Triple-negative breast cancer (TNBC) is usually the most malignant and aggressive mammary epithelial tumor characterized by the lack of expression for estrogen receptors and progesterone receptors, and the absence of epidermal growth factor receptor (HER)2 amplification. Corresponding to 15–20% of all breast cancers and well-known by its poor clinical outcome, this negative receptor expression deprives TNBC from targeted therapy and makes its management therapeutically challenging. Type 2 diabetes mellitus (T2DM) is the most common ageing metabolic disorder due to insulin deficiency or resistance resulting in hyperglycemia, hyperinsulinemia, and hyperlipidemia. Due to metabolic and hormonal imbalances, there are many interplays between both chronic disorders leading to increased risk of breast cancer, especially TNBC, diagnosed in T2DM patients. The purpose of this review is to provide up-to-date information related to epidemiology and clinicopathological features, risk factors, diagnosis, biomarkers, and current therapy/clinical trials for TNBC patients with T2DM compared to non-diabetic counterparts. Thus, in-depth investigation of the diabetic complications on TNBC onset, development, and progression and the discovery of biomarkers would improve TNBC management through early diagnosis, tailoring therapy for a better outcome of T2DM patients diagnosed with TNBC. Diabetes is a serious metabolic disorder characterized by sustained hyperglycemia, hyperinsulinemia, and hyperlipidemia due to insulin resistance, which potentially enhances the risk of cancer, and especially triple-negative breast cancer (TNBC) in women . DiabetePatients with T2DM have an increased risk of developing TNBC, a heterogeneous group of breast carcinomas with most of them characterized by a fast-growing and aggressive subtype with different genetic profiles. Some rare TNBC cases are low-grade and poorly invasive with an indolent clinical outcome; however, TNBC is frequently diagnosed at a later stage, associated with a larger tumor size, a higher tumor grade, poorly differentiated invasive ductal carcinoma histological subtype, increased rate of lymph metastasis, and associated with a poor 5-year survival ,22. TNBC−) and progesterone receptor (PR-negative or PR−) expression and the absence of human epidermal growth factor receptor 2  overexpression detected by immunohistochemistry (IHC) staining. However, recently, a negligible percentage corresponding to less than 1% of ER and PR expression has been detected [TNBC, a highly invasive heterogeneous subtype with the poorest outcome, is clinically and molecularly defined by the lack of estrogen receptor , and cyclin-dependent kinase inhibitor 2A (CDKN2A) gene mutations are often found in BL1, BL2, and ML subtypes while the LAR TNBC subtype harbors TP53, PTEN, and phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA) gene mutations. Considering TNBC clinical behavior, prognosis, and multidrug resistance, a PTEN-reduced/PI3K-high/mammalian target of rapamycin (mTOR)-high expression is suggested to constitute a high-risk profile of TNBC progression [HRAS, KRAS, and platelet-derived growth factor receptor-alpha (PDGFRA). The two other TNBC subtypes contain genomic alterations resulting in mutations in immune-related genes such as nuclear factor kappa-B inhibitor alpha (NFKBIA) and tumor suppressor APC [The main genetic mutations, reported to be strongly associated with TNBC development, are breast cancer genes. Both subtypes . The tumgression . The metssor APC . Added tAlthough the specific etiology of TNBC has not been described, many predisposing risk factors such as gender; age; genetic mutations and family history; reproductive history; ethnicity; and precipitating risk factors, including body mass index (BMI), physical inactivity, sex hormonal imbalances, vitamin supplements, alcohol intake and smoking, and metabolic syndrome, have led to TNBC occurrence . While TThe main predisposing risk factor for all types of breast cancer including TNBC is the gender as women are more predisposed to develop breast cancer than men, who represent only 0.02 percent of all breast cancer cases . BRCA1/2The age factor is a crucial parameter in TNBC onset and development as TNBC risk increases with the age. The median age of women presenting TNBC is around 60, much younger than those diagnosed with other breast cancer subtypes ,45. The FTO), the interleukin-6 (IL-6) gene, and heat-shock protein 60 encoded by HSPD1 gene [FTO rs3751812, IL-6 rs1800796, and HSPD1 rs2605039. It was revealed that T2DM individuals with CC genotype of IL-6 rs1800796 or GG genotype of HSPD1 rs2605039 were strongly associated with increased risk of breast cancer (OR (95%CI): 2.53  and OR (95%CI): 6.40 , respectively) compared to non-T2DM individuals [HSPD1 rs2605039, suggesting HSPD1 influence on breast cancer survival [Several genetic-based studies, especially large-scale, genome-wide association studies (GWAS), have been conducted to understand the etiology of complex diseases such as breast cancer and T2DM. Large-scale GWAS of T2DM and breast cancer have provided evidence for shared susceptibility genes, including a fat mass and obesity-associated gene  associated with first primary invasive breast cancer risk in 817 Caucasian women while other SNPs  were strongly associated with breast cancer-specific mortality [SLC30A8 gene encodes zinc transporter ZnT-8 and its variants were correlated with improved glucose tolerance, protection from T2DM development, and reduction of T2DM risk [IRS2 gene, observed in 49% of all TNBC patients [CDKN2A/B (FDR = 5.6 × 10−5 and 7.5 × 10−5) genes than other subtypes, providing the basis for future genomic-driven targeted therapies [CDKAL1, mainly reported as a susceptibility gene for T2DM through sustained systemic glucose homeostasis and adipose mitochondrial function regulation, has been linked to luminal-AR TNBC subtype [TCF2/HNF1B involved in β-cell development and function, and is reported to be negatively associated with overall survival in breast cancer  [Additional genetic polymorphisms-related studies reported more diabetes-related risk SNPs  . The ideBRCA1/2, is the major risk factor associated with breast cancer and corresponds to 5–10% of all breast cancers [+ subtype, and ER−/PR−/HER2+ breast cancer was the most pronounced compared to women with no affected degree relatives [BRCA1 or BRCA2 mutation, half of them developed breast cancer  and faced a twofold increase risk of diabetes in the 15 years after breast cancer diagnosis [IGF1R, main key players in the pathogenesis of T2DM-related complications and in breast cancer onset and progression [Hereditary breast cancer, particularly the susceptible germline mutation  cancers ,66. Seveelatives . Analyzielatives . There ielatives . After riagnosis . T2DM deiagnosis . Primariiagnosis . In addigression ,70.TNBC etiology is also due to reproductive lifestyle such as early age at menarche, late menopause and remaining nulliparous , late age at first completed pregnancy, and lack of breastfeeding, increasing by twofold the risk of TNBC ,72. A re− and TNBC was reported between European-American or non-Hispanic white women and African American women, indicating a 20% increased risk of developing breast cancer in T2DM White women while increased risks of 43% and 92% of ER− development in T2DM African American women and their non-obese counterparts, respectively [Population-based analyses estimated that the annual average of TNBC incidence corresponds to 15 in 100,000 women, especially non-white premenopausal (less than 40 years) individuals developing TNBC . Recent ectively .Breast density is often a hereditary trait, which is measured by mammography representing the volume of fibroglandular (dense) tissue relative to fatty (non-dense) tissue. Breast density can also increase by using menopausal hormonal therapy and having a low body mass index ,91,92. TSeveral observational epidemiological studies based on measurements of breast density provided evidence that the association between T2DM and mammography density is differential depending on the type of diabetes treatment . Another2 while underweight, overweight, and obese subjects present BMI value lower than 18.5 kg/m2, in the range of 25–29.9 kg/m2, and greater than 30 kg/m2, respectively. However, it has been well documented that obesity is associated with decreased risk of premenopausal breast cancer and increased risk of postmenopausal breast cancer [+ breast cancer and a reduced risk of TNBC [BMI is a common index derived from the weight and height of the individual, estimating the amount of body fat and indicating the healthy weight status of the person. Three categories have been classified based on BMI value such as healthy weight corresponding to an average of 23 kg/mt cancer ,98. Due  of TNBC ,101,102.Based on epidemiological and observational studies conducted in large cohorts, recreational physical activities are one of the most modifiable factors to be encouraged to perform for reduction or even prevention of the risk of breast cancer, especially TNBC . PreviouTNBC etiology may be also due to hormonal imbalances caused by oral contraceptives or menopausal replacement therapy. The existence of a strong link between hormonal imbalances and the risk of breast cancer development in women has been well defined unlike in men, as breast cancer like ovarian cancer is known as female hormone-responsive cancer. The long exposure of ovarian hormones, endogenous estrogen, and progesterone may cause breast cancer onset. Pre-clinical studies and observations suggested that estrogen might promote the development of ERα-negative breast cancer subtypes, including TNBC . Althoug3—also known as cholecalciferol, presents protective effects through its interactions with several proteins involved in cell growth arrest, DNA damage, and in the inactivation of cathepsin L-mediated degradation of p53 binding protein 1 (TP53BP1) [Most of the vitamins are