cancer risk. The odds ratio (SNPstats . All comCYP11A1 gene were significantly associated with breast cancer susceptibility among Guangdong Han Chinese women, and the interaction of these two SNPs was associated with elevated risk. The haplotype CCCCGA of our six SNPs also had a protective effect against the development of breast cancer. The biological mechanisms behind these associations remain unknown. Therefore, further comprehensive investigations of steroid hormone biosynthesis and metabolism gene variations combined with other risk factors are required to identify biomarkers for inherited breast cancer susceptibility.In summary, our results showed that rs2959008 and rs2279357 of the


Background. Breast cancer is a major public health issue and the most commonly diagnosed cancer for women worldwide. Despite lower incidence rates than those living in Western countries, breast cancer incidence among Chinese women has increased dramatically in the past 20 years. Nevertheless, there is a paucity of studies reporting the attitudes toward and practices of breast cancer screening among Chinese women. Methods. This cross-sectional study examined the practices, knowledge, and attitudes toward breast cancer screening (BCS) on a convenience sample of 400 Chinese women. Results. Among study participants, 75% of the women never had a mammogram and the top three barriers reported were low priority, feeling OK, and lack of awareness/knowledge toward breast cancer screening. The results from the logistic regression model showed increased self-efficacy; having performed monthly self-exams, and having had clinical breast exams in the past two years were significant correlates while demographic variables were not correlated with screening behaviors. Conclusion. The findings provide a foundation to better understand beliefs and practices of Chinese women toward BCS and highlight the critical need for general public, health professionals, and the health care system to work collaboratively toward improving the quality of breast cancer care in this population. Breast cancer is a major public health issue and the most commonly diagnosed cancer for women worldwide –3. DespiA recent study conducted by Ziegler and colleagues 2008) projects that based on a representative sample of Chinese women from both urban and rural areas, the age-standardized breast cancer incidence rate in China will be 87.8 per 100,000 women by the year 2021  projects.Not only is the overall incidence rate for breast cancer rising in China, but also, another intriguing fact is that women in China are diagnosed with breast cancer at an earlier age than Caucasians. In Hong Kong, the incidence of breast cancer occurs at the highest rate at the age of 40 years  while foEarly detection of breast cancer through regular screening modalities and enhanced treatment has been found to decrease mortality rates by 25–30% , 20. DurThe purpose of this descriptive study was to explore the relationships of sociodemographic characteristics, knowledge, and beliefs about breast cancer (BC) and their practices to the self-reported practice of BC screening (BCS) among women in China. The specific aims were to (a) describe the sociodemographic characteristics, knowledge, beliefs, and mammography screening practices of women in China ages 40 and older and (b) identify correlates of BCS practices.In this cross-sectional study, the data was collected from the Wuhan, the capital city of Hubei Province, an important central city as the political, economic, scientific and technological, cultural, and financial center in inland China. The study sample consisted of 400 Chinese women aged 40 years and older. This study protocol was reviewed and approved by the Institutional Review Board for conducting this research.The study participants were recruited at the gatherings of community centers, parks, and temples. The study aim was explained to eligible women, who were assured that their participation would be voluntary and confidential. After consents were obtained, the participants completed the survey, which took approximately 20–30 minutes.The study measurement survey included two parts: (a) Chinese Mammogram Screening Beliefs Questionnaire (CMSBQ) and three subscales were used in current study: (1) perceived self-efficacy , (2) perceived benefits subscale , and (3) perceived barriers subscale . Possible responses to the items ranged from 1 (strongly agree) to 4 (strongly disagree). Each subscale was scored by calculating the means of all item scores. The findings of benefits and barriers subscales among Asian women demonstrated excellent supportive psychometric properties with promising Cronbach's alphas above 0.70, and the results from the confirmatory factor analysis supported construct validity with good model fit indices , 24. TheData analyses were performed using the SPSS statistical software package (Version 18.0). Descriptive statistics, including percentages, means, and standard deviation (SDs), were calculated. Next, multivariate logistic regression analysis was performed to examine the associations of demographics, knowledge, beliefs, and behavioral factors with the probability of having received mammograms/ultrasounds after accounting for all other factors in the model. These associations were expressed in the form of adjusted odds ratios with 95% confidence intervals (CI).Qualitative analytic techniques were employed for the analysis of the open-ended questions. The primary researcher and two research associates reviewed and coded participants' responses separately to uncover emerging themes and categories of the responses. The coding of the responses was then discussed and verified by two senior investigators with expertise in cancer control and qualitative research. Once consensus on coding was reached by the research team, the set of themes/categories were finalized. To complete each content area, quotes from the responses have been highlighted to illustrate the category. Discrepancies between the two researchers were resolved through iterative discussions until consensus was reached. Both the content and frequency of the responses were then analyzed. The sociodemographic characteristics of the study participants are presented in  As shown in The study tool asked the respondents to name up to three of the most important reasons that prevented women from having regular breast cancer screening. In this sample, 346 women (86.5%) gave 807 responses related to their perceptions on barriers toward BC screening.Ten themes emerged from the participants' responses to the question of barriers to breast cancer screening . If a won = 145, 42%), and examples of responses included “awareness is not emphasized/enough,” “don't