This study aimed to identify predictors of nonadherence to breast cancer screening guidelines in an urban screening clinic among high‐ and average‐risk women in the United States.not receiving recommended supplemental imaging for high‐risk women. We used t‐tests and chi‐square tests to examine bivariate associations with guideline‐congruent screening, and probit regression to regress guideline‐congruence unto breast cancer risk, breast density, and their interaction, controlling for age and race.We reviewed records of 6090 women who received ≥2 screening mammograms over 2 years at the Karmanos Cancer Institute to examine how breast cancer risk and breast density were associated with guideline‐concordant screening. Incongruent screening was defined as receiving supplemental imaging between screening mammograms for average‐risk women, and as p < 0.01). Among average‐risk women, incongruent screening was more likely among those with dense versus nondense breasts . Among high‐risk women, incongruent screening was more likely among those with nondense versus dense breasts . The significant main effects of density and high‐risk on increased incongruent screening were qualified by a density by high‐risk interaction, showing a weaker association between risk and incongruent screening among women with dense breasts  versus nondense breasts . Age and race were not associated with incongruent screening.Incongruent screening was more likely among high‐ versus average‐risk women (97.7% vs. 0.9%, Lack of adherence to evidence‐based screening guidelines has led to underutilization of supplementary imaging for high‐risk women and potential overutilization for women with dense breasts without other risk factors. In the Breast Cancer Surveillance Consortium, a population‐based cohort study, Hill et al. found that only 6.3% of high‐risk women received supplemental screening with breast MRI in accordance with screening guidelines (guideline‐congruent).Supplemental screening of average‐risk women has been shown to increase false positive results,Whereas prior studies have evaluated potential predictors of supplemental breast cancer screening among high‐risk women recruited either from a high‐risk screening clinic,22.1Eligible participants included Black and White women who presented to the Karmanos Cancer Institute (KCI) for at least two screening mammograms between January 1, 2016 and March 31, 2018. 2D screening mammograms were offered to women with nondense breast tissue, and 3D screening mammograms were offered to all women with dense breasts at the KCI. We chose to include only Black and White patients in this analysis because they comprise the largest groups of women seen at the KCI mammography screening clinic. We chose the start date to obviate any effects of Michigan's recently adopted breast density notification law (June 2015) on supplemental breast cancer screening behaviors. The end date reflects the termination of use of mammography workflow software at the KCI which automatically estimated each patient's breast cancer risk score (PenRad Technologies). Women were excluded if they had any diagnostic breast imaging (with and without a subsequent biopsy) in the interval between their two mammograms or if they developed breast cancer during the two‐year study period. Women were also excluded if they had a prior history of invasive breast or ovarian cancer, ductal carcinoma in situ (DCIS), lobular carcinoma in situ (LCIS), or atypical ductal or lobular hyperplasia. Consequently, our dataset consisted of mammogram and supplemental breast imaging results specifically for screening purposes among asymptomatic women with no prior breast or ovarian cancer‐related history. Of note, this paper describes the proportion of women who received supplementary screening at the KCI and reflects referrals that could have been made by any of the participants' health care providers. Therefore, the referrals that occurred at the KCI mammography clinic do not represent a standard of care referral process developed at the KCI. The study was approved by the Wayne State University IRB.2.2BRCA status, abnormal breast biopsy history, Ashkenazi inheritance, and family/personal history of breast cancer. Women were classified as high‐risk if their lifetime breast cancer risk was ≥20%; intermediate‐risk if their lifetime risk was between 15% and 20%; and average‐risk if their lifetime risk was <15%. Women who had a BI‐RADS Density classification of C or D  were classified as having dense breasts, whereas women with a BI‐RADS Density of A or B  were classified as having nondense breasts. Breast cancer risk and breast density data were recorded based on the most recent clinic encounter before the end of the study period.The age and race of women in the study population were extracted from clinic data. The age was divided by 10 and mean‐centered to facilitate analyses. The Tyrer‐Cuzick version 7 model of lifetime risk of breast cancer was included in the mammography report and was interpreted as an indicator of breast cancer risk for this analysis. The Tyrer‐Cuzick version 7 model is a risk assessment tool that incorporates the following variables to estimate lifetime risk of breast cancer: age, height, weight, age at menarche, parity, menopausal status, hormone therapy use, 2.3Women were classified as having supplemental breast screening if their clinic records indicated any breast imaging  for screening purposes in the interval between their screening mammograms. Whether women were undergoing diagnostic or screening mammograms was determined by whether the imaging appointment was categorized as diagnostic or screening within the electronic medical record system.Since screening guidelines for intermediate‐risk women leave greater latitude for shared decision‐making regarding screening regimens, we restricted the current analysis to women with high‐ and average‐breast cancer risk so that we could more accurately determine whether breast cancer screening practices were congruent or incongruent with risk‐based screening guidelines. High‐risk women were classified as incongruent if they did not receive supplemental imaging between screening mammograms; average‐risk women were classified as incongruent if they received supplemental imaging between screening mammograms.2.4t‐tests. All analyses were performed using the SPSS version 25.Using the full samples, we described the proportion of women who had supplemental breast cancer screening and examined the relationship between supplemental screening and potential explanatory factors including breast cancer risk, age at the time of the screening, and breast density using chi‐square and The analysis was restricted to high‐ and average‐risk women for whom we had data on breast density. We used a series of path analysis with probit regression to examine predictors of incongruent screening in three steps. In the first step, we regressed the indicator for incongruent screening for age at screening and racial group; and in the second step, we introduced the main effects of breast density and breast cancer risk (high vs. average as measured by the Tyrer‐Cuzick model). In the final step, we introduced a risk by density interaction to examine whether the effects of risk on incongruent screening were influenced by breast density. Models were fit with lavaan in R version 4.1.2.32 (2) = 49.16, p < 0.01) and were also more likely to be older compared with Black women . Overall, 23.8% of the women in the sample were identified by radiologists as having dense breasts. White women were more likely to have dense breasts compared to Black women (31.3% vs. 21.1%: χ2 (1) = 64.11, p < 0.01).Our sample consisted of 4462 Black and 1628 White women who had multiple screening events between January 1, 2016 and March 31, 2018. Most women (86.5%) were at average‐risk for breast cancer, while 7.7% were at intermediate‐risk, and 5.8% were at high‐risk. The clinical and demographic characteristics of the average‐ and high‐risk women in our sample are listed in Table 2(2) = 9.22, p = 0.01). Younger women  and women with dense breasts  = 80.65, p < 0.01) were also more likely to have supplemental breast cancer screening. There were no racial differences in the rate of supplemental screening.Sixty‐one women (1%) in our sample had supplemental breast cancer screening. Of the women who received supplementary imaging, 32 were classified as having dense breasts; five were classified as having nondense breasts; and breast density data was missing for the 25 remaining women. Overall, women at high‐ (2.3%) and intermediate‐ (1.7%) risk were more likely to have supplemental breast cancer screening than women at average‐risk . Women were more likely to be incongruently screened if they were younger , had dense breasts  = 59.80, p < 0.01), or were White  = 15.85, p < 0.01).Restricting the analysis to average‐ and high‐risk women, 390 (6.9%) of these women were incongruently screened which included 343 (97.7%) out of all high‐risk women and only 47 (0.9%) out of all average‐risk women (2 (1) = 62.09, p < 0.01) or if they were younger . High‐risk women were more likely to be incongruently screened if they had nondense breasts  = 7.04, p < 0.01), or if they were older . There was no significant impact of race (Black vs. White) on the rate of incongruent screening for average‐ or high‐risk women.We also looked at factors associated with incongruent screening stratified by high‐ versus average‐risk status. Average‐risk women were more likely to be incongruently screened if they had dense breasts ; however, the effects of age and race were no longer significant with the addition or risk and high density in subsequent steps. The significant main effects of increased breast density and high‐risk status on a higher rate of incongruent screening as shown in step 2 were qualified by a density by high‐risk interaction indicated in the final model listed in step 3. Probing this interaction indicated that the association between breast cancer risk and incongruent screening was weaker among women with dense breasts  compared with those without dense breasts . In other words, women with dense breasts were less likely to be incongruently screened regardless of breast cancer risk. Examining the marginal probabilities consistent with the final model  of women were supplementally screened, 6.9% of women who were either at average‐ or at high‐ risk for breast cancer had screening which was inconsistent with guidelines. Notably, 97.7% of high‐risk women in our sample were incongruently screened, meaning they did not receive supplementary screening when indicated. Guidelines indicate that high‐risk women should be screened with supplemental MRI in addition to screening mammogram,Identifying barriers to guideline‐congruent screening among high‐risk women informs interventions to directly address inadequate supplementary screening management for at‐risk populations. The few studies that have explicitly examined barriers to supplemental screening among high‐risk women suggest that lack of knowledge  and guideline‐inconsistent recommendations from their physicians contribute to low rates of supplemental screening.Regarding knowledge, data from a sample of more than 11,000 women showed that knowledge of objective breast cancer risk factors are not associated with mammography screening attendance.Further, our multivariable analysis showed that women with dense breasts were less likely to be incongruently screened regardless of breast cancer risk. This weaker association between breast cancer risk and incongruent screening among women with dense breasts may be due to overscreening among average‐risk women with dense breasts. These findings support the ongoing uncertainty about recommendations for breast cancer screening for patients with dense breasts, despite increased density being an established risk factor. Given that there are currently no standardized guidelines directing management of breast cancer screening in the setting of dense breasts, clinicians may be inclined to recommend supplementary imaging regardless of recommendations since increased breast density is a known risk factor for breast cancer.Guidelines also indicate that average‐risk women should refrain from supplemental screening unless clinically indicatedWhile increased breast density is an independent risk factor for breast cancer, the extent to which breast density affects risk for breast cancer is not absolutely established. Relative risk conferred by breast density in part depends on a woman's other risk factor patterns.Our study showed no racial differences in incongruent breast cancer screening in our study sample. This differs from prior studies which reported that Black women were more likely to be incongruently screened compared with their White counterparts following passage of density notification laws in MI.4.1These data are limited in that they originated from one screening center in an academic, metropolitan setting, which may limit generalizability. Nonetheless, as