well known for their anticancer properties that can impede TNBC development and progression. In vitro studies have reported the anticancer effects of vitamin C and vitamin B3-derived nicotinamide on TNBC development and progression by decreasing pro-angiogenic hypoxia-inducible factor HIF-1α and increasing pro-apoptotic reactive oxygen species levels, respectively ,117. TheTP53BP1) . VitaminTP53BP1) ,120,121.TP53BP1) . IncreasTP53BP1) . Based oTP53BP1) ,124. In + breast cancer, their association with TNBC risk is more controverted [Lifestyle, including alcohol intake and smoking, has a major impact on promoting breast cancer malignancy through the overproduction of estrogens, alterations in estrogen receptors, oxidative stress, DNA damage, genotoxicity, epithelial–mesenchymal transition promotion, and over-activation of epidermal growth factor receptor (EGFR)/ErbB2 signaling ,126. Altroverted ,128,129.roverted . In controverted . In addiroverted .Metabolic syndrome, a cluster of biological irregularities resulting in hypertension, dyslipidemia, obesity, insulin resistance, and subsequently diabetes, was reported to be a robust risk factor for TNBC in a case–control study of 555 West African women . A retro+ breast cancer but an increased risk of developing TNBC [T2DM, essentially hyperglycemia and known as a common comorbidity of obesity, is considered an independent prognostic risk factor leading to an increased risk for TNBC onset, development, recurrence, and metastasis . Many eping TNBC . Due to ing TNBC .Glycemic control based on glycosylated hemoglobin (Hgb) A1c measurements indicate the three categories of glycemic control: well controlled (HgbA1c < 7.0), moderately controlled (HgbA1c 7.0–9.4), or poorly controlled (HgbA1c ≥ 9.5). TNBC incidence increases with worsening glycemic control in T2DM patients .There is also an urgent need to identify potential novel circulating plasma biomarkers, including micro-RNA (miRNA) and long non-coding LncRNA, circulating tumor DNA (ctDNA), metabolites, and circulating tumor cells and exosomes as non-invasive screening for TNBC diagnosis in T2DM patients.The cell-free miRNAs, a large family of small 20–22 nucleotides, and LncRNAs, longer than 200 nucleotides, are both considered non-coding RNAs, which play a pivotal regulatory role at the post-transcriptional level leading to either mRNA degradation or translational suppression of more than 30% of genes in the human during development ,137. RecBlood-based circulating tumor DNA (ctDNA) and cell-free nucleic acids have been known as “liquid biopsies”, referring to alternative biomarkers in the biofluids, which avoid the invasive procedure of tissue biopsies ,148,149.The exosomes are nanosized lipid bilayer extracellular vesicles, which serve as a cargo containing a complex of mixtures including various types of nuclei acids  and signaling proteins that are released after binding to recipient cells. An increasing body of evidence revealed the multiple exosome biological functions in TNBC tumorigenesis processes, including tumor initiation, metastasis, drug resistance, and immune escape . In addiThe metabolomics can provide clinically valuable information about metabolic alterations, including metabolic reprogramming, in a cancer state requiring large supplies of macro-nutrients and can lead to new insights into the pathology of TNBC and the discovery of potential therapeutic targets aiding in the development of effective therapies . Recentl+/HER2− breast cancer [The circulating tumor cells (CTCs) represent another feature of biomarkers as these tumor cells have been detached from the primary tumor or even from metastatic tumors, shed into the vasculature or lymphatic system and found disseminated in the patient’s blood circulation. Many studies have evidenced CTC presence in TNBC patients at different stages  with the detection of enhanced nuclear PARP-1 activity and observed an increased enumeration of CTCs in a metastatic advanced stage of TNBC and a drop of CTC percentage after neoadjuvant chemotherapy ,172. At t cancer . A retrot cancer . Of notet cancer .In this review, current and potential tissue biomarkers of TNBC with metabolic or pathological functions related to T2DM, including their impact in T2DM/TNBC patients, are mentioned in BRCA1 and BRCA2 germline mutations lead to impaired cell cycle and DNA repair, which represent 70–80% and 16–23% of all TNBC cases [BRCA1 promotes methylation, causing downregulation of the gene expression profile due to homologous recombination deficiency (HRD) and was associated with elevated risk of developing TNBC [BRCA1 mutation carriers of developing TNBC than BRCA2 mutation carriers do [BRCA1/2 germline mutations enhance TNBC sensitivity to DNA-damaging agents, resulting in a prolonged survival of these carriers diagnosed with TNBC [BRCA1/2 carriers compared with carriers without cancer, especially for women with high BMI [While BRCA1 has been reported for its metabolic actions, BC cases . A recening TNBC . Clinicoriers do ,181. Thehigh BMI .The epidermal growth factor receptor (EGFR) family  constitutes a group of transmembrane protein receptors classified into a subfamily of four closely related ErbB receptor tyrosine kinases  for specific extracellular protein ligands such as EGF or even transforming growth factor α (TGFα) . Upon acThe vascular endothelial growth factor (VEGF) is the most potent pro-angiogenic growth factor mainly expressed in hypoxic conditions by the hypoxic stress transcription factor hypoxia-inducible factor alpha (HIF-α). VEGF is a family of ligands  which are specific to the VEGFR family, and results in the stimulation of endothelial cell proliferation and migration, increase of vascular permeability, and enhances tumor cell extravasation. Predominantly expressed in vascular cells including endothelial and smooth muscle cells, the main actors of the formation of new blood vessels from pre-existent vasculature, VEGF is the main factor regulating metastasis formation. In diabetic conditions, VEGF mainly mediates endothelial dysfunction, including diabetic vasculopathy . VEGF is− breast cancer, including TNBC and especially in ductal TNBC, a high Ki-67 expression level is considered as a poor prognostic with predictive potential as it is associated with poor outcomes while high mitotic activity in the tumor shows a very good clinical response to combination chemotherapy [+/HR− subtype) were subjected to lipid profiling and IHC staining measuring Ki-67 score [Ki-67, also known as MKI67 (Marker of Proliferation Ki-67), is a nuclear antigen detected during cell cycle interphase  and mitosis while Ki-67 is absent from the cells in a quiescent phase, making Ki-67 the main cell proliferation biomarker routinely used in clinical practice . The grootherapy . A recen67 score . Serum d67 score . Another67 score .TP53 gene cause loss of functions of cell cycle checkpoint controls and impairment of the DNA damage repair system, promoting genomic instability which lead to uncontrolled cell proliferation with replication of damaged DNA replication, resulting in cancer onset, development, and in cancer progression by stimulating invasion and metastasis [TP53 mutations are frequently detected in 65 to 80% of the cases of TNBC patients compared to in luminal breast cancer cases (12%) and were associated with poor prognosis, suggesting p53 as a biomarker [TP53 mutations scoring in patients’ TNBC tissues and patients’ response to conventional neoadjuvant chemotherapy, a recent precision oncology-related study using the Evolutionary Action Scoring system reported the strong association between TP53 mutations and worst outcomes among 96 TNBC patients with residual disease following chemotherapy [TP53 mutations in hyperglycemic conditions and a potentiation of p53 mutants of insulin-induced AKT1 activation by binding and inhibiting the tumor suppressor disabled homolog 2-interacting protein (DAB2IP) in the cytoplasm [TP53 Pro72Arg polymorphism) from a case–control study performed on a Chinese population, the p53 serum level was determined to be strongly correlated with clinical and biochemical parameters in 225 T2DM in comparison to 255 non-diabetic control individuals [The p53  is a stress-sensitive nuclear tumor suppressor transcription factor protein that plays a vital role in controlling cell cycle, apoptosis , and in preserving DNA repair and genomic integrity, which gives it its nickname of “guardian of the genome” . Non-cantastasis ,204. TP5iomarker ,206. Altotherapy . Experimytoplasm . Recentlividuals ,210.Poly(ADP-ribose)polymerase (PARP) is a nuclear enzyme that plays a prominent role in the catalysis of the ADP-ribose units polymerization, units derived from the ADP donor nicotinamide adenine dinucleotide (NAD)+. PARPs constitute a family of eukaryotic enzymes responsible for the maintenance of cellular homeostasis through their involvement in various biological processes, including DNA damage repair (homologous recombination and non-homologous end-joining-based repair mechanisms), transcription, genomic stability, chromatin structure modulation, recombination, cell replication, cell adhesion and motility, innate immunity, and transformation . At the p < 0.005) in grade II presenting ER+/PR+/HER2− and not in grade I or even in advanced grade III  S6 Kinase 1 (p70S6K1) is the downstream effector of lipid kinase PI3K/Akt/mTOR complex 1 (PAM), one of the most important and active pathways in breast cancer including TNBC and in metabolic alterations such as T2DM ,223. Therade III B. As forrade III . VatsebaMetformin, chemically known as 1,1 dimethyl-biguanide hydrochorine, is a member of the class of guanidines derived from the plant Galega officinalis (French lilac) and is frequently used as the first-line antihyperglycemic drug to lower glycemic level and manage T2DM without causing hypoglycemia in T2DM subjects. Indicated as an adjunct to diet and exercise, metfomin exhibits various hypoglycemic properties through reduction of hepatic glucose production, decreasing the intestinal absorption of glucose and through enhancement of insulin sensitivity by increasing both peripheral glucose uptake and utilization . HoweverT2DM subjects have an increased risk of developing breast cancer, especially TNBC, a fast growing and aggressive malignant tumor usually presenting poor clinical outcome due to the lack of targeted therapies, which requires to be screened and detected early. This review lists the main clinicopathological features, related risk factors, current and potential biomarkers for clinical screening, diagnosis, prognosis, monitoring, and development of novel therapies including immunotherapies, and ongoing clinical trials for TNBC patients with T2DM. So far, immunotherapies intensively implemented in clinical trials to improved TNBC outcome require challenging molecular and cellular investigations for T2DM patients with exhausted immune systems and dysfunctional tumor microenvironments ,242. Ris