have related knowledge (on breast cancer or screening),” “don't know how to do screening,” and so forth. Among these responses, 22 (6%) were simply stated, “I'm too old to get screened.” About 33% of responses were related to the cost of having a mammogram; several women indicated that after retirement, they do not have insurance arrangements getting mammograms. Less than 10% of the valid responses could be classified as “logistics,” including “do not know where to go for a screening, “difficulty when doing the exams in a big hospital,” and “traffic/transportation.” Interestingly, “radiation exposure” was the least frequently identified barrier (1%) in the study sample.As shown in R2 of Cox and Snell and Nagelkerke showed that 14%–21% of variance of BCS practice was explained by the selected variables, and the model had an overall correct prediction of 71%. Among variables included in the model, demographic variables  did not have significant prediction with screening behavior while increased self-efficacy; having performed monthly self-exams and having had clinical breast exams in the past two years were significant predictors of having had a mammogram or an ultrasound in the previous five years. Among these variables, for those who reported having recent clinical breast exams done in the past two years, those who performed monthly breast self-exams and those who reported higher self-efficacy were more likely to get the mammogram or ultrasound done in the past five years , that is, women with higher knowledge scores were more likely to indicate that they plan to get screening in the future. Logistic regression analyses were conducted modeling the odds of ever having screening done within the previous five years. The While breast cancer incidence has been historically less prevalent in Asian countries compared to the West, recent statistic data reveal breast cancer incidence rates have had a 20–30% increase in China in the past decade according to China's urban cancer registries . To our The results provided evidence that few Chinese women in the study sample participated in any BCS modalities ; specifically, only 30% of women reported they had CBE done in the past two years and 70% of women never had a mammogram. The rates reported in the current study were even lower than the rates of (43–64%) in previous studies conducted in other regions of China  , 27 and Nevertheless, the mediating effects of psychosocial and cultural variables on the impact of breast cancer intervention in low-to-middle income countries (LMC) are still understudied . The perIn terms of knowledge and attitudes related to breast cancer screening, Chinese women participants in the current study had much lower knowledge levels about breast cancer and risk factors, perceived less susceptibility, and reported greater barriers to screening and lower self-efficacy compared to study findings on immigrant Chinese women in the U.S.  The studThe study participants were also cognizant about their lacking awareness in breast cancer. The data lent support for the recent release of the Consensus Statement from the Breast Health Global Initiative that indicated a lack of public awareness in the importance of early detection of breast cancer in LMCs . IdentifThe study found positive associations between ever having had a mammogram, having CBE within the past two years, and performing monthly breast-exams, that is, adoption of one screening practice was positively correlated with other breast screening modalities in the current study. This finding was consistent with a previous study conducted in the U.S. among immigrant Asian women . While C Financial cost emerged as one of the top five barriers for Chinese women in this study to access mammography screening. This finding is consistent with other studies done in Hong Kong and immigrant Chinese women in the U.S. Women's lack of resources, combined with cultural and attitudinal barriers, along with limited access to affordable mammography services directly affect the likelihood of BCS since women with low incomes may have many other competing priorities related to survival that make breast cancer screening a low priority. The responses of study participants suggest that they might be amenable to engage in more BCS if they were more aware of what was available to them and if they still have access to preventive services such as mammograms after they retire. In 2008, the Chinese Ministry of Health started a program on early detection and treatment of cancer sponsored by central financial assistance that provide CBE, mammography, and ultrasound for women in 30 provinces in China. Efforts to develop programs and/or policies to improve breast cancer screening will be more successful if the implementation includes strategies to increase cancer knowledge, awareness, and information and complementary efforts to help women address emotional, socioeconomic, and other barriers to clinical breast exam and mammography use.The current study had several limitations. First, due to the nature of self-report data, there may have been over- or under-reporting of screening practices. Future studies with a combination of self-report and verification from objective chart reviews are warranted to ensure the accuracy in reporting socially desired behaviors, such as cancer screening. Second, because the use of convenience sample and small sample size, this approach can lead to sampling error; therefore, the results cannot be generalized to other regions/provinces in China, a country with more than 1.3 billion citizens with diverse socioeconomic and cultural backgrounds. Nevertheless, based on the demographics in the current study sample, the study recruited relatively diverse groups of women with a wide range of income and educational levels.  Changes in breast cancer incidence and distribution of breast cancer patterns in Chinese women present opportunities for developing and implementing effective public health programs to promote BCS for women in China. It is essential that public health campaigns focus on the importance of early detection and prevention that target both general public and health care professionals working directly with their clients. The findings of this descriptive correlational study provide a foundation to better understand beliefs and attitudes of Chinese women toward BCS. The concept of screening while asymptomatic is lacking and a low priority toward BCS was observed in the current study. As a result, this study highlights a critical need among general public, health professionals, and the health care system to work collaboratively toward narrowing the gap in BCS and to improve the quality of breast cancer care in this population.