the screening clinic's population is majority Black, and represents the large and diverse screening population in the Metro Detroit, we feel that our results are at least generalizable to the population in the downtown KCI's catchment area. Our data did not account for variation due to physicians who referred women to the screening clinic for mammograms; therefore, the extent to which incongruent screening might be due to sociodemographic characteristics of referring physicians is unknown. Additionally, while supplementary screening for women at high‐risk for breast cancer has long been a well‐established guideline, it is important to note that any screening recommendation made by a clinician depends on a wide variety of factors not measured in this study, such as patient willingness to complete the imaging, social or financial barriers, or medical comorbidities which may influence whether a clinician thinks supplementary imaging is in the patient's best interest. Furthermore, our inclusion criteria may have resulted in some degree of selection bias in that we may have missed women who had a screening mammogram, followed by supplemental screening in the interval, but who did not return for a screening mammogram within the recommended two‐year interval .Future studies may account for physician characteristics and other sources of variation in women's screening behaviors . Finally, we did not measure the women's perceptions of breast cancer risk, or attitudes toward supplemental screening; hence, potentially important individual‐level psychological predictors were absent in the data collected for this study. Despite these limitations, the demonstration of the scope of guideline‐incongruent screening among high‐risk women, and of the ostensible influence of breast density notifications on guideline‐incongruent screening among average‐risk women contribute to the strength of these data. Further, a major strength of our study is the large sample size of Black women compared to prior studies that assess the utilization of guideline‐concordant screening for breast cancer.5Whereas health organizations promote evidence‐based guidelines for supplemental breast cancer screening in service of patient health, the lack of adherence to these guidelines is concerning. This concern is pressing for women who are at high‐risk for breast cancer since they were most likely to be incongruently screened, yet they are potentially most likely to benefit from the increased surveillance proffered by supplemental screening with MRI. These findings support the need for interventions to support guideline‐congruent behavioral decision‐making related to supplemental breast cancer screening so that we may ensure that women are more likely to partake in secondary screening measures in line with evidence‐based recommendations.Alexandra Wehbe: Conceptualization (lead); data curation ; formal analysis ; investigation (lead); methodology ; project administration (lead); resources ; software ; writing – original draft (lead); writing – review and editing (lead). Madeleine Gonte: Conceptualization ; investigation ; writing – original draft ; writing – review and editing . Suzanne C. O'Neill: Conceptualization ; data curation ; investigation ; methodology ; writing – review and editing . Alit Amit‐Yousif: Conceptualization ; data curation ; investigation ; methodology ; writing – review and editing . Kristen S. Purrington: Conceptualization ; formal analysis ; methodology ; validation ; writing – review and editing . Mark Manning: Conceptualization ; data curation ; formal analysis (lead); investigation ; methodology ; writing – original draft ; writing – review and editing . Michael Simon: Conceptualization ; investigation (lead); supervision (lead); writing – review and editing .There were no sources of funding.The authors have no conflicts of interest to disclose.This study was approved by the Institutional Review Board (IRB). According to IRB guidelines, neither approval from the ethics committee nor informed consent from the study populations is required for this study.This study did not use material from other sources.


Globally, the burden of breast cancer has increased significantly in recent decades. Emerging evidence suggested that endocrine-disrupting chemicals (EDCs), which have the potential to interfere with the function of normal hormones, may play a crucial role in this trend. However, the potential relationships were inconsistent in various studies.In our study, we sought to fully evaluate the currently available epidemiological evidence to ascertain whether certain EDC congeners and their metabolites are related to breast cancer risk. Following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines, we conducted a comprehensive literature search of original peer-reviewed publications in three electronic databases: PubMed, Web of Science, and Embase. Publications that covered xenobiotic EDC exposures and breast cancer–confirmed histological results or antecedent medical records or reporting to health registers were taken into consideration.The final result of the literature search was 6,498 references, out which we found 67 publications that matched the requirements for meta-analysis and eight publications for qualitative trend synthesis. In this meta-analysis, statistically significant associations revealed that (i) 1-chloro-4-benzene  and its major metabolite 2,2-bis(4-chlorophenyl)-1,1-dichloroethylene  were somewhat related to a greater risk of breast cancer. However, this relationship only existed in blood serum but not in adipose tissue. (ii) Breast cancer risk was increased by exposure to chlordane and hexachlorocyclohexane. (iii) Five polychlorinated biphenyls  can increase the risk of breast cancer. (iv) One phthalate congener (BBP) and one per- and polyfluoroalkyl substance congener (PFDoDA) were negatively associated with breast cancer risk. Unfortunately, heterogeneity was not well explained in our review, and a limited number of available prospective studies investigating the associations between EDC exposure and breast cancer were included in our meta-analysis. To elucidate the overall associations, future large, longitudinal epidemiological investigations are needed.https://www.crd.york.ac.uk/PROSPERO/, identifier CRD 42023420927. The global burden of breast cancer is increasing significantly. According to GLOBOCAN 2020 2021), an estimated of 2.3 million new breast cancer cases were diagnosed in 2020, which contributed to the most female cancer deaths globally  and Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines .The available research on EDC exposure and breast cancer was identified through a comprehensive peer-reviewed of original literature search in three electronic databases, namely, PubMed, Web of Science, and Embase, from 1961 to May 2023. The identified search terms were divided into three search blocks: the first dealt with the EDC exposure, the second covered the outcome (breast cancer), and the last covered study design (case−control and cohort study). A manual search of the included article’s reference lists was subsequently performed. The search protocol provided the search specifications and respective hits in each search block  were included in the meta-analysis. The following 10 compound groups of EDCs were investigated in the included publications: (i) BPA, (ii) dioxins, (iii) parabens, (iv) phthalates diesters and their metabolites, (v) flame retardants, (vi) PAHs, (vii) PCBs, (viii) organochloride pesticides, (ix) PFAS, and (x) triclosan.Breast cancer: Breast cancer–confirmed histological results or antecedent medical, records, or reporting to health registers were taken into consideration.Risk estimates  and their 95% confidence intervals (95% CI) as an outcome according to higher versus lower levels of EDC exposure contacts within the given study.Only cohort studies and case−control studies were included in our analysis.Criteria for exclusion of studies were as follows:Research studies conducted on animals, case reports, cross-sectional researches, reviews, conference proceedings/abstracts, letters, editorials, and comments were not included in our analysis.Publications that discussed prescription hormones, did not report risk estimates, or reported repeated estimates from other publications were excluded.Publications that reported DDT, PCBs, PFAS, and phthalate congener summary estimates but no specific risk estimates were excluded.Self-reported breast cancer was excluded.The process of data extraction was performed independently by KR and YS, and any inconsistency was resolved by JL. The following information was extracted from each publication including author, location, study design, number of cases and referents, biospecimens, exposure contrast, and substance. Risk estimates with 95% CIs were recorded for each measured compound. When risks according to several levels of exposure were reported, The risk estimate of the highest versus lowest levels was chosen. If a study reported that the OR value was in both unadjusted and adjusted models, then we gave preference to the adjusted OR value.2-test statistic and p-value. Significant heterogeneity was defined as I2 > 50% or value of p < 0.10. Low, moderate, and high degrees of heterogeneity were defined with I2-values of 25%, 50%, and 75%  and sample types were human specimens. Separate forest plots for each EDC exposure were conducted to illustrate summary ORs with 95% CIs. The random-effects model was used to summarize the risk estimates. A meta-analysis was performed independently when ≥3 studies reported the compound. Heterogeneity was assessed using the degree of I and 75% . SubgrouThe process of risk of bias and quality assessment was performed independently by two authors (KR and YS), and any inconsistency was resolved by a third author (JL). Each study was assessed for the completeness of reporting using a standardized form adapted from . There aThe relationship between pesticides and breast cancer has received the most attention because of their persistence in the environment . However2 = 77.7%, P < 0.001) , whereas the blood serum p,p′-DDT was associated with an increase in breast cancer   , and the OR for blood serum was close to unity but not significant    (There were twenty-eight case−control articles and eight nested case−control studies (< 0.001) Figure 22 = 77.4%). The heterogeneity was not affected by subgroups of sample type . The heterogeneity was affected by subgroups of sample type and study design. In blood serum, the concentrations of HCH were associated with a significant increase in breast cancer . The summary OR for HCH in adipose tissue was significantly reduced  with no heterogeneity  (Sixteen case-referent studies provided twenty-two risk estimates for hexachlorocyclohexane (HCH). There were 12 case−control articles and four nested case−control studies . No significant increase were observed in other pooled ORs. The results are summarized in There were 17 publications that reported the associations between other pesticide exposure and breast cancer. 