Objective This study evaluated the risk of the hereditary breast and ovarian cancer (HBOC) syndrome in patients with breast cancer by using the Family History Screening 7 (FHS-7) tool, a validated low-cost questionnaire with high sensitivity able to screen the HBOC risk in the population.Methods Women diagnosed with breast cancer (n = 101) assisted by the Unified Health System at the 8th Regional Health Municipal Office of the state of Paraná answered the FHS-7, and the results were analyzed using IBM SPSS Statistics for Windows, Version 25.0. software .Results The risk of HBOC was 19.80% (n = 20). Patients at risk exhibited aggressive tumor characteristics, such as high-grade tumors (30%), presence of angiolymphatic emboli (35%), and premenopausal at diagnosis (50%). Significant associations between the prevalence of high-grade tumors were observed in women younger than 50 years at diagnosis with HBOC (p = 0.003).Conclusion Our findings suggest a possible family inheritance associated with worse clinical features in women with breast cancer in this population, indicating that HBOC investigation can be initially performed with low-cost instruments such as FHS-7. BRCA1,BRCA2, andTP53.Breast cancer is the major cause of women's death in the world. In Brazil, 60,000 new cases are estimated for 2020.Age at diagnosis under 50 years old, pre-menopausal status, and being overweight at the time of diagnosis represent independent risk factors for the disease.However, genetic testing for HBOC is not affordable for many patients, and the implementation of cancer counselling and testing in the public health system represents a challenge due to its high cost. A simple, low-cost, sensitive, and easily applicable instrument in primary care units would be particularly useful in underserved communities, where the identification and referral of high-risk individuals is difficult.In this context, a Brazilian study conducted by Ashton-Prolla et al.As far as we know, the FHS-7 instrument has never been applied in the population of Southern Paraná. Thus, the present study aims to use the FHS-7 tool to screen breast cancer patients treated in a public oncology reference center to outline the syndrome profile in this population and investigate any clinicopathological correlation.n = 350) attended in a public oncology hospital located in Francisco Beltrão, Paraná, which is the reference for 27 municipalities in the Southwest of Paraná under the 8th Regional Health Municipal Office of the State of Paraná and the Unified Health System, from January 2016 to August 2018. The patients were selected during routine doctor appointments, after analysis of image exams and/or biopsy results.This is an observational, descriptive, and prospective transversal study, approved by the ethics committee of the Universidade do Estado do Paraná (CAAE 35524814.4.0000.0107). Patients (n = 101), and the inclusion criteria consisted in patients that were able to fully answer the questions, dichotomized as yes or no, in the FHS-7 questionnaire  who volunteered and signed a consent form ;p-values were reported). For these analyses, the software IBM Statistical Package for the Social Science for Windows version 25.0  was used. The statistical significance adopted for the analyses was 5%. Only the significant associations were reported in the study.The variables were analyzed for statistical assumptions of normality (Shapiro-Wilk test) and homoscedasticity (Levene test). The results were assessed with the Grubbs test for the detection of outliers, and no outliers were detected. The frequency of the variables was determined as percentages, and the Person Chi-square and linear logistic regression tests were used to verify putative associations between the frequencies of the categorical variables (β andn = 81). The remaining patients (n = 20), the HBOC-risk group, answered positively to at least one of the instrument questions, which results in a prevalence of 19.8% for HBOC risk in the studied population. All patients included were Caucasian, agreeing with the characteristics of European colonization in the region studied.Among the patients, 80.2% answered negatively to all questions of the FHS-7 questionnaire, gathered in the “sporadic cancer” group  in the sporadic cancer group , when compared with the HBOC-risk group . Regarding the overexpression of HER2 receptors, there was a predominance in the HBOC-risk group in comparison to the sporadic cancer group . The clinicopathological characterization of patients is recorded inBoth groups showed a similar distribution regarding molecular subtypes, age at diagnosis, menopause status and BMI classification as overweight/obese. The HBOC-risk group presented high-grade tumors more frequently than the sporadic cancer group , as well as intratumoral angiolymphatic emboli (35% vs 33% from the sporadic group) and lymph node involvement (25% vs 23% from the sporadic cancer group).p-values of β are described inRegarding the HBOC-risk group, we found significant association between high histological grade and the occurrence of triple-negative tumors and high histological grade at diagnosis with age at diagnosis (under 50 years). Lymph node involvement was found to be significantly associated with the presence of angiolymphatic emboli in patients as well as being menopausal at diagnosis. The associations observed in the sporadic cancer group show a direct relationship between subtype and histological grade and between angiolymphatic emboli and lymph node involvement. The beta andBRCA1/2genes is associated with the occurrence of triple negative breast tumors and are higher in Caucasians when compared with Hispanics.BRCA1/2.As far as we know, this is the first Brazilian study to apply the FHS-7 tool for screening the risk of HBOC in women with breast cancer in the Southwest region of Paraná, where most of its population is composed by European ancestry and inbreeding due to the geographic peculiarities of rugged relief. The prevalence of mutations in then = 5). No involvement was found concerning male relatives.The FHS-7instrument has high sensitivity for large-scale HBOC risk screening when compared with other molecular biology tools.BRCA1gene protects breast tissue from genomic instability induced by estrogen, which could be associated with the better prognosis observed in tumors with positive hormonal receptors.Regarding hormone receptors, patients in the HBOC-risk group had positive estrogen and progesterone receptors in only 35% and 45% of the cases, respectively, which may impair the response capacity to anti-hormone therapy and could impact the prognosis. In contrast, the sporadic cancer group showed an index of 46% for estrogen and 52% for progesterone. The normal functioning of theIncreased high-grade tumors were found in the HBOC-risk group (45%), representing an important factor of worse prognosis and lower survival.BRCA1andBRCA2mutation carriers, which are strongly associated with HBOC.In addition, the presence of intratumoral angiolymphatic emboli may predict the likelihood of lymph node invasion and could be a poor prognostic finding.BRCA-mutation carriers due to impaired DNA repair against lesions promoted by free radicals.Regarding age at diagnosis, older women had high-grade tumors. However, aging leads to the accumulation of free radicals, molecules capable of maintaining genomic instability that promotes cellular undifferentiating in breast tumors.It is argued that even in the presence of lymph node involvement or tumors with aggressive subtypes, postmenopausal patients have a better prognosis when compared with those diagnosed younger than 35 years old.Exclusive associations were found in the HBOC group. Positive significant associations were identified concerning age at diagnosis and histological grade, indicating that older patients from the HBOC group have more undifferentiated tumors. The expression of BRCA1 is associated with familial breast tumors history;The limitations of the study include the small sample size, which precluded a more refined statistical design and the assessment of relevant outcomes, such as survival and relative risk. A strength of the study is that this kind of data are scarce in literature and add knowledge since they bring new information about the clinicopathological characteristics of women with breast cancer at HBOC risk in a geographical area where this syndrome seems to have a high prevalence.The present study used an investigation tool to evaluate the prevalence of HBOC risk in patients from the Southwest region of Paraná, whose prevalence exceeded the one found by other published studies in similar populations. Besides, patients at risk were associated with clinicopathological factors related with aggressive disease. The study indicates that the FHS-7 questionnaire could be used for the identification of patients at high risk for HBOC syndrome, enabling early detection and effective cancer prevention as well as genetic counselling.