Inadequate understanding of risk among counselees is a common problem in familial cancer clinics. It has been suggested that graphical displays can help counselees understand cancer risks and subsequent decision-making. We evaluated the effects of a graphical presentation in addition to a frequency format on counselees’ understanding, psychological well-being, and preventive intentions.Design: Multicenter controlled trial.Setting: Three familial cancer clinics in the Netherlands.Participants: Unaffected women with a breast cancer family history (first-time attendees).Intervention: Immediately after standard genetic counseling, an additional consultation by a trained risk counselor took place where women were presented with their lifetime breast cancer risk in frequency format (X out of 100) (n = 63) or frequency format plus graphical display (10 × 10 human icons) (n = 91).Main outcome measures: understanding of risk , psychological well-being, and intentions regarding cancer prevention. Measurements were assessed using questionnaires at baseline, 2-week and 6-month follow-up.Baseline participant characteristics did not differ between the two groups. In both groups there was an increase in women’s risk accuracy from baseline to follow-up. No significant differences were found between women who received the frequency format and those who received an additional graphical display in terms of understanding, psychological well-being and intentions regarding cancer prevention. The groups did not differ in their evaluation of the process of counseling.Women’s personal risk estimation accuracy was generally high at baseline and the results suggest that an additional graphical display does not lead to a significant benefit in terms of increasing understanding of risk, psychological well-being and preventive intentions.http://ISRCTN14566836Current Controlled Trials  BRCA1 or BRCA2 mutation. The cumulative lifetime cancer risk for carriers of a BRCA1/BRCA2 gene-mutation is 60 to 80 percent [Risk communication in breast cancer genetic counseling aims to improve women’s understanding of cancer risks in order to facilitate informed decision-making . The ris percent , and theSeveral studies have shown that unaffected women with a family history of breast cancer tend to overestimate their breast cancer risk, even after genetic counseling, although underestimation also occurs ,5. InappIt has been shown that the way in which risks are presented may influence individuals’ interpretations of risk and their subsequent decisions ,10. RiskPeople often find it difficult to understand risks, especially when risks are presented as numerical estimates . The useAlthough graphical displays are suggested to enhance quantitative risk communication, empirical evidence from clinical settings is scarce and few studies have been in the form of controlled trials aimed at individuals at high cancer risk. In a prospective randomized controlled trial, Ghosh et al.  showed tThe main objective of this study was to evaluate the effect of a graphical risk communication format in addition to a frequency format in genetic breast cancer counseling with regard to the healthy counselees’ understanding of risk (risk accuracy and perception of the risk), psychological well-being, and intentions regarding cancer prevention. Based on the arguments discussed previously, it was expected that graphical displays added to a frequency format were expected to lead to better understanding ,22, and The BRISC study was designed as a controlled trial with repeated measures using questionnaires. For the present study, to analyze the effect of graphical presentation, two presentations of lifetime risk information to women were compared: frequency format vs. frequency format plus a graphical display of risk. For the graphical display of risk information, icon arrays were used. The effect of time-frame (age-related risk information) on outcome measures – also part of the BRISC study – will be considered in future analyses, given that this was a different kind of intervention where risks were not – or not only – presented in a different format but also different risks were presented. More details can be found elsewhere .Power calculation indicated that 60 women per condition were needed to detect a clinically relevant difference of 10-20% change in risk accuracy between the presentation formats . UnaffecBRCA1/BRCA2 mutation; the risk passing the mutated gene to children; and the women’s personal breast cancer risk as based on her family history.Immediately after a standard genetic counseling session with the clinical geneticist or genetic counselor , an additional consultation by a trained risk counselor took place in which risks were communicated in one of two ways, namely: 1) in a frequency format ; or 2) in a frequency format plus a graphical display of risk (10×10 human figure icons)  or one of three increased risk categories: ‘slightly increased’ (11-20%), ‘moderately increased’ (20-30%) or ‘highly increased’ (30-40%)), using percentages when presenting risks. The risk counselor was informed about the counselees’ breast cancer risk by means of a “checklist after standard counseling” that was filled out by the genetic counselor. The checklist also included information on whether the counselee or a relative had been offered DNA testing. The study was blinded, i.e. the genetic counselors did not know which group a woman was allocated to.With regard to understanding of risk, women’s risk accuracy and risk perception was assessed. Risk