2 = 74.0%; OR, 1.33; 95% CI, 1.10–1.60; I2 = 52.9%)  (2 = 24.6%)  11. The 2 = 33.0%; P = 0.1888). However, the overall ORs for DBP, DEHP, DEP, and DIBP were not statistically significant (The characteristics of the studies included in our review are shown in 2 = 21.7%). The results are summarized in The characteristics of the studies included in our review are shown in 2 = 45.1%).There were only four publications for polybrominated diphenyl ethers (PBDEs) included in our meta-analysis. The characteristics of the studies are shown in The characteristics of the studies included in our meta-analysis are shown in ad-hoc grouping comparison of median values in cases and controls rather than divided by tertiles and quantiles (or more detailed) or by grouping of levels. For example, this case–control study that only contrasted the median values in cases and control has assessed as high risk of exposure contrast  upgraded PCBs to group 1 “Carcinogenic to humans” in 2015, on the basis of sufficient evidence of an excess risk for melanoma . In receThere are numerous unavoidable and accidental causes of exposure to BPA, phthalates, PBDEs, and PFASs in daily life. We found that one PFAS congener (PGDoDA) and one phthalate congener (BBP) were passively linked with the risk of breast cancer. However, only three publications addressing PGDoDA matched the requirements for meta-analysis. More studies are needed to identify the association. The impact of other phthalates, PFAS, BPA, and PBDE on breast cancer was not sufficiently supported by the results.In addition, there were four compound groups of EDCs that were not eligible for meta-analysis because of limited studies. Further larger population-based studies are needed to clarify the real relationship between environmental EDCs and breast cancer.There were only two studies addressing the relationship between parabens and breast cancer. Wu et al. found that breast cancer was inversely associated with total parabens  in a nested case−control study in 2021 . Parada We have rigorously assessed the epidemiological data on 10 compound groups of EDCs  and breast cancer and, finally, summarized risk estimates for six compounds . To our knowledge, we have for the first time summarized the relationship between HCH and breast cancer, and found that HCH was positively related to breast cancer risk. Meanwhile, the relationships between several common EDC congeners and breast cancer have been updated. such as DDT, PCBs, and phthalates. This facilitated a better understanding of the association between each type of EDCs and breast cancer. Unfortunately, heterogeneity was not well explained in our review, and a limited number of available prospective studies investigating the associations between EDC exposure and breast cancer were included in our meta-analysis. More attention was given to women, not men, perhaps because breast cancer is more common in women. To elucidate the overall associations, future large, longitudinal epidemiological investigations are needed.In this meta-analysis, statistically significant associations revealed that (i) p,p′-DDT and its major metabolite p,p′-DDE were somewhat related to a greater risk of breast cancer. However, this relationship only existed in blood serum but not in adipose tissue. (ii) Breast cancer risk was increased by exposure to chlordane and HCH. (iii) Five polychlorinated biphenyls  can increase the risk of breast cancer. (iv) One phthalate congener (BBP) and one PFAS congener (PFDoDA) were negatively associated with breast cancer risk. Our meta-analysis suggested that exposure to a few specific EDCs was identified as a risk factor for breast cancer. More effective preventive measures should be taken to control the environmental pollution of EDCs.The original contributions presented in the study are included in the article/HL: Software, Data curation, Formal analysis, Writing – original draft. YKS: Data curation, Formal analysis, Writing – review & editing. LR: Data curation, Formal analysis, Writing – review & editing. JL: Data curation, Formal analysis, Writing – review & editing. YFS: Data curation, Formal analysis, Writing – review & editing. CM: Data curation, Formal analysis, Writing – original draft. CH: Conceptualization, Data curation, Project administration, Writing – original draft.


IntroductionBreast cancer (BC) is one of the most common cancers worldwide and it considerably increases morbidity and mortality globally. Screening methods, such as self-examination, clinical examination, and mammography, can help in early detection and treatment, which will help in improving survival rates and reducing mortality. While regular screening of the breast is essential to detect the earliest stages of breast cancer, not all women adhere to regular breast screening.MethodsA cross-sectional study was conducted in Jeddah, Saudi Arabia, between December 2021 to July 2022. using an online self-administered questionnaire. The total number of responses (n = 328), Data was analyzed using SPSS 25.ResultsIn this study, out of the 328 respondents, 18.9% reported undergoing regular mammography, 14.3% reported having regular clinical breast examinations, and 38.1% reported practicing regular breast self-examinations.. In addition, the participants' most known warning signs of BC were a lump under their armpit (69.1%). the most perceived barrier to breast self-examination (BSE) was Doing a breast examination will make her worry about what is wrong with her breast (47%). whereas the most barrier to clinical breast examination (CBE) was embarrassment (45.9%). On the other hand, the main barriers that prevented women from having mammograms were embarrassment (36%) and pain (32.6%).ConclusionThe most perceived barrier to BSE was women’s concern, while embarrassment and painful procedures were significant barriers to performing mammography and CBE. Therefore, adult females in Jeddah need educational programs to improve their knowledge and increase public awareness of breast cancer screening for early detection. Breast cancer (BC) has surpassed lung cancer as the most commonly diagnosed cancer among women, with an estimated 2.3 million new cases, accounting for approximately 11.7% of all cancer diagnoses. 5]. Acco. Acco5].Breast self-examination (BSE), clinical breast examination (CBE), and mammography are three BC screening (BCS) techniques that are helpful in identifying the disease early . Breast Even while routine breast screening is necessary to identify BC in its initial stages, not all women do it . AccordiPrevious studies have identified various factors deterring women from participating in BC screening, such as emotional issues, lack of awareness about the consequences, and avoidance of the process ,19. DiffThe barriers to BC screening for different screening techniques among women in the city of Jeddah were not considered in earlier studies of comparable scale. The purpose of this paper is to understand some of the social, physical, and cultural barriers that are playing a role in assisting early detection, preventive measurements, and reducing the incidence of BC among women in Jeddah, Saudi Arabia, 2021.This is an analytic cross-sectional study aimed to assess the barriers to using different screening methods for breast cancer among adult females in Jeddah, Saudi Arabia, in 2021. This study was approved by the institutional review board of King Abdulaziz University (Reference No 29-22). Between December 2021 and July 2022, an online survey was conducted among adult women aged 20 years and above residing in Jeddah. The survey was distributed via popular social media platforms such as WhatsApp and Snapchat, utilizing publicly listed phone numbers obtained from a national telecom company. Participant confidentiality and anonymity were ensured, and the online format allowed for private responses. The questionnaire used neutral language and employed snowball sampling to gather diverse participant responses. Initially, 345 responses were received, but after excluding individuals who did not meet the criteria of being adult women aged 20 years or older residing in Jeddah, we obtained a final sample size of 328 responses out of the calculated sample size of n=377, resulting in a response rate of 87%. The sample size calculation was done using Rao software with a 95% confidence interval and a 5% margin of error based on the population size .The survey used in this study was obtained from a previous research project without modifications . To ensuThe third section contained the Modified Champion Health Belief Model Scale (CHBMS) which is a valid and reliable tool to measure susceptibility, seriousness, barriers to BSE, barriers to CBE, and mammography barriers, as well as confidence in BSE efficacy and health motivation . ParticiStatistical analyses were done using IBM SPSS Statistics for Windows software version 26  to test and evaluate the hypothesis. Descriptive statics of frequency tables, cross-tabulation, and percentages were constructed using the software. The relationships between performing BSE on a regular basis and participants' age, marital status and educational level were conducted using the Chi-squared test (χ2). Furthermore, multivariate logistic regression analysis of the barriers to BCS was done with a 95% CI to assess the risk factors (independent predictors). A p-value of less than 0.05 was considered statistically significant.A total of 328 women took part in this study. Most participants (55.2%) had an age ≥45 years, 69.8% were married, and 79.6% had a university level of education and above. Of the participants, 46.3% had a family member or friend who experienced breast cancer and only 5.8% had ever been diagnosed with breast cancer Table . Knowledge of BC and risk factors of BCThe most known warning signs of BC by the participants were a lump in your breast 69.1%), discharge or bleeding from the nipple (55.7%) and changes in the shape of the breast or nipple (43.6%). And the most known risk factors of BC were having a close relative with breast cancer (49.5%) and having a history of breast cancer 44.9%  shows that participants who underwent mammography or clinical breast examination on a regular basis had a significantly higher percentage of having age ≥45 years and being married (p=<0.05). On the other hand, a non-significant relationship was found between undergoing mammography or clinical breast examination on a regular basis and participants' educational level (p=>0.05).Multivariate logistic regression analysis was done to assess the risk factors (independent predictors) of having any barrier to breast examination Table . It was The aim of this study was to assess the level of knowledge of breast cancer and screening measures as well as some of the sociodemographic barriers that play into the decision of screening for BC among women in Jeddah, Saudi Arabia. Participants had a good level of understanding of BC, including its symptoms and risk factors. However, practising BCS was low for self-examination, clinical screening, and mammography, which curbs early detection and recovery. This is in line with similar studies in the literature ,18.Breast cancer screening is provided free of charge to eligible females in Saudi Arabia within the recommended screening age group . It playWe used the CHBMS questionnaire to investigate barriers to BCS among women. The factors that influence BSE behavior include barriers, health motivation, perceived benefits of BSE, and susceptibility. Most participants were aware of how serious BC is, so fear and anxiety were common themes and screening for BC is worrisome, so most women avoid it. While self-confidence in the ability to perform BSE was high, most participants cannot remember to periodically do the examination and felt embarrassed to have the doctor examine them and/or do mammography. This finding is consistent with similar studies in the literature -22. A stRegarding preventive practices to detect BC, one of the key findings is that having BCE is statistically dependent on age above 45 and a history of BC diagnosis. Furthermore, conducting a self-examination was also statically dependent on age above 45, having a history of breast cancer, and higher levels of education. Furthermore, education was also a strong factor, along with age and previous history of BC diagnosis, for women to go under mammographic examination regularly, which is in line with a previous study conducted in Saudi Arabia by Al-Wassia et al. Education level was found to be strongly associated with higher mammography knowledge . In contThe study has several limitations. First, the use of the snowball sampling method carries a significant risk of selection bias. The reliance on participants to refer others from their social networks may introduce a non-random element into the sample, potentially excluding individuals who are less connected or have different characteristics. Therefore, caution should be exercised when generalizing the findings to the broader population of adult women in Jeddah, or other regions in Saudi Arabia. Second, although various measures were implemented to control for social desirability bias, such as ensuring anonymity, conducting the survey online, and using neutral language, it is important to recognize that complete elimination of this bias is challenging, and self-reported data may still be influenced by it. Third, there is a possibility of reporting bias since the data relied on self-reports. Fourth, as the data was cross-sectional, causality cannot be established, and caution is advised when interpreting the results. Lastly, while this study examined the factors influencing breast cancer screening uptake among adult women in Jeddah, Saudi Arabia, it is important to acknowledge that certain key determinants were not specifically investigated. Factors such as the quality of screening services, the skill level of healthcare providers, cultural beliefs surrounding breast cancer screening, and the availability of social support for women were not explored in depth. Future research should focus on examining these aspects to better understand their influence on screening uptake and to develop targeted interventions that address these crucial determinants.While the level of knowledge regarding BC among women in this study was high, utilization of different screening techniques, despite their availability, was low. To address this, a comprehensive approach is needed. Women should be educated about the benefits of breast cancer screening and encouraged to undergo it. Initiatives should focus on improving women's comfort during screening. Additionally, assessing the quality of breast cancer screening services in Saudi Arabia is important to ensure they meet global standards. Collaboration among healthcare providers, policymakers, and stakeholders is crucial to develop and implement comprehensive screening programs that target emotional barriers and avoidance.