The inflammatory and insulinemic potentials of diets have been associated with colorectal cancer risk. However, it is unknown whether the plasma metabolite profiles related to inflammatory diets, or to insulinemic diets, underlie this association. The aim of this study was to evaluate the association between metabolomic profile scores related to the food-based empirical dietary inflammatory patterns (EDIP), the empirical dietary index for hyperinsulinemia (EDIH), and plasma inflammation  and insulin (C-peptide) biomarkers, and colorectal cancer risk. Elastic net regression was used to derive three metabolomic profile scores for each dietary pattern among 6840 participants from the Nurses’ Health Study and Health Professionals Follow-up Study, and associations with CRC risk were examined using multivariable-adjusted logistic regression, in a case-control study of 524 matched pairs nested in both cohorts. Among 186 known metabolites, 27 were significantly associated with both the EDIP and inflammatory biomarkers, and 21 were significantly associated with both the EDIH and C-peptide. In men, odds ratios (ORs) of colorectal cancer, per 1 standard deviation (SD) increment in metabolomic score, were 1.91 (1.31–2.78) for the common EDIP and inflammatory-biomarker metabolome, 1.12 (0.78–1.60) for EDIP-only metabolome, and 1.65 (1.16–2.36) for the inflammatory-biomarkers-only metabolome. However, no association was found for EDIH-only, C-peptide-only, and the common metabolomic signatures in men. Moreover, the metabolomic signatures were not associated with colorectal cancer risk among women. Metabolomic profiles reflecting pro-inflammatory diets and inflammation biomarkers were associated with colorectal cancer risk in men, while no association was found in women. Larger studies are needed to confirm our findings. Colorectal cancer is a major cause of morbidity and mortality, in the United States and globally . SeveralFurthermore, we found that the EDIP and EDIH are robustly associated with an increased risk of colorectal cancer, with a stronger association shown in men than in women ,12,13,14Metabolites are the final product of preceding molecular processes related to genomics, transcriptomics, and proteomics ,23,24. Mn = 121,701) enrolled female-registered nurses (married at the time) aged 30–55 years in 1976 .,43.42,43Three EDIP-related metabolomic signatures were created: (1) a metabolomic profile score including metabolites associated mainly with the EDIP, irrespective of their association with biomarkers (EDIP-only metabolome), (2) a metabolomic profile score comprised of metabolites associated mainly with inflammatory biomarkers, not considering their associations with the EDIP (BIOM-only metabolome), and (3) a metabolomic profile score reflecting the metabolic potential of both an inflammatory dietary pattern, and circulating inflammatory biomarkers (metabolomic dietary inflammatory pattern (MDIP)). Similarly, three EDIH-related metabolomic signatures were created: (1) a metabolomic profile score including metabolites associated mainly with the EDIH, irrespective of their association with biomarkers (EDIH-only metabolome), (2) a metabolomic profile score comprised of metabolites associated mainly with C-peptide, not considering their associations with the EDIH (CPEP-only metabolome), and (3) a metabolomic profile score reflecting the metabolic potential of both an insulinemic dietary pattern, and circulating C-peptide (metabolomic dietary index for hyperinsulinemia (MDIH)).Next, the associations between the EDIP (diet), the EDIH (diet), and the six metabolomic profile scores  in 1 standard deviation (SD) increment, and colorectal cancer risk, were examined, separately by sex, using conditional logistic regression analyses. Basic models were adjusted for only the matching factors, including age (continuous), sex , time of blood collection (continuous), race/ethnicity (white or non-white), and menopausal status for women . The multivariable model was further adjusted for BMI (continuous), physical activity (continuous), regular aspirin use (yes or no), smoking status , family history of colorectal cancer (yes or no), endoscopy (yes or no), alcohol intake (continuous), and total calorie intake (continuous). An additional multivariable model further adjusting for coffee intake (which contribute to a lower EDIP and EDIH), but which had metabolites related to caffeine, which may not influence inflammation, was conducted. To gain insight into potential metabolic pathways, individual metabolites of MDIP and MDIH were examined in relation to colorectal cancer risk. All analyses were performed using an R package for metabolomic signature generation, and SAS version 9.4  for all other analyses.2, respectively. The majority of participants were White. There were no differences in characteristics among the participants for the metabolomic, and colorectal cancer case-control study, analyses. Compared with the controls, the cases tended to be more likely to be overweight or obese, not use NSAIDs, have a positive family history of colorectal cancer, and be less likely to have had a previous lower gastrointestinal endoscopy.Selected participant characteristics, in each of the two cohorts, and by colorectal cancer case-control status in the colorectal cancer subsample, are summarized in Among 186 known metabolites, 82 metabolites for the EDIP, 43 for the inflammatory biomarker z-score, 120 for the EDIH, and 25 for C-peptide from the elastic net regressions were identified. A total of 27 metabolites were commonly associated with both the EDIP and the inflammatory biomarker z-score, and 21 metabolites were commonly associated with both the EDIH and C-peptide. The EDIP (diet) had correlations of 0.42 with EDIP-only, 0.21 with BIOM-only, and 0.20 with MDIP metabolomic scores. The EDIH (diet) had correlations of 0.44 with EDIH-only, 0.14 with CPEP-only, and 0.15 with MDIH metabolomic scores.Metabolomic profiles based on both pro-inflammatory diets and inflammation biomarkers were associated with colorectal cancer risk in men . The ORsMetabolomic profiles reflecting insulinemic diets and the C-peptide biomarker were not significantly associated with colorectal cancer risk . In men,When the individual associations between the common metabolites of the EDIP and inflammatory biomarkers, and colorectal cancer risk, were examined  in men, In the current study, an integrated analysis of the association between inflammatory and insulinemic dietary patterns, plasma inflammation, and insulin biomarkers, plasma metabolomics, and colorectal cancer risk, was conducted. We identified 27 metabolites associated with both an inflammatory dietary pattern, and plasma inflammation biomarkers, termed the common metabolome for inflammation (MDIP); and 21 metabolites associated with both an insulinemic dietary pattern, and C-peptide levels, termed the common metabolome for hyperinsulinemia (MDIH). When we examined the association between the identified metabolomic signatures, and colorectal cancer risk in a nested case-control study, metabolomic profiles reflecting pro-inflammatory diets and inflammation biomarkers were associated with colorectal cancer risk in men. However, we found no association for metabolomic profiles reflecting insulinemic diets (MDIH) and C-peptide in men. Moreover, the metabolomic signatures for insulin and inflammation were not associated with colorectal cancer risk among women.n = 394) than in men (n = 130). Thus, sample size alone is not likely to explain the sex differences. In fact, we had lower power in men than in women to detect associations.As reported earlier, we did observe associations with increased risk for EDIP and EDIH among both men and women in the full cohort. Consistent with previous studies, we found that the EDIP and EDIH were positively associated with colorectal cancer risk in a nested case-control study with metabolomic data, though confidence intervals were wide , given the much-smaller sample sizes. For men, signatures for MDIP (robustly) and MDIH (suggestively), and biomarker-only, were associated with the risk of colorectal cancer. However, these metabolomic patterns were not associated with colorectal cancer in women. The limited sample size for the metabolomic analyses, resulting in statistical instability, may have played a role in the inconsistent results, though the number of case-control pairs was greater in women . It is possible that metabolomics may be more closely associated with insulin resistance than with high levels of insulin , although we could not directly evaluate this speculation. If a circulating level of insulin (a mitogen and anti-apoptotic hormone) is the direct causal factor, as suggested by a Mendelian randomization study , then diLarger studies are required to confirm if the stronger associations we saw in men are robust, or due to random fluctuation resulting from a limited sample size. Some data do suggest that the association between insulin resistance and colorectal cancer is stronger in men than in women . In addiWe