accuracy was measured with regard to two types of risks: 1) lifetime population breast cancer risk: a risk within plus or minus 10% of the communicated risk was defined as accurate, and 2) the woman’s own lifetime risk of getting breast cancer as compared to the counseled risk: a risk within their estimated risk category as provided by the risk counselor was defined as accurate. For some women the risk changed during the study due to the receipt of genetic test results and in those women accuracy was based on self-reported test results at 6-month follow-up.Risk perception was assessed using two items on a 7-point rating scale: “How likely do you think it is that you will get breast cancer during your lifetime” (very unlikely (1) - very likely (7); ‘Perceived likeliness’), and “Based on your feelings, how high is your lifetime risk of developing breast cancer” (very low (1) - very high (7); ‘Risk as feeling’).Psychological well-being was assessed using an adapted version of the Lerman Cancer Worry Scale (CWS)  . The scaOverscreening was addressed by looking at preventive intentions at 6-month follow-up. Intentions with regard to “having breast screening by a physician every 6 months”, and “having a screening mammography every year” were assessed on a 7-point scale (definitely not (1) to definitely (7)). This was best assessable only in the subgroup with lowest breast cancer risk (i.e. 10-20% risk) where this type of preventive screening is not recommended according to the Dutch guidelines, and thus low preventive intentions were considered as required based on actual risk.The process of counseling as perceived by the women was evaluated using the one-dimensional Dutch version of the Perceived Personal Control questionnaire (PPC) (α = .85) .Socio-demographic characteristics assessed included age, level of education, marital status, number of children, parents’ country of birth, religious activity, family history of breast cancer, i.e. the number of family members that have or have had breast cancer including the family relationship. At 6-month follow-up, women were asked about the results of DNA testing.P-values less than .05 were considered statistically significant. Characteristics that differed at baseline between presentation formats were entered as covariates in the further analyses. Logistic regression analyses were performed to assess differences in the proportion of women with accurate risk estimates (adjusted for pre-test (T0) risk accuracy) between frequency format vs. frequency format plus graphical display in risk accuracy at 2 weeks (T1) and 6 months (T2). Differences were reported with odds ratios (ORs) and 95% confidence intervals (CIs). Analysis of covariance (ANCOVA) was performed to test the effects of presentation format on perceived likeliness, risk as feelings, psychological well-being, preventive intentions and evaluation of counseling, by comparing pre-test- (T0) with post-test- (T1/T2) outcome measures. Mean differences in outcomes scores between presentation formats were reported with 95% CIs for significant differences.Comparisons of baseline characteristics between groups were made using chi-square tests for categorical variables and t-tests for continuous variables. Over a two-year period, 557 women (55% of the eligible counselees) gave their informed consent. After consent, 110 women were excluded, mainly because they did not meet the inclusion criteria or because they cancelled the appointment for the additional risk consultation. Another 37 women were excluded after being allocated to one of the additional risk presentation formats because the data on risk estimation/category were not yet available by the closure of the study. In total, 63 women received a frequency format and 91 women received a frequency format plus graphical display. The remaining 256 women received a different format (e.g. age-related risks), and the corresponding effect of time-frame will be presented elsewhere.Baseline questionnaires were missing for one woman in the frequency format group, and four women in the frequency format plus graphical display group; these women were excluded from the analyses, leaving 149 women in the analyses. Baseline demographic or other participant characteristics did not differ between the two groups (Table BRCA mutations (lifetime risk changed to 60-80%), three of whom had previously been informed that they had a slightly or moderately increased risk. For five women in both the frequency format group and the frequency format plus graphical display group, the risk changed from moderately or highly increased to a population risk of 10% due to true negative test results. Five women with true negative results were already in the lowest risk group (i.e. risk estimation did not change).At 6-month follow-up, 18 women in the frequency format group, and 23 women in the frequency format plus graphical display group reported that they had received DNA test results or were still waiting for their results. For 13 of these women, the risk category changed based on the results of DNA testing. In total, seven women reported that they had had a pathogenic mutation (true positive results) after DNA testing for In both groups there was an overall increase in the women’s accuracy of risk estimation from baseline to follow-up. At 2-week- and 6-month follow-up, no difference in risk accuracy was found between women who received the frequency format and those who received the frequency format plus graphical display Table .Cancer-specific worry and state anxiety decreased