Laboratory studies have indicated that a cholesterol metabolite and selective estrogen receptor modulator, 27-hydroxycholesterol (27HC), may be important in breast cancer etiology and explain associations between obesity and postmenopausal breast cancer risk. Epidemiologic evidence for 27HC in breast cancer risk is limited, particularly in multiethnic populations.In a nested case–control study of 1470 breast cancer cases and 1470 matched controls within the Multiethnic Cohort Study, we examined associations of pre-diagnostic circulating 27HC with breast cancer risk among African American, Japanese American, Native Hawaiian, Latino, and non-Latino White postmenopausal females. We used multivariable logistic regression adjusted for age, education, parity, body mass index, and smoking status. Stratified analyses were conducted across racial and ethnic groups, hormone receptor (HR) status, and use of lipid-lowering drugs. We assessed interactions of 27HC with steroid hormones.27HC levels were inversely related to breast cancer risk , but the association was not statistically significant in the full model. Directions of associations differed by racial and ethnic group. Results suggested an inverse association with HR-negative breast cancer . 27HC interacted with testosterone, but not estrone, on risk of breast cancer; 27HC was only inversely associated with risk among those with the highest levels of testosterone .This is the first US study to examine circulating 27HC and breast cancer risk and reports a weak inverse association that varies across racial and ethnic groups and testosterone level.The online version contains supplementary material available at 10.1186/s13058-023-01693-6. Breast cancer is the most common cancer and the second leading cause of cancer death among US females . Obesity27-Hydroxycholesterol (27HC) is a cholesterol metabolite, the most-abundant circulating oxysterol, and an endogenous selective estrogen receptor modulator (SERM) –21. 27HCThere are substantial racial and ethnic disparities in breast cancer risk in the US; breast cancer incidence rates range from a high of 172.1 per 100,000 person years among Native Hawaiian to a low of 92.1 among Hispanic females , 33. AddTo our knowledge, there is no epidemiologic study that has assessed the role of circulating 27HC and breast cancer risk across groups defined by race and ethnicity or examined the interaction of 27HC with lipid or steroid hormone levels. Thus, we conducted a nested case–control study within the Multiethnic Cohort to examine associations of pre-diagnostic, circulating 27HC with breast cancer risk among African American, Japanese American, Latino, Native Hawaiian, and non-Latino White females (1470 cases and 1470 controls). Our primary aim was to assess differences in associations across groups defined by race and ethnicity and to examine interactions of 27HC with lipids and steroid hormones. We also examined associations of 27HC with breast cancer according to hormone receptor (HR) status and use of lipid-lowering drugs. Our secondary aim was to assess differences in associations of lipids and steroid hormones with breast cancer risk across racial and ethnic groups.The Multiethnic Cohort (MEC) is a prospective cohort of 215,251 individuals  45–75 years of age at enrollment between 1993 and 1996 . Particin = 192, 15%) were matched by sequentially relaxing criteria to comprise age at blood draw and birth year ± 3 years (6.6%), time of blood draw ± 4 h and < / = 1 adjacent fasting group (4.8%), age at blood draw and birth year ± 5 years (3.0%), time of blood draw ± 6 years and < / = 2 fasting groups adjacent (0.4%), and finally, time of blood draw ± 8 years and < / = 4 fasting groups adjacent (0.2%). Thus, the final study population consisted of 1470 matched pairs.Participants diagnosed with invasive breast cancer were identified through linkage to the Hawaiʻi and California tumor registries of the Surveillance, Epidemiology, and End Results Program of the National Cancer Institute. Case and control selection are illustrated in Additional file Our main exposure of interest was circulating 27HC. We also assessed circulating lipids , steroid hormones , and sex hormone binding globulin (SHBG).Biomarker assays were conducted at the University of Hawaiʻi Cancer Center Analytical Biochemistry Shared Resource under the supervision of Dr. Adrian Franke. Plasma samples were prepared according to Honda et al. with slight modifications  Additio. 27HC anLaboratory personnel were blinded to case–control status. Matched pairs were assayed in the same batch; quality control pairs were included within each batch, and duplicate pairs were included between batches. All coefficients of variation (CVs) are available in Additional file Covariates included socioeconomic, behavioral, and reproductive factors assessed at baseline or blood collection and consisted of factors that were hypothesized confounders of associations between 27HC and breast cancer risk or previously observed to be associated with breast cancer risk , alcohol consumption , Alternate Mediterranean Diet total score  , smoking status and pack-years , hormone replacement therapy , and lipid-lowering drug use .Covariates collected at the time of blood collection were age at blood draw, body mass index  status was obtained from cancer registries .We calculated the mean, SD, and range of each analyte among control and case populations in the overall study population and across racial and ethnic groups. Distributions of raw values of biomarkers were right skewed; thus, the log-transformed values of biomarkers were used for analysis. We generated frequency distributions of race and ethnicity, HR status, lipid-lowering drug use, and covariates among cases and controls.We used the quasi-Newton method to determine optimal parameters for multivariable conditional logistic regression to estimate odds ratios (ORs) and 95% confidence intervals (CIs) for associations between log-transformed levels of circulating analytes and breast cancer risk . We examP-values < 0.1  involving 27HC, lipid levels, and steroid hormones to breast cancer diagnosis . Thus, the association between 27HC and breast cancer risk was assessed across four quartiles of testosterone based on levels among controls . Only matched pairs with testosterone levels within the same quartile of testosterone were used in this analysis.To assess joint effects between 27HC and other analytes, an interaction term for 27HC and each lipid, steroid hormone, and SHBG were assessed separately in the full model. Using the Wald test, only the model with the interaction term for 27HC and testosterone levels was significant at To assess whether 27HC associations with breast cancer risk were independent of steroid hormones, full models were additionally adjusted for estrone and testosterone. We also assessed whether associations between 27HC and breast cancer diagnosis differed by BMI status .To summarize our results on the association between 27HC and breast cancer risk with those of the mostly White, European EPIC-Heidelberg Cohort , we usedp-values were two-sided; p-values < 0.05 were considered statistically significant. All analyses except for meta-analyses were conducted using SAS version 9.4 . Meta-analyses were conducted with Stata v17 .All ρ = 0.14) and triglycerides (ρ = 0.28). Given that estrone and estradiol were correlated (ρ = 0.72) and over 45% of estradiol observations were below the LLOD, while less than 0.5% of estrone observations were below the LLOD, we restricted further analysis to the estrone biomarker. Frequency distributions of subgroups are presented in Table Levels of 27HC, lipids, steroid hormones, and SHBG among controls and cases are presented in Table p < 0.1 . Results also suggest a weak inverse association of 27HC for HR-negative breast cancer . The p-value for a test of heterogeneity across HR status was p = 0.14.Results from fully adjusted models of the association between 27HC and breast cancer risk are presented in Fig. p = 0.013). Thus, we examined associations between 27HC and breast cancer risk in the overall study population across quartiles of testosterone . We did not observe interactions between 27HC and estrone, SHBG, or lipid biomarkers.We observed a significant interaction between 27HC and testosterone levels (p = 0.02) and for lipid-lowering drug use (p = 0.02).Figure Additional file Estimates of associations between 27HC and breast cancer risk did not change with further adjustment for levels of estrone and testosterone , with a similar but not statistically significant association among non-Latino White females within the MEC and postmenopausal females within EPIC-Heidelberg  and among HR-positive cases within the Multiethnic Cohort and ER-positive postmenopausal cases within EPIC-Heidelberg   . ResultsNotably, our results did not confirm those from animal studies that suggest 27HC would confer increased risk for breast cancer –21. It iWhile most of our estimates of the association between 27HC and breast cancer risk across racial and ethnic groups were not statistically significant, they do suggest differences in the association between 27HC and risk across racial and ethnic groups. Specifically, higher levels of 27HC may be inversely associated with breast cancer risk among Japanese American, Latino, and Native Hawaiian females and positively associated among White and African American postmenopausal females. However, while this suggests that our hypothesis that associations would vary across racial and ethnic groups may be valid, our hypothesis that high 27HC levels would shed light on racial and ethnic disparities in breast cancer risk is not supported by our results. For example, 27HC is inversely associated with risk among Native Hawaiian females who have the highest breast cancer incidence rate as well as Latino females who have the lowest breast cancer incidence rate. Given the complexity of endocrine signaling, it may be that the influence of 27HC on breast cancer development may be modified by other endocrine molecules; our observation of an interaction between 27HC and testosterone in breast cancer risk supports this possibility.Testosterone levels have been linked to higher breast cancer risk –30, 45; A 2015 meta-analysis of prospective cohort studies reported an inverse association between HDL-C and risk among postmenopausal females ; these studies included only White females . Our stuNotably, ours is the first study of 27HC among a multiracial, multiethnic population and thereby sheds light on differences in the mechanism of the impact of cholesterol on breast cancer risk across groups. There has only been one prospective cohort study among African American females that examined total cholesterol and found no association, but that study included females across a wide age range and used self-reported total cholesterol  . On the Our study of breast cancer risk within the Multiethnic Cohort with prospective assessment of 27HC, lipids, and steroid hormones and information on other covariates  offers substantial data to evaluate hypotheses regarding 27HC as a risk factor for breast cancer. However, we note some limitations. Despite being the largest and most diverse cohort population in the US to date to examine 27HC and breast cancer risk, assessing the mechanisms behind associations between 27HC or lipid levels and breast cancer risk may require larger populations, particularly in consideration of groups for which these associations differ  and in consideration of the potential for differences in risk across breast cancer subtypes.Ours is the first US study to examine associations between 27HC and breast cancer risk and suggests a weak inverse association that varied across groups within a multiethnic population. The inverse association of 27HC with breast cancer risk was strongest in females with the highest levels of testosterone. Results from this study extend our understanding of the role of lipids and their metabolites in breast cancer risk as well as racial and ethnic differences in risk of postmenopausal breast cancer.Additional file 1. Supplemental methods, tables, and figures.