observed evidence of sex differences in the relationship between metabolomic signatures of an inflammatory dietary pattern and biomarkers, and colorectal cancer risk. A significant positive association was found only in men, and not in women. When we examined the individual metabolites for MDIP with colorectal cancer, we also found differences in the metabolites associated with colorectal cancer risk by sex. In men, 1-methylguanosine, N2.N2-dimethylguanosine, and N-carbamoyl-beta-alanine were positively associated, and C36:2 PE plasmalogen was inversely associated, with colorectal cancer risk, but C2 carnitine and biliverdin were positively associated with colorectal cancer risk in women. These findings suggest potentially different biological mechanisms linking dietary inflammatory potential to colorectal cancer development by sex, or may reflect random variation. Several modified nucleosides in urine, including 1-methylguanosine and N2.N2-dimethylguanosine, have been identified since the mid-1980s as biochemical indicators of malignant disease , includiThere were a number of strengths to our study design. The sample for the derivation of metabolomic profile scores was sufficiently large to produce reliable patterns. We utilized a robust statistical methodology to identify the important metabolites. Elastic net regression has the advantage of effectively dealing with highly correlated variables , while maintaining the quality of model selection. Our cohorts contained comprehensive covariate data, which allowed the control of potential confounding factors. We used state-of-the art interdisciplinary techniques to integrate metabolomic, biomarker, and dietary data, to identify potential mechanistic pathways linking dietary patterns and colorectal cancer risk. Finally, the repeated measuring of diet reduced potential measurement error.However, our study had several limitations. Firstly, we had a relatively small colorectal cancer nested sample (524 case-control pairs), which may have reduced the statistical power to detect significant differences. Of relevance to this, similar-sized but statistically significant associations for diet variables were observed in the full cohort, indicating that our power may have been limited in the nested case-control study. Although important on a population basis, the relative risks were relatively moderate in magnitude. Secondly, metabolomic measurement was conducted once for each participant. However, our previous study showed high stability for most of the metabolites over 1 to 2 years within individuals. Of note, this approach in our cohort did produce robust findings for both insulin and inflammation metabolites with risk of type 2 diabetes, a disease that is strongly associated with these biological processes. Colorectal cancer has multiple etiologic factors, of which insulin resistance and inflammation may be one of many, so associations would be expected to be weaker than for diabetes. It is also possible that associations are stronger for molecular or anatomical sub-types of colorectal cancer, but we could not examine this with adequate power . MoreoveIn conclusion, we found that metabolomic profiles reflecting pro-inflammatory diets and inflammation biomarkers were associated with colorectal cancer risk in men. Results for hyperinsulinemic profiles were suggestive in men. However, no association was observed between the metabolomic signatures and colorectal cancer risk in women. Larger studies are needed to confirm our findings.


Ductal carcinoma in situ (DCIS) is recognised as a precursor to invasive breast cancer (IBC), while lobular carcinoma in situ (LCIS) is considered a risk factor for subsequent IBC. To investigate whether the factors that increase the risks of DCIS and LCIS also predispose to IBC, we analysed risk factors for 3075 DCIS cases, 338 LCIS cases, and 1584 controls aged 35–60. Results showed that breastfeeding after childbirth decreased risks of DCIS and LCIS, similar to the association with IBC. Post-menopausal hormone replacement therapy (HRT) increased the risks of DCIS and LCIS, especially in long-term use (10+ years), with a stronger association with LCIS. However, neither parity nor an increasing number of births decreased the risks of DCIS or LCIS, as they do IBC. The study shows both similarities and differences in the risk factors affecting in situ breast cancer and IBC, and results suggest that regular surveillance is warranted in post-menopausal women taking long-term HRT.Ductal carcinoma in situ (DCIS) is widely accepted as a precursor of invasive ductal carcinoma (IDC). Lobular carcinoma in situ (LCIS) is considered a risk factor for invasive lobular carcinoma (ILC), and it is unclear whether LCIS is also a precursor. Therefore, it would be expected that similar risk factors predispose to both DCIS and IDC, but not necessarily LCIS and ILC. This study examined associations with risk factors using data from 3075 DCIS cases, 338 LCIS cases, and 1584 controls aged 35–60, recruited from the UK-based GLACIER and ICICLE case-control studies between 2007 and 2012. Analysis showed that breastfeeding in parous women was protective against DCIS and LCIS, which is consistent with research on invasive breast cancer (IBC). Additionally, long-term use of HRT in post-menopausal women increased the risk of DCIS and LCIS, with a stronger association in LCIS, similar to the association with ILC. Contrary to findings with IBC, parity and the number of births were not protective against DCIS or LCIS, while oral contraceptives showed an unexpected protective effect. These findings suggest both similarities and differences in risk factors for DCIS and LCIS compared to IBC and that there may be justification for increased breast surveillance in post-menopausal women taking long-term HRT. Breast cancer is the most common cancer in women, accounting for over 2 million new diagnoses globally each year . AlthougDCIS is a non-invasive form of breast cancer which arises in the ducts of the breast and accounts for about 15% of breast cancer diagnoses in the UK, with over 7000 women diagnosed per year . HoweverIn contrast, LCIS is clinically and radiographically occult and generally an incidental finding on biopsy (~3% of needle biopsies)  or assocAfter the introduction of the breast screening programme, the number of women diagnosed with in situ disease in the UK has almost tripled since the 1990s . It is lThis study aims to summarise the risk factors that have been previously shown to predispose to DCIS and LCIS and to determine whether the risks associated with these risk factors are similar for DCIS and LCIS, and to associations previously found with IBC. This will be explored using a large cohort of DCIS and LCIS cases recruited to the ICICLE (A study to Investigate the genetiCs of In situ Carcinoma of the ductaL subtypE) and GLACIER (A study to investigate the Genetics of LobulAr Carcinoma In situ in EuRope) studies, which collectively recruited 2121 controls and 5900 cases between 2007 and 2012.Although there have been a number of studies looking at risk factors predisposing to DCIS, the studies looking at LCIS are limited and are likely underpowered to detect significant associations due to small study populations.It is well documented that the risk of IBC is much higher in those with a family history of breast cancer than those without, with a previous study of over 100,000 UK women estimating that 15% of women diagnosed with breast cancer have a first-degree family history of breast cancer, corresponding to a ~1.8-fold increase in breast cancer risk (either in situ or invasive) .A similar association is also seen with DCIS ,39,40,41BRCA1, BRCA2, TP53, and more recently, PALB2 and CHEK2, is well established. It is understood that the presence of genetic mutations is more prevalent in those diagnosed at a younger age, particularly in oestrogen receptor-negative IBC (ER-negative IBC), with all women in the UK being offered genetic testing if diagnosed with ER-negative IBC aged 60 or under [BRCA1 and BRCA2 mutations are found in around 10% of IBC diagnoses and confer a 5–10-fold increase in the odds of developing breast cancer [PALB2 gene mutations are found in <1% of breast cancers [TP53 mutations in up to 3% of women diagnosed with breast cancer under 30 [CDH1 gene have been specifically associated with ILC, often with a history of diffuse gastric cancer [BRCA2 carriers than BRCA1 carriers [BRCA1 or BRCA2 genes [The increased risk of IBC due to rare inherited genetic mutations in genes such as or under . BRCA1 at cancer . Rarer m cancers , and TP5under 30 . Mutatioc cancer , and ILCcarriers . HoweverA2 genes .BRCA2 and CHEK2 mutations and a weaker association with PALB2, BRCA1, and TP53, although the frequency of BRCA mutations appears to be less frequent in DCIS than invasive disease [In many countries, women with DCIS do not qualify for genetic testing despite studies (including ICICLE ) showing disease ,52. Appr disease , increasCHEK2 gene and increased risk of LCIS, although this mutation was rare (3% of LCIS