over time in both groups. No significant differences were found between the risk format groups with regard to women’s cancer worry (CWS) and STAI scores Table .For the subgroup of women with no or slightly increased risk of breast cancer, there was no difference between the groups in behavioral intentions with regard to breast cancer screening by physical examination or yearly mammography Table . In bothNo significant differences were observed between scores on PPC of women in the two groups Table .This study investigated the effect of a graphical presentation format in addition to a frequency format among healthy women with a family history of breast cancer attending genetic counseling. There was an overall increase in women’s accuracy of lifetime risk estimation from baseline to follow-up. However, no significant differences were found between the group who received the frequency format compared to the group who received the frequency format with a graphical display in terms of understanding of risk, psychological well-being and preventive intentions. No differences were found in women’s evaluation of the counseling process.The BRISC study is a field study; in other words, it is a clinical trial which offers a unique opportunity to evaluate effects of different formats of communicating risks to counselees having to make real-life decisions. The study was a multicenter trial encompassing three out of nine Dutch familial cancer clinics. Women were allocated to receiving a particular risk communication format depending on the time they entered the study. Not only were the direct effects of communication assessed but also the effects at 6-month follow-up. In interpreting the results, one must, however, realize that any effect of different formats for risk communication on people’s perception and decision-making is bound to be small, in particular since this study involved a standard counseling session before the risk communication consultation (intervention) where the risks already had been discussed. Moreover, the sample was powered to detect a difference in risk accuracy. However, within each condition women were already quite accurate with regard to their own risk estimate leaving little room for improvement in both groups. Furthermore, for some women in both groups the risk status had changed as a result of genetic testing, as one would expect to happen in real life. In these cases, risk was based on the self-reported test results, which might differ from their actual risk status. For logistical reasons, actual test results were not recorded in this study. Moreover, the study measured women’s intentions to preventive behavior, and it is well known that there is often a discrepancy between intentions and actual behavior. Some caution should be taken in generalizing the results to all women with familial cancer since women with low education and women of ethnic minority groups were underrepresented. It is well known that in general women who come to the familial cancer clinics are higher educated women and therefore not representative for the whole population. Also, at 6 months, loss to follow-up was 25% and 22% of the women in the frequency format and frequency format plus graphical display group, respectively. To study differences in preventive intentions, as we intended to do for a small subgroup of women with low risk, a larger sample is probably needed.This study showed no significant effects of an additional graphical display to a frequency format on the outcome measures. However, at 6-month follow-up, women who received the additional graphical display had higher risk perceptions, although not significantly higher, and, for women in the lowest risk category, higher intentions to have yearly mammography compared to women who received the frequency format only. There is mixed evidence regarding graphical displays improving understanding or aiding decision-making . EarlierIn this study, only icon arrays were evaluated, while other graphical displays such as bar charts may also be used. Waters et al. , for exaIt has been shown that inadequate perception of risk may lead to screening that is not consistent with the recommendations for their actual risk category . The preAlthough risks are generally assumed to be important for decision-making, the results suggest that the way in which risks are presented does not influence women’s intentions, either because the presentation format has no effect on their understanding of the risks, or because women do not consider risks important for their decisions. For counselees, the risk level, in whatever form it is presented, may be less relevant compared to other factors, e.g. emotions such as worry and pre-existing beliefs . It has Up till now, in most studies, the majority of unaffected women with a family history of breast cancer overestimated their breast cancer risk ,5. In coThe results of this controlled trial suggest that a graphical presentation in addition to a frequency format has no effect on understanding of risks, psychological impact and screening intentions. Further research is needed to establish whether this is indeed the case.The authors declare that they have no competing interests.LH, JO, CvA, FM and DT designed the study. CO coordinated the data collection. LC and PK planned the analysis. LC performed the statistical analysis. LH wrote the first draft of the manuscript. All authors contributed to the final manuscript and have approved it.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1472-6947/13/55/prepub