Background Interleukin-10 (IL-10) is a cytokine with a vast variety of functions, but its role in cancer development and progression is not yet clear. It is involved in two of the hallmarks of cancer: vascularization and immune modulation. IL-10 inhibits angiogenesis and hence is antitumorigenic. But it also can suppress the immune system and be tumorigenic.Objective Evaluating the role of IL-10 (-1082 A/G) gene promoter single-nucleotide polymorphism (SNP) in breast cancer susceptibility and progression in Georgian women.Methods A case–control study was performed on a total of 128 women, with 64 of them being histologically confirmed to have breast cancer and 64 healthy controls. SNP genotyping was performed with TaqMan assay with real-time polymerase chain reaction. And pathology report, containing proliferative activity and breast cancer hormonal status, was obtained after surgery of the case individuals. Statistical analysis was done to investigate the significance of data obtained from genotyping and histology reports.Results Statistical analysis revealed that the difference in frequency of genotypes was not statistically significant between cases and controls . The comparison of proliferative activity of cases with AA genotypes and AG/GG genotypes showed no statistical difference . Although when put into a plot (box and whiskers), patients with AG/GG genotype have outliers with very high proliferative activity.Conclusion This study shows that -1082 A/G SNP in the promoter region of the IL-10 gene is not associated with breast cancer risk in Georgian women. Breast cancer is the most frequently diagnosed cancer among women and is the second leading cause of cancer mortality in females. In 2019, breast cancer accounted for 30% of estimated new cancer cases in the United States.BRCA1,BRCA2,PTEN,TP53,LKB1/STK11, andCDH1are among high-risk susceptibility genes, whileCHEK2,TGFβ1,CASP8, andATMare among low- to moderate-risk group. Currently, ongoing case–control studies are dedicated to finding more of such genes.Today, 5 to 10% of all breast cancers are caused by known germline mutations. These genes are divided into high-risk and low- to moderate-risk susceptibility groups. Genes likeEven though the human immune system has advanced ways of averting cancer development, neoplastic cells employ multiple strategies to evade immune detection. One such strategy is producing immunosuppressive factors that dampen the function of T cells, dendritic cells, and natural killer cells. Nitric oxide (NO), IL-6, IL-10, tumor growth factor-beta, indoleamine 2,3-dioxygenase, arginase-1, prostaglandin E2, vascular endothelial growth factor, and cyclooxygenase-2 are all examples of such factors that help cancers evade immunity. Among these factors is IL-10 which can impede the function of CD4+ and CD8+ T cells and promote immunosuppressive cells like TregHowever, the role of IL-10 in cancer is not so straightforward. IL-10 expression has also been linked to the downregulation of major histocompatibility complex class I expression on the cell surface. This change enhances the natural killer cells' detection of tumor targets, leading to the lysis of tumor cells.Due to its conflicting immunologic effects, the role of IL-10 in cancer remains uncertain. IL-10 gene is located on chromosome 1 (1q31-1q32), and SNPs in its promoter region, -1082 A/G, can alter its expression and therefore can lead to alterations in cancer susceptibility and cancer progression.In this case–control study, we aim to investigate the involvement of this SNP in breast cancer pathogeneses and its clinical manifestations in the Georgian women population.The study is approved by the Ethics Committee of Tbilisi State Medical University, Tbilisi, Georgia. Signed informed consent was collected from every study participant. The study design is a case–control study. Breast cancer patients who were consecutively admitted to the surgery department in two different oncology hospitals (The First University Clinic and Cancer Research Center) in Tbilisi, Georgia, were recruited from 2017 to 2019. Age-matched women who were regularly involved in breast cancer screening in an outpatient clinic in Tbilisi, and were healthy, were asked to volunteer as control group members. Clinical information of patients was collected from medical notes.The eligibility criteria for the cases were:Biopsy-confirmed diagnosis of breast carcinoma, with no previous history of breast surgery, and no preoperative chemotherapy or radiation therapy.The age range of 30 to 80 years.Ability to understand the purpose of the study and provide informed consent.Female sex.Being ethnically Georgian.Eligibility criteria for controls were as followed: No history of previous cancer diagnosis, no family history of cancer, and minor illnesses were acceptable . The rest of the criteria (2–5) are identical to that of the cases.Due to incomplete data, inappropriate material, or nonmatching diagnosis, 11 patients were later excluded from the study. In total, 64 patients were involved in the study along with 64 healthy controls.Blood samples were collected in a vacutainer tube containing ethylenediaminetetraacetic acid. Genomic deoxyribonucleic acid (DNA) was extracted from the whole blood using DNA purification kits . DNA concentration was measured using the fluorometer-based method .SNP genotyping was performed using the TaqMan assay (Thermo Scientific). Each TaqMan SNP genotyping assay contained sequence-specific forward and reverse primers to amplify the polymorphic sequence of interest and two TaqMan minor groove binder probes with nonfluorescent quenchers: one VIC-labeled probe to detect Allele 1(A) sequence and one FAM-labeled probe to detect Allele 2(G) sequence.Real-time polymerase chain reaction (PCR) was performed based on standard protocols. The final volume of PCR reaction was 25 µL, which contained DNA polymerase, forward and reverse primers with a final concentration of 200 nM, probes with a final concentration of 250 nM, and 50 ng of genomic DNA. PCR conditions for amplification included polymerase activation at 95°C for 10 minutes (hold), denaturation at 95°C for 15 seconds, and annealing/extension at 60°C for 1 minute (cycle 40). The real-time PCR instrument software plot the results of the allelic discrimination (AD) data as a plot of Allele 1 (VIC dye) versus Allele 2 (FAM dye). The AD plot represents each sample well as an individual point on the plot. A typical AD plot shows homozygote clusters, a heterozygote cluster, and no-template controls. The points in each cluster are grouped closely together and each cluster is located well away from the other clusters.The study and control groups were analyzed separately. All statistical analyses were performed by GraphPad Prism 9.3.1 for macOS .p < 0.05.Statistical significance for differences in genotype frequencies was determined by chi-square and Fisher's exact test, and the level of significance was put att-test was used to compare the difference in mean proliferative activity of patients with AA genotype and patients having the G allele (AG and GG).An unpairedTo evaluate associations between the SNPs and the risk of cancer, odds ratios (ORs) and 95% confidence intervals (CIs) were calculated using unconditional logistic regression analysis. All statistical tests are planned to be two-sided.p = 0.7478). Neither IL-10 1082 AA  nor IL-10 1082 AG  were associated with breast cancer. Overall, there was no association between breast cancer risk and the IL-10 -1082 A/G genotype . Although when data are put into a box and whiskers plot, there are obvious outliers in proliferative activity of patients with AG/GG genotype  polymerase 1 (PARP-1). It has been shown that PARP-1 acts as a transcription repressor, with preferential binding for the A-allele than the G-allele. This difference results in less expression in the A allele of this gene.This SNP has been studied in different populations and malignancies, and the results have been conflicting. A study investigated the result of 22 studies in 13 different malignancies regarding the SNPs of the IL-10 gene and found a positive association between IL-10 genotype and disease susceptibility or progression. In some cancers like cutaneous malignant melanoma and prostate cancer, the genotypes that were associated with less expression of IL-10 were a risk factor. However, in other cancers like cervical cancer and hepatocellular cancer, higher expression of IL-10 was a risk factor. The results are especially more conflicting in breast cancer.One study on the Indian populationIn this study, the difference we found in the frequency of genotypes between our cases and controls, who were all Georgian women, was not statistically significant. Our findings are in accordance with the finding of two studies on ChineseWhile these differences might be explained by the dual function of IL-10, as an immunosuppressant (cancer-promoting) and as an antiangiogenic (cancer-inhibiting) agent, it also might be a result of differences in ethnicity.This study also compared the proliferative activity between cases harboring the G allele with those only having the A allele in IL-10 -1082 SNP for the first time. The proliferative activity was measured by Ki-67 which is a monoclonal antibody that recognizes proliferating cells.This study showed no real association between -1082 A/G SNP and breast cancer risk in Georgian women. Much larger studies including different ethnicities and studies combining genotype and gene expression are required to improve our understanding of the role of IL-10 in cancer development, as this current study comes with limitations in size and a lack of full phenotype status . Further understanding of IL-10's role in cancer development can aid us in new cytokine immunotherapies in malignancies.


Background: Triple-negative breast cancer (TNBC) is the most malignant subtype of breast cancer, and studies have found an association between the Myb proto-oncogene like 2 (MYBL2) gene and TNBC development; however, the specific mechanisms underlying development remain unknown. Recent studies have reported the association of alternative splicing (AS) with cancer, providing new approaches to elucidate the carcinogenesis mechanism. This study aimed to identify MYBL2 AS-related genetic variants that influence the risk of developing TNBC, providing new ideas for probing the mechanism of TNBC and novel biomarkers for TNBC prevention.Methods: We conducted a case-control study of 217 patients with TNBC and 401 cancer-free controls. The CancerSplicingQTL database and HSF software were used to screen for MYBL2 AS-related genetic variants. The association of sample genotypes with the risk of TNBC development and with clinicopathological features was analysed via unconditional logistic regression. Combining multiple platforms, the candidate sites were subjected to biological function analysis.Results: Two AS-associated SNPs, rs285170 and rs405660, were identified using bioinformatics analysis. Logistic regression analysis showed that both rs285170  and rs405660  exhibited protective effects against TNBC under the additive model. Stratification analysis showed that these two SNPs had more significant protective effects in the Chinese population aged ≧50 years. Additionally, we found that rs405660 was associated with the risk of lymph node metastasis  in TNBC. Functional analysis revealed that both rs285170 and rs405660 are associated with splicing of exon 3 and that the exon 3-deleted spliceosome does not increase breast cancer risk.Conclusion: We found for the first time that MYBL2 AS-related genetic variants are associated with reduced TNBC susceptibility in the Chinese population, especially in women aged ≧50 years. In recent years, the incidence of breast cancer has been increasing annually; according to the American Cancer Society, the number of new cases of breast cancer worldwide was 2.26 million in 2020, which is the highest number of new cancer cases recorded to date . Breast Genetic susceptibility to breast cancer is increasingly investigated by post-genome-wide association studies, so that current research focuses on identifying functional genetic variants that affect breast tissue carcinogenesis and elucidating the underlying mechanisms via molecular biology experiments. Alternative splicing (AS) is a key molecular event during gene expression, and it involves generation of RNA exons from pre-mRNA transcription binding via selection of different splice sites, resulting in different mRNA splicing isoforms. It is an intricate mechanism of coordinated transcriptional regulation that enables a single gene to generate different mRNAs, thereby greatly increasing the diversity of the transcriptome and proteome. This phenomenon is present in almost all genes encoding proteins. AS occurs widely in human tissues; however, abnormal splicing events may lead to the development of various neurological diseases and cancers . At leasMYBL2) gene, a member of the MYB family of transcription factors and one of the key genes regulating cell cycle progression, is expressed in almost all proliferating cells and regulates cell survival and differentiation, while also mediating aberrant cell progression in cancer  gene, and in-vitro assays showed that variants B and C are associated with invasiveness in breast cancer cell lines. A causal relationship between breast cancer risk-associated splicing events and SNPs has been confirmed in minigene experiments  analysis between SNPs was performed using the Haploview 4.2 software. The potential AS function of these SNPs was further predicted in the HSF online software, and the minor allele frequency (MAF) of these SNPs in the southern Chinese population was detected using the 1000 Genomes Project. SNPs with an MAF >0.1 were screened, and redundant SNPs were removed according to HSF score and LD analysis.All potential splicing-causing SNPs of This study recruited 217 women with TNBC and 401 cancer-free women (controls) at the Affiliated Hospital of Zunyi Medical University from 2018 to 2021. All cases were histopathologically and immunohistochemically confirmed for TNBC; the immunohistochemical results met ER (-), PR (-), and HER-2 (-), and the patients had no previous history of other malignancies. The control group comprised cancer-free women attending the physical examination department of this hospital during the same period according to the table of random digit, and they were matched with the case group based on age (±5 years). Briefly, 2 ml of peripheral blood was drawn for subsequent testing. General information and pathological data of the study participants were obtained by querying medical records and conducting interviews. Data entry was completed using the EpiData 3.1 software. All data were entered in duplicate, and then, inter-input comparisons were performed to ensure the accuracy of data entry. Participants who had never smoked or smoked less than one cigarette per day were defined as “never smoked” and all other cases were defined as “smoked.” Alcohol consumption twice or more per week for at least one year was defined as “drinking” and other cases were considered as “no drinking.” The ethics committee of the Affiliated Hospital of Zunyi Medical University approved the study (No. KLLY-2021-200). All participants provided written informed consent.The collected peripheral blood samples were used to extract genomic DNA using the BloodZol Kit . The concentration of the DNA solution was diluted to 50 ng/µL and stored at −20°C. Sample DNA was genotyped using the CFX96 real-time PCR detection system  and TaqMan probes . Duplicate experiments were performed using 5% of samples randomly selected from cases and controls; the results of the two experiments were compared for consistency.MYBL2 splice isoforms among different tissues were analysed using the TSVdb platform (http://tsvdb.com/index.html) to determine whether the splice isoforms are associated with breast cancer risk.We used the CancerSplicingQTL database to analyse candidate sites, the exon in which they were predicted to act, the splicing pattern, and the splice site, to determine what spliceosome the candidate site might affect. The expression differences in 0: Exposure rates of risk factor in controls; p1: Exposure rates of risk factors in the cases; r: Ratio of control sample size to case sample sizeCalculation of power was performed using the following formula:p < 0.05 were considered statistically different. All data were analysed using the SPSS 26.0 software .It was calculated that 200 cases and 400 controls would provide 91% test power when OR = 2, α = 0.05, and MAF = 0.1. Chi-square tests and two independent-sample t-tests were used to compare whether demographic statistical characteristics were significantly different between cases and controls. The chi-square goodness-of-fit test was used to test whether the genotype distribution in controls was consistent with the Hardy–Weinberg equilibrium, and a logistic regression model was used to calculate the odds ratio (OR) and 95% confidence interval (CI). Values of MYBL2 AS in BC, and the specific screening pipeline was as described below.Through bioinformatics analysis , we obtaMYBL2 gene, which finally resulted in 199 breast cancer associated SNPs potentially affecting AS in MYBL2 in CancerSplicingQTL , and input into Haploview. The r2 values among loci  between cases and controls. After stratification based on age, there was still no significant difference in age between cases and controls. There were also no significant differences in smoking and alcohol consumption between cases and controls. The TNM staging characteristics are described in A total of 217 TNBC cases and 401 cancer-free controls were included in our case-control study, with a mean age of 50.41 years (±10.93 years) in the case group and 50.28 years (±11.57 years) in the control group. There was no significant difference in age (Typing experiments were performed in which 5% of the samples were randomly selected for repeated experiments with an anastomosis rate of 100%. The distribution of rs285170 typing results of cases and controls is shown in p = 0.007) and additive model  both revealed that the C allele was a protective factor for TNBC; rs285170 (T > C) was a protective mutation for TNBC. Furthermore, we found that the rs405660GG genotype was significantly associated with a decreased risk of TNBC . The G allele exhibited protective effects against TNBC in the population under the dominant model  and additive model   .p = 0.001), while the dominant  and additive  models similarly showed significant differences.An age-stratified analysis was performed based on whether participants were aged ≧50 years. Notably, rs285170 was not significantly associated with TNBC susceptibility in participants aged <50 years; however, it showed a greater and significant difference in participants aged ≧50 years as compared to the whole population. In participants aged ≧50 years, the TC genotype was significantly associated with a decreased risk of TNBC  and additive  models  models .p = 0.005) and additive  models  in TNBC ; however) models . HoweverMYBL2 AS. The splice site was chr20:42310424, and both affected MYBL2 exon 3 skipping; that is, generation of a spliceosome with exon 3 skipping (MYBL2 splice isoforms in breast cancer was next analysed on the TSVdb platform. In this database, we performed an analysis of the exonic expression landscape of MYBL2 in breast cancer, with the final output shown in MYBL2 and the 14 exons on MYBL2 in normal, orthotopic, and metastatic breast cancer tissue, and the difference between the two splice isoforms is shown in Functional prediction of our candidate SNPs in CancerSplicingQTL indicated that both rs285170 and rs405660 were associated with skipping . The expBRCA1/2 germline mutations are associated with approximately 15%–20% of TNBC cases; nevertheless, the susceptibility genes and mutation sites have only been specified in few patients with TNBC (TNBC is a highly proliferative subtype of breast cancer that is often associated with poor prognosis. The treatment strategy of breast cancer varies based on molecular typing; however, as TNBC lacks expression of ER, PR, and HER-2, it has low hormone sensitivity, and the prognostic markers and therapeutic targets remain unknown. Compared with other invasive breast cancers, TNBC has an earlier onset and a higher recurrence rate, and it is also more likely to spread and metastasise . TNBC isith TNBC .MYBL2 is an important regulatory gene in cell cycle progression and it affects cell proliferation and differentiation. Several studies have found an association of MYBL2 with malignant tumour development, such as breast, bladder, lung adenocarcinoma, and ovarian cancer (MYBL2 S427G variant has been reported to be associated with an increased risk of basal-like breast cancer in the African American population; however, the specific mechanism has not been clarified (MYBL2 in breast cancer development and found that miR-143-3p targets the MYBL2 3′-untranslated region, which inhibits MYBL2 expression and promotes proliferation and inhibits apoptosis of breast cancer cells. A MYBL2 splice variant lacking exon 9A has been found to function as a suppressor protein and counteract transactivation mediated by the whole exon-expressed MYBL2 (MYBL2 and breast cancer risk (MYBL2. These two SNPs are likely to influence the risk of TNBC by affecting AS of MYBL2. To our knowledge, this is the first study conducted worldwide on these two SNPs and the first systematic analysis of the association of MYBL2 AS-related genetic variants with AS risk.n cancer . The MYBlarified . Chen aned MYBL2 . Althouged MYBL2 . Previoucer risk . TherefoMYBL2 expression in endometrial cancer tissues, which supports a relationship between the increased risk of endometrial cancer and MYBL2 expression levels in premenopausal women (MYBL2 expression; however, ER, PR, and HER-2 are not expressed in TNBC cases. Therefore, to explain whether MYBL2 expression in TNBC is associated with hormone level changes, studies should confirm the existence of associations between hormone level changes and MYBL2 expression. In addition, unlike rs285170, where the homozygous rs405660 variant showed significant protection in the full-age analysis, the association of the homozygous rs285170 variant with TNBC risk was not significant. This may be explained by the low MAF of rs285170 with few homozygous mutant genotypes in our sample. Follow-up studies using larger sample sizes may obtain significant results. In addition, we found that rs405660 was significantly associated with a decreased risk of lymph node metastasis in TNBC, and this finding may provide a reference for subsequent clinicopathological studies on MYBL2 in association with TNBC.We found that the C allele of rs285170 was associated with decreased susceptibility to TNBC in 217 of our cases, and the same results were confirmed in dominant and additive models. However, no significant difference was found in participants aged <50 years, and when this group of the cohort was excluded, the difference was more pronounced in the group aged ≧50 years. The G allele of rs405660 also exhibited a similar protective effect. A certain characteristic of the population aged ≧50 years is expected to play a role in this protective effect. Combined with the age characteristic analysis, we speculate that a joint effect relationship exists between menopausal status and mutations at this locus. After menopause, women experience certain changes in levels of hormones such as oestrogen and progesterone, and menopausal status is often found to be a risk factor for breast cancer . ERβ expal women . CurrentMYBL2 exon 3 skipping. Whole exon spliceosomes significantly increase breast cancer susceptibility, but the exon 3-deleted spliceosome was not significantly different between breast cancer tissues and adjacent non-cancerous tissues. It reduced susceptibility to breast cancer compared with the whole exon spliceosome. This is consistent with the population association analysis results that MYBL2 AS-related genetic variants can reduce the risk of breast cancer.On the basis of population association analyses, we performed biological functional analyses based on TCGA data. The results showed that both rs285170 and rs405660 were associated with MYBL2, and determined linkage loci in the SNPs most likely to exhibit AS function. Our findings may provide a reference for AS-related research. Our study had a few limitations. First, our cases were all recruited as inpatients in hospitals, and there may be a certain Berkson’s bias. Secondly, since the AS-related SNPs, splicing exons, splicing sites, and splicing patterns obtained in this study were all predicted by bioinformatics, we recommend that subsequent studies supplement biological functional verification, such as analysis of SNP association with MYBL2 splice isoform expression levels and minigene construct or subcutaneous tumour bearing experiments in nude mice.In this study, we proposed a novel bioinformatic analysis strategy to search for genetic variants associated with AS, systematically analysed all SNPs potentially causing AS in MYBL2 AS-related genetic variants in TNBC susceptibility in the Chinese population using bioinformatics analysis. We found an association between rs285170 and rs405660 and the development of TNBC. The SNPs rs285170 and rs405660 may produce different splicing isoforms by altering the transcription of MYBL2, which in turn affects the risk of TNBC. In addition, we found that rs405660 was associated with the risk of lymph node metastasis in TNBC. Our findings elucidate the mechanisms underlying TNBC development and cancer in general.To the best of our knowledge, this is the first study to systematically investigate the role of


Biomarkers for early detection of breast cancer may complement population screening approaches to enable earlier and more precise treatment. The blood proteome is an important source for biomarker discovery but so far, few proteins have been identified with breast cancer risk. Here, we measure 2929 unique proteins in plasma from 598 women selected from the Karolinska Mammography Project to explore the association between protein levels, clinical characteristics, and gene variants, and to identify proteins with a causal role in breast cancer. We present 812 cis-acting protein quantitative trait loci for 737 proteins which are used as instruments in Mendelian randomisation analyses of breast cancer risk. Of those, we present five proteins  that show a potential causal role in breast cancer risk with confirmatory results in independent cohorts. Our study suggests that these proteins should be further explored as biomarkers and potential drug targets in breast cancer. Proteomics of blood samples is a promising avenue for cancer diagnosis. Here, the authors conduct Mendelian randomisation analysis of protein levels across multiple cohorts, and identify 5 proteins that show promise as biomarkers for the long-term risk of breast cancer, and as potential drug targets. Genome-wide and exome-wide genetic association studies have successfully identified over 300 breast cancer susceptibility loci4 but the mechanisms underpinning most loci and specific gene variants remain uncharacterised, which limits the translation of genetic susceptibility loci to new therapies and precision medicine tools4.Breast cancer is globally the most common cancer in women and is associated with significant morbidity and mortality5. In comparison to genome-wide association studies (GWAS), MR exploits a more confined test space, which increases statistical power, and inherently supports causal gene identification. MR can be further supported by genetic colocalization analysis of exposure and outcome6. The relevance of MR has been evaluated and supported by retrospective analyses of drug targets with a proven aetiological or causal role in disease from randomised controlled trials (RCT)8.Mendelian randomisation (MR) offers an alternative approach to the mapping and understanding of aetiologically important pathways in cancer risk and development. MR aims to elucidate causal relationships between modifiable risk factors and disease based on the analysis of genetic variants in observational data9. Second, circulating proteins can be measured with high throughput and precision using a variety of advanced methods11. Third, recent studies have shown that a majority of circulating proteins are associated with cis-acting protein quantitative trait loci (pQTL) i.e., located within 1 Mbp from the protein-encoding gene13. Fourth, individual cis-pQTL explains relatively large proportions of variance in the protein, making them statistically powerful instrumental variables for causal inference using MR14. Hundreds of pQTL for plasma proteins have been identified, but so far, no studies have reported pQTL in an entirely female population19.Circulating proteins possess many of the characteristics suitable for the discovery of breast cancer biology using MR. First, the plasma proteome has been shown to reflect both normal physiology and pathogenic biological processes in cancer20 and the UK Biobank21. Lastly, we followed up on significant proteins identified in the MR analysis by visualising and evaluating colocalization of the protein and breast cancer genetic associations and evaluated potential causal relationships with established and emerging breast cancer risk factors, also using MR  using the Olink PEA Explore assay in plasma samples taken from 598 women who were free of a breast cancer diagnosis at the time of sampling. We 1) performed a genetic association analysis of protein levels to identify cis-pQTL and (2) used the cis-pQTL as instrumental variables in MR analysis of breast cancer in the Breast Cancer Association Consortium (BCAC) case-control meta-analysis of breast cancer risk, and (3), replicated MR findings in a second breast cancer case-control meta-analysis of FinnGen performeOut of 730 plasma proteins evaluated using MR, genetically elevated levels of five proteins were associated with breast cancer risk, namely CD160, 2’-deoxynucleoside 5’-phosphate N-hydrolase 1 (DNPH1), layilin (LAYN), Leucine rich repeat containing 37 member A2 (LRRC37A2) and toll-like receptor 1 (TLR1), which were confirmed in an independent set of data. Our results suggest that these five proteins are aetiologically relevant for breast cancer development. Pending further validation, these findings may point to drug target opportunities or stratification biomarkers in breast cancer.22. The sample for the present substudy was selected for the purpose of evaluating plasma protein biomarkers in relation to incident breast cancer within 2 years from blood sampling. The selection included samples from 299 women in the Southern Sweden (Skåne) region who received a breast cancer diagnosis within 2 years after a blood draw and 299 random controls from the same region, who, as of 2021, had remained breast cancer-free. No difference between cases and controls was seen for median age, body mass index or percent women receiving hormone replacement therapy at the time of blood draw. The proportion of smokers and women with a family history of breast cancer was more common among cases . The aim of the project is the identification of risk factors for breast cancer23.We chose to analyse the plasma samples using an affinity proteomics approach. While targeted methods, such as the Olink PEA approach, are inherently biased towards the subset of proteins that are measured, we attempted to maximise the possibility for discovery by measuring as many proteins as possible. Hence, we used the recently launched version of Olink’s Explore I and II panels, which includes 2949 proteins  in post-menopausal women and higher placental alkaline phosphatase (PLAP) levels in smokers25. Some less described correlations included lower plasma levels of glycodelin (PAEP) and chordin like 2 (CHRDL2) and higher levels of glycoprotein hormone alpha polypeptide (CGA) in post- and peri-menopausal women, and lower levels of osteomodulin (OMD) in women using (HRT).To examine observational relationships between protein levels and clinical characteristics of the KARMA women, we regressed each measured protein against seven factors , hormone replacement therapy (HRT), peri- and post-menopause and current smoking). All associations are shown in Supplementary Data The replication of known trait-to-protein associations suggests that the data quality was satisfactory and that additional trait-to-protein associations are enabled by the expansion of the number of detectable proteins.p < 2.77 × 10−4, Supplementary Fig. 2 < 0.1) across 737 proteins, henceforth referred to as cis-pQTL (Supplementary Data n = 523) but several pQTL were also observed for Explore II proteins (n = 289). Some of the cis-pQTL showed effect sizes well above 1 standard deviation, including the nucleotidase NT5C , acylphosphatase (ACYP1)  and carboxypeptidase Q (CPQ) . We conclude that pQTL are readily detected for proteins on both Explore I and II panels, providing potential MR instruments for 737 proteins.To identify genetic instruments for the downstream causality testing using MR, gene variants within a range of 1 Mbp up and downstream of genes encoding each of the 2929 unique proteins were tested for association with levels of the corresponding protein. Significant associations  (>0.8), were looked up in previously published studies reporting cis-pQTL based on the Somascan proteomics platform5. We did not identify genetic proxies for seven of the proteins with cis-pQTL in KARMA, resulting in the testing of 730 protein exposures. Of those, seven proteins surpassed the statistical threshold for significance (p < 7.5 × 10−5) in the discovery study -positive compared to ER-negative breast cancer Table . However28. LRRC37A2 showed MR evidence for later age at menarche and earlier age at menopause in two independent outcome datasets, and also for higher IGF-1 levels To summarise, the MR analysis showed that genetic elevation of CD160, DNPH1, LAYN, LRRC37A2 and TLR1 associated with breast cancer risk, and with similar effects on ER+ and ER− cancers.p-values for breast cancer risk but were localised in the same, size-limited, genomic region. We considered the cis-pQTL to be colocalised with breast cancer risk  and exhausted CD8+ T cells for several cancer types including breast cancer43. In our study, the genetic elevation of LAYN protein levels is associated with increased breast cancer risk, suggesting a LAYN inhibitor would be desired for the treatment of breast cancer. However, mechanistic studies will be required to confirm the direction of effect proposed by the MR evidence and to validate LAYN as a drug target in breast cancer.Our search for drug targets highlighted the connection between LAYN and Hyaluronic Acid. LAYN encodes Layilin, which is a talin-binding transmembrane and integral membrane protein functioning as a receptor for Hyaluronic acid (HA), with a role in cell adhesion and motility44. Zhu et al. demonstrated an absence of association with breast cancer for genetically elevated levels of C-reactive protein45 and Shu et al. reported a wider MR analysis, instrumenting 1469 proteins using Somascan-based pQTL in the INTERVAL cohort, of which genetic instruments for 26 proteins were found to be associated46. Bouras et al. instrumented 47 inflammatory cytokines and reported that genetically increased levels of CXCL1 and decreased levels of MIF associated with breast cancer47. Of the 28 proteins previously reported in breast cancer MR studies, our study included post-QC data on 22 proteins and a cis-pQTL was identified in our study for five of them . We replicated cis-pQTL reported for RELT, ENG and SCG3 in ref. 26, which were based on the Somascan protein assay, but were unable to replicate pQTL reported for TFPI, ISLR2  were associated with a weak but significantly increased risk of breast cancer, whereas IGF-binding protein-3 was unassociatedOur study has both strengths and limitations. One of the strengths is the large number of proteins tested for cis-pQTL and that the cis-pQTL used to instrument genetic elevation using MR were identified in women only, which should provide better estimates in MR for female breast cancer. Another strength is that the protein exposures meeting statistical significance in our discovery MR, using data from the BCAC consortium as outcome, were replicated in the independent case-control analysis that combined breast cancer cases and controls in FinnGen and the UK-Biobank.p-value threshold for significance for the number of independent variants in each cis-region. Effect sizes observed in KARMA were highly concordant with an overlapping set of 33 cis-pQTL for proteins measured with Olink PEA that were previously reported in a study several times larger than the present study. To evaluate the robustness of cis-pQTL identified in KARMA, we sought replication for an overlapping set of 569 proteins measured with Somascan. Of those, 2/3 (374/569) were replicated, which is on par with the expected replication rate given differences in protein analysis methods16.However, our study had a limited sample size for discovering cis-pQTL with smaller effect sizes. Therefore, we cannot exclude that additional proteins on the Olink Explore II panels harbour significant cis-pQTL but remained undetected in the KARMA sample. In addition, several of the cis-pQTL with very large effect sizes such as ENTPD6 and NT5C, have a minor allele frequency of less than 3% and their effect sizes may well be inflated because of the so-called “winner’s curse. To decrease the false-negative error rate we only included variants in cis to decrease the multiple-test burden and corrected the In conclusion, by applying an MR approach to a broad range of circulating proteins we found that genetically elevated CD160, DNPH1, LAYN, LRRC37A2 and TLR1 were associated with breast cancer. This suggests that these five proteins play an aetiological or causal role in breast cancer, providing a basis for further functional evaluation of their potential as drug targets.https://etikprovningsmyndigheten.se/en/22. From KARMA we identified 299 women diagnosed with breast cancer, which occurred within 2 years of blood draw, and who were residents in Southern Sweden. We used the matchit function from the MatchIt library implemented in R to match 299 controls from the KARMA study to the incident cases by randomly drawing women without incident breast cancer so that the median age at blood draw in cases and controls were similar (median matching). Blood samples were collected at baseline. All blood samples were handled in accordance with a strict 30-h cold-chain protocol, which required that all blood samples were transported on ice and were processed and aliquoted within 30 h from the draw. The sample collection included 16 plasma aliquots, one aliquot of extracted DNA and two aliquots of whole blood for backup. In total, EDTA blood samples from 69,440  study participants were collected.The KARMA cohort consists of 70,877 women performing a screening or clinical mammogram at four hospitals in Sweden during the period October 2010–March 2013. Women consented to both risk and prognosis of breast cancer including collection and storage of questionnaire data, mammograms, matched health care register data, and biological samples. The study was approved by the Stockholm Ethical Review Board, Plasma proteomics was performed in samples from the 299 BC cases and 299 BC free controls from KARMA, using the Olink Explore I and II panels  according to the manufacturer’s protocol. Explore combines the Proximity Extension Assay (PEA) technology with Next-generation sequencing (NGS).In brief, the PEA technology uses matching pairs of oligonucleotide-labelled antibody probes. The PEA probes bind to target antigens producing a binding complex where the complimentary oligonucleotides exist in close proximity to each other, enabling the formation of a target sequence. The dual targeting of probes has been proven to produce outstanding specificity enabling a high degree of multiplexing while maintaining sensitivity and a broad dynamic range. In the Olink Explore protocol, the target sequence is amplified in a double PCR reaction and purified before the NGS. The sequence data is processed and normalised to produce Olinks relative quantification unit Normalised Protein eXpression (NPX). The produced DNA signal functionally works as a proxy for the protein levels present in the sample. Further details on the Olink Explore protocol and internal quality control are available in the Supplementary Methods The Olink QC-system includes negative controls, used to monitor the background noise and to set the limit of detection (LOD). Supplementary Fig. A principal component analysis of all data was performed to detect outliers and to inform potential sample exclusions. Of the 598 samples that were included in the analysis, one sample was excluded entirely, two samples were excluded from the analysis of Explore ONC-II and two samples were excluded from the analysis of the Explore INF-II panel data . The average number of variants per cis-region was 6249 . A false-discovery rate (FDR) at 5% provided a similar estimate (p < 5.54E-04).Proteins >75% of NPX values below LOD were filtered out before the pQTL analysis, yielding a total of 2476 proteins in the analysis. Values below LOD were included. The pQTL discovery analysis was performed using an additive model with adjustments for age, BMI and 10 genetic PCs in PLINK 2.0. To preserve statistical power for pQTL identification, only variants within a 1 mega-base pair window of the protein-coding gene were tested for association with respective circulating protein levels. To manage multiple test correction, while limiting false negatives, the total number of variants per cis-region were calculated as well as the number of independent variants (R49 (https://mrcieu.github.io/TwoSampleMR/) to test for proteins with a potential causal role in breast cancer. Independent cis-pQTL (r2 < 0.001) were used as instrumental variables (IV), and GWAS of breast cancer risk from the BCAC consortium were used as outcome, which included data from 122,977 breast cancer cases and 105,974 controls. In the case of a single independent IV Wald Ratio was applied, otherwise, inverse‐variance weighted estimates were reported. The threshold for statistical significance was defined as (7.5 × 10−5) to account for multiple testing. The replication analysis was performed in a meta-analysis of FinnGen R9 and the UK-biobank, which included 25,807 cases and 355,307 controls. Only the seven proteins that met statistical significance in the BCAC discovery analysis were included in the replication analysis, and hence a nominal p-value of 0.05 was considered statistically significant.We performed Two-sample MR using the R package TwoSampleMRFurther information on research design is available in the Supplementary InformationPeer Review FileDescription of Additional Supplementary Files DocumentSupplementary Data 1-6Reporting Summary