cases) [CDH1 mutations and unilateral pure LCIS, but germline CDH1 mutations have been found in women with bilateral LCIS (with/without ILC) [Data on pure LCIS is very limited, but previous research involving cases from the GLACIER study has shown an association between pathogenic germline mutations in the S cases) . There wout ILC) .Over the past decade, over 300 low-risk single nucleotide polymorphisms (SNPs) have been shown to predispose to IBC. A total of 313 of these SNPs have been combined to develop a polygenic risk score (PRS), which shows a greater than 30% increased risk of breast cancer for those that fall into the highest 1% of the PRS ,58,59,60IBC incidence rates increase with increasing age . SimilarThere has been a lot of research into the relationship between childbirth and the risk of IBC. Studies have shown that parous women are at decreased risk of developing ER-positive breast cancer and that increasing parity is associated with decreased risk of ER-positive breast cancer, but these associations were not found with ER-negative breast cancer ,66,67,68Some studies have found that the decreased risk was only apparent over a certain number of births, such as three or more births ,63,69, fDue to the small numbers, there is little data on the relationship between parity and LCIS alone. Two studies that included both DCIS and LCIS showed no evidence of an association between LCIS and being parous or parity number ,38 but dIncreasing age at first birth has also been shown to increase the risk of IBC ,67,73. SThe association between older age at first live birth and higher odds of developing breast cancer is also seen with DCIS ,40,63,70Breastfeeding has been found to be protective against IBC ,76,77. HEarly menarche and late menopause are associated with an increased risk of IBC due to the increased years of exposure to endogenous hormones, and this association is stronger in ILC compared to IDC . HoweverThere has, however, been evidence of an association between age at menopause and risk of developing DCIS. Previous research has found that women aged 55 or over at menopause had a significantly increased risk of developing in situ breast cancer ,71, withLifestyle behaviours such as smoking and alcohol consumption are modifiable risk factors that have been associated with increased IBC risk ,84,85,86Having a higher BMI has been associated with post-menopausal ER-positive IBC , and theIncreased hormone exposure can also come from the use of exogenous hormones, such as hormone replacement therapies (HRT) or some forms of contraception.Various studies have, so far, found evidence of a small increased risk of breast cancer with oral contraceptive use . One stuHowever, previous studies have shown clear evidence of an association between HRT usage and the increased risk of IBC, particularly ILC, with combined oestrogen–progestogen HRT ,42,43,94The ICICLE and GLACIER studies were two UK-based case-control studies recruiting participants between 2007 and 2013, with the aim of investigating genetic predisposition to DCIS and lobular breast cancer, respectively (MREC references: 08/H0502/4 and 06/Q1702/64).Cases were recruited to both studies at over 100 hospitals throughout the UK, and women aged 60 or younger at the time of diagnosis were eligible for the studies. For inclusion in the ICICLE study, cases had to be diagnosed with pure DCIS with no ipsilateral invasion >1 mm; for the GLACIER study, cases could be diagnosed with ILC, with LCIS alone, or LCIS concurrent with IBC. Controls were eligible for the study if they had no known family history of in situ breast cancer or IBC up to 2nd degree and also had no personal history of either in situ breast cancer or IBC. Cases identified control participants, as cases were asked to approach non-blood relatives and friends to act as controls and there were no age restrictions on controls.Of the ~5900 cases, 293 from ICICLE and 180 from GLACIER were found to be ineligible after recruitment based on the histopathology of their first tumour and were, therefore, excluded from the case-control analysis. Additionally, 12 controls were excluded due to missing date of birth or not sending the blood samples required for genotyping. After exclusions, there were 2109 controls, and 5422 cases between the two studies, with the cases categorised according to their primary diagnosis into these subgroups; 3136 with DCIS, 1560 with ILC, and 726 with LCIS, of which 386 LCIS cases were alongside an ipsilateral invasive tumour of other morphology (mixed type or IDC).For the purposes of this current study, only women aged ≥35 and ≤60 at diagnosis (for cases) or study entry (for controls) were included. The upper age limit was defined as 60 due to the ICICLE and GLACIER studies only recruiting cases aged 60 or under, although controls of any age were accepted. Additionally, the lower age limit of 35 was chosen as the controls had a different age distribution than the cases, and a small number of cases were younger than 35. Only the pure DCIS and LCIS cases were included, leaving 3075 DCIS and 338 LCIS cases, in addition to 1584 controls .As part of the ICICLE and GLACIER studies, all participants were asked to fill out a questionnaire at study entry, which included questions on past medical history, use of exogenous hormones, ethnicity, and reproductive history, which provided data on risk factors . For casThe number of births was calculated as the number of full-term births, including stillbirths, as reported by participants. Twins counted as one birth. Family history of cancer was considered as positive if 1st–3rd-degree relatives had reported a history of any cancer, and family history of breast cancer if 1st–3rd-degree relatives had a history of breast cancer. Where participants had not answered questions on childbearing/number of births or HRT use, these were considered not present (1.5% for number of births and 3% for HRT).Women were asked questions on menstrual history as part of the questionnaire, and menopausal status was inferred from answers to these questions using the following rules: women who were still having periods were considered pre-menopausal, and women who reported that their periods stopped naturally or after removal of ovaries were post-menopausal. This was inferred based on methodology in the Million Women Study, which was a large prospective cohort study of nearly 2 million women in the UK, looking at breast cancer risk ,97. For Variables with missing data over 10%  were imputed by multiple imputations using chained equations over 40 iterations. Variables used for the imputation included age at diagnosis, contraceptive use, HRT use, presence of rare genetic mutations, year of diagnosis, cancer morphology, parity, and age of menarche. Due to a lack of patterns in the data, missing data was imputed for most but not all missing data points. When attempting to impute the type of HRT taken, convergence was not achieved, and so this variable was not imputed. After multiple imputations, age at menopause had 1% missing, length of HRT use had 1% missing, and length of contraceptive use had 5% missing.Associations between in situ breast cancer groups and each of the risk factors were explored using individual logistic regression models compared to controls, adjusting by the age at study entry/diagnosis and then separated by inferred menopausal status. A case-only analysis was then undertaken to compare risk factors for DCIS and LCIS, using LCIS as the control group. A further multivariate logistic regression model was created, including the variables of interest that had displayed associations with either DCIS or LCIS. Logistic regression models were also run for the association between in situ breast cancer and oral contraceptive use, adjusted by year at diagnosis/study entry, and for the associations between in situ breast cancer and breastfeeding and HRT, stratified by age categories . In addiAge was considered as a confounder in all logistic regression models due to the likeliness of age being associated with multiple variables, such as contraceptive use, parity and number of births, menopausal status, and HRT use, and due to the difference in age distribution between the cases and controls. These logistic regression models were restricted to parous women only when looking at age at birth, breastfeeding, and the number of births, restricted to women who took HRT only when looking at the length of time taking HRT, and restricted to women who took the oral contraceptive (OC) pill only when looking at the length of time taking the OC pill. These logistic regression models were then presented as a forest plot. The “unknown” category of breastfeeding was removed from the forest plots due to wide confidence intervals and no significant association.All analyses were performed using STATA/MP 17.0.p < 0.0001 on the t-test comparing all in situ cases to controls), and a higher proportion of DCIS cases were inferred as post-menopausal at study entry.The characteristics of 3075 DCIS, 338 LCIS, and 1584 controls are shown in Logistic regression models were used to explore the associations between each of the risk factors and developing in situ breast cancer, initially considering all women and then in pre-menopausal and post-menopausal women separately. Each model for the different risk factors was adjusted by age at diagnosis for cases or age at study entry for controls. Logistic regression models were then run with cases only to assess differences in risk factor associations in DCIS and LCIS, with LCIS used as the control group. The results from the regression models for cases compared to controls are presented as forest plots in There were statistically significant increases in the odds of both DCIS and LCIS with increasing age that the period stopped, with a 2% increase in risk per increase in year , but no association with age of menarche . There wp = 0.054), albeit no strong association with parity. In parous women, breastfeeding significantly decreased the risk of developing both DCIS  and LCIS  regardless of menopausal status. Breastfeeding after childbirth was not associated with one in situ breast cancer over the other, neither overall nor when separated by menopausal status. When looking only at the pre-menopausal DCIS cases, this association was of borderline statistical significance  but was significant in pre-menopausal LCIS, conferring 39% reduced odds of LCIS (95% CI: 0.38–0.97). Breastfeeding in parous women was significantly associated with reduced odds of both DCIS and LCIS in post-menopausal women.There was no evidence of an association between DCIS and parity, number of births, or age at first birth, either when looking at all cases or when separated by menopausal status. However, there was a borderline association between an increased risk of LCIS and an increasing number of births  and LCIS by 62% , with even stronger associations in those who took long-term HRT (for 10+ years). HRT is sometimes offered to pre-menopausal women who are approaching menopause. Despite only a small proportion of pre-menopausal women taking HRT , this was associated with an increased risk of both developing DCIS  and LCIS . In post-menopausal women, HRT use significantly increased the risk of LCIS  but not DCIS .Long-term HRT use (10+ years) also significantly increased the risks of both DCIS  and LCIS  in post-menopausal women, albeit with a wide confidence interval in LCIS due to small numbers. Case-only analysis revealed that long-term HRT usage was more strongly associated with LCIS than DCIS , and this association was further strengthened when restricted only to post-menopausal cases . There were no associations between the type of HRT and developing either type of in situ breast cancer. A sensitivity analysis was conducted to address the potential differences in trends of exogenous hormonal use over time, excluding cases diagnosed before 2005. where, the association with long-term HRT use was still observed in both DCIS and LCIS .There appeared to be a protective effect of using oral contraceptives in both DCIS  and LCIS (OR: 0.72 95% CI: 0.51–0.99) compared to those not taking contraceptives. However, when the year of diagnosis was added to the logistic regression model (year of study entry with controls), this association was no longer significant in DCIS or LCIS . AdditioA positive family history of breast cancer was also more strongly associated with LCIS over DCIS , but this association was not significant when stratifying by menopausal status.The associations were further explored in a multivariate logistic regression model, including breastfeeding, parity, length of time taking HRT, age at diagnosis/entry for controls, and age when periods stopped . After aWhen exploring associations with pathological subtypes of DCIS as a secondary analysis, the oral contraceptive pill did not show any evidence of an association with specific DCIS grades but did show an association with ER-DCIS, indicating that the contraceptive pill use was associated with reduced odds of ER- DCIS specifically . HoweverThis current study has shown that HRT use increases the risk of both types of in situ breast cancer, especially when taken for 10+ years and even when taken in pre-menopausal women. This supports previously found associations in IBC and in situ disease, with similar findings of stronger associations with the lobular subtype ,101,102.This is the first study to show a protective effect of breastfeeding against DCIS and LCIS, both when adjusted by age at diagnosis/study entry in parous women only, and on the multivariate analysis including HRT use, parity, and age period stopped. Additionally, when stratified by age, statistically significant associations were in the same direction, so this association is unlikely just due to the difference in age between cases and controls . DespiteThe lack of a protective effect with the number of births could be due to women in this study not having a high enough number of births to observe an association, as some studies have only seen associations of breast cancer in more than three children ,69,70,71For both DCIS and LCIS, there was no association with age at menarche and only a very small association with age at menopause for DCIS, supporting the findings of other studies of DCIS, but in contrast to IDC. DCIS and LCIS had very similar risk profiles, with only the length of HRT use and family history having a stronger association with LCIS than DCIS. The strong association of family history with LCIS was also observed by Claus et al. .p = 0.059) association with increased risk of DCIS, but not LCIS. The reason for the protective effect in post-menopausal women is not clear, but could be explained by potential confounding factors, bias, or artefact. As participants filled out questionnaires upon study entry and had to recall their past exposures, bias could affect this association, especially as cases may be more likely to remember medical history or risk factors related to developing breast cancer compared to controls. This is further supported by the slightly higher proportion of missing data on contraceptive use in controls. However, there was no other way to collect objective data on the risk factors included in this study. This association could also potentially be explained by differences in the oral contraceptive pill offered over time, which is supported by results in The finding of a protective effect of the oral contraceptive pill against in situ breast cancer was unexpected and is not in line with previous literature, which either shows a weak or no association. Interestingly, we did not see this association in pre-menopausal women, where we actually saw the opposite effect with pre-menopausal women taking the oral contraceptive pill for longer than 10 years having a borderline (Despite adjusting for age in all of the models, due to the difference in age distribution between the cases and controls, it is possible that the study could be influenced by birth cohort effects. However, when investigating this by matching cases to controls 1:1 , the assWe chose to also analyse patients by inferred menopausal status, and this status was inferred based on the methodology used in the Million Women Study ,97. SomeFrom published literature, it would be expected that the risk factors predisposing to DCIS would be similar to those predisposing to IDC, but with less data on LCIS, there were not the same expectations of similarities with risk factors predisposing to ILC. We have shown that the risk factors associated with LCIS in our study were similar to those found previously for ILC, with a clear association with HRT and long-term use in post-menopausal women and breastfeeding in parous women. Unlike ILC, there was no association with age of menarche and no protective effect of increasing the number of births. However, with only 338 in the LCIS cases, it is possible that the study had low statistical power to detect some associations in the LCIS group, and important associations could have been missed. The smaller number of LCIS cases could be a consequence of LCIS being less frequent and possibly also due to LCIS diagnoses being mostly incidental findings, in contrast to DCIS.This study has found some similarities in risk factors pre-disposing to both forms of in situ breast cancer and compared to established associations with IBC from previously published studies, e.g., with breastfeeding after childbirth and post-menopausal HRT use. It has shown that some risk factors are more strongly associated with LCIS over DCIS, such as length of HRT use and family history. It would be important to repeat these analyses in incident cases only and further examine the association between decreased risk of in situ disease and contraceptive use in post-menopausal women. However, with observed associations between increased risk of in situ breast cancer and HRT use, women should be informed of these risks with long-term HRT and encouraged to reduce the duration of HRT use. There is an argument for increased radiographic surveillance in women taking long-term HRT.


Regular breast self-examination is the most cost-effective method for early detection of breast cancer, especially in resource-limited countries. However, breast self-examination practice was low among reproductive-age women.This study aims to evaluate breast self-examination practice and associated factors among women of reproductive age in southeast Ethiopia.p-value <0.05 during multivariable logistic regressions were considered as significantly associated with the dependent variable. Thematic data analysis was carried out for the qualitative study.A convergent parallel mixed-methods study design was conducted on 836 women of reproductive age. An interviewer-administered questionnaire was used for the quantitative part of the study and supplemented by focus group discussions. A database was created using Epi-info version 3.5.3, and analyzed using SPSS version 20. Bivariate and multivariable logistic regressions were done to examine the effect of explanatory variables. Variables with Out of 836 total participants, only 20.7% had ever heard about breast self-examination practice. Also, only 13.2% of the mothers had practiced breast self-examinations. Even though the majority of the participants during focused group discussions had knowledge about breast cancer screening, most of them stated that breast self-examination was not practiced among them. Maternal age, mother’s level of education, and previous history of breast examination by health professionals were significant predictors of breast self-examination practices.This study reported a low prevalence of breast self-examination practice. Therefore, enhancing women’s education and encouraging breast examination by health professionals are essential to increase the proportion of women performing breast self-examination. Breast cancer (BC) is typically asymptomatic when the tumor is small and easily treated. Therefore, it is very essential for women to follow suggested screening rules in detecting BC in the early stage . Before Age is the most important factor for BC risk in women. It will affect women in their most productive years of life. More breast cancer cases are identified in developing countries than those in developed countries among women of reproductive age (15–49 years) . The othThe most imperative strategies in achieving early detection of BC are mammography and physical examination of the breasts by qualified health workers, or clinical breast examination (CBE) and breast self-examination (BSE) , 9, 10. In 2018, breast cancer represents approximately 24.2% of all new cancer cases and 15% of 4.2 million deaths due to cancer globally, of which sub-Saharan Africa accounts for 8.1% of breast cancer cases and 11.8% of breast cancer deaths. It is the most prevalent cancer in developing countries , 9, 13. A convergent parallel mixed-methods study design was conducted from 1 March 2017 to 30 May 2017 in Bale Zone. Robe town is the capital city of Bale Zone, which is 430 km from Addis Ababa, the capital city of Ethiopia. The Bale Zone has 20 districts with farming communities predominating. The study was conducted in Ginnir, Sinana, and Madda Walabu districts, which are located at 560 km, 430 km, and 630 km southeast of Addis Ababa, respectively. According to the last census survey (2007) projection in 2017, the estimated population of Ginnir, Sinana, and Madda Walabu districts were 159,766, 154,170, and 127,426, respectively. Women of childbearing age numbered 35,357 for the Ginnir district, 34,118 for the Sinana district, and 28,200 for the Madda Walabu district . The GinIn the selected household, adult participants responded to the questionnaire.All selected women of childbearing age (15–49 years) living in the study area and residing for more than 6 months in the selected district and interest in participating in the study were included in both qualitative and quantitative sections.Non-Ethiopians, pregnant women, women with known breast cancer, seriously ill women during data collection, and women not willing to participate in the study were excluded in both quantitative and qualitative studies. Participants in the quantitative study were also excluded from the qualitative study.A single population proportion formula was used to determine the sample size for the quantitative data. A sample size of 841 was calculated based on an estimated proportion of BSE practice of 53.6% ; the desSix focus group discussions (FGDs) were conducted for the qualitative part, which was supplementary to the quantitative study, with two FGDs in each district. A total of 59 discussants participated . Five FGA multi-stage sampling technique was used to select the study participants. In the first stage, three districts were selected randomly. The kebeles in the selected districts were identified and stratified into urban and rural kebeles. The selected urban and rural districts were selected by using a lottery method. The selection of kebeles was dependent on their distance from the capital town of the district, taking the hospital as a center. For the three districts, the furthest kebeles on average were 45 km from the center. Three kebeles closest to the town (at approximately 15 km), three kebeles from the middle region (at the second 15 km), and three kebeles furthest from the town (at the third 15 km) were selected randomly. A list of all households with eligible mothers was identified; finally, we used a sampling frame to pick the study participants using the simple random sampling method.For the qualitative data, purposive sampling technique was used to select participants. From each selected district, two groups of childbearing-age women not included in the quantitative study were asked to participate in the FGD.For quantitative data collection, a standardized questionnaire was initially developed in the English language after reviewing different literature reflecting the purpose of the study –24, thenTwelve diploma nurses collected interview data guided by three supervisors. The training of data collectors and supervisors mainly focused on issues such as data collection tools, field methods, inclusion–exclusion criteria, and record keeping. The investigators coordinated the interview process and reviewed completed questionnaires on a daily basis to ensure completeness and consistency of the data. The questionnaire was pre-tested on 5% of the sample outside the selected district for this study.The FGDs were held in the community meeting room and in health posts at the participant’s districts. The participants were women of childbearing age (15–49 years old) not included in the quantitative study and recruited purposively. An interview guide was used to conduct the FGD and a tape recorder was used to capture the conversation. The interview guide was originally prepared in English, then translated to the local language (Afan Oromo), and translated back to English to check the consistency. Each FGD was conducted by two trained female degree holder nurses; one moderated the discussion while the other took notes and recorded the tape. One gatekeeper  for each FGD was assigned.p ≤ 0.25 were retained for subsequent multivariable logistic regression. Strength of association was tested using adjusted odds ratio (AOR) and 95% confidence interval (CI). The significance level considered for multivariable logistic regressions was p ≤ 0.05.The quantitative data was entered into a database using Epi-info, version 3.5.3, and it was exported to SPSS version 20 for data analysis. First, descriptive statistics were used to describe the data. Then, simple logistic regression analysis was carried out; those variables that were significant at For qualitative data, tape recordings were transcribed, organized in narrative forms congruent with the respondents’ own words, and analyzed under selected themes based on the question guide and summarized manually; i.e., thematic type of data analysis was carried out for the qualitative study.A total of 836 mothers completed the questionnaire for a response rate of 99.4%. The mothers had a mean ( ± SD) age of 31.09 ± 7.34 years. Forty-five percent of the study participants were illiterate. The majority [744 (89.0%)] of the study participants were married. Ninety percent of women were housewives. Regarding the husband’s level of education, more than half [439 (52.5%)] had finished primary school. The majority [463 (55.4%)] of respondents had a monthly income below the poverty line . The majority [475 (56.8%)] of the study participants had a television or radio. The majority [210 (74.9%)] were rural dwellers. Health centers or hospitals were very close to 667 (79.8%) of the study participants  Table 1.The majority [576 (68.8%)] of the respondents had sufficient knowledge about breast cancer screening. Regarding sources of information, television and radio were the main sources [98 (56.6%)]. This finding was supported by comments from a 19-year-old merchant mother who reported, “…in our place there is no awareness creating activities or education by doctors on breast problem: presence of the disease, its consequences, its sign and symptoms, and its risk factors, and an option for treatment. We heard information from the television and radio. Some of us even heard the presence of its management today from this discussion.”A middle-aged adult civil servant also stated that, “…I heard my neighbor complaining of a breast disease; I heard also that the disease was cancer. Many women have suffered with disease of the breast, yet I haven’t seen any breast disease on my body…. When I was a child, I heard a neighbor who died due to breast disease. Her breast was wounded, and she was referred and taken far out of this area to receive better treatment in referral hospitals. Even though she had visited many hospitals, she died as the consequence of the disease.”Only 187 (22.2%) participants knew the appropriate timing to perform BSE. The knowledge of how BSE is performed was known to 491 (58.7%) of the study participants. The majority [650 (77.8%)] of the mothers responded that performing BSE is important. Above 90% of respondents had visited health facilities at some time for sickness. However, only 37% of the mothers had a CBE by health workers during the visit Table 2.During another FGD, a housewife in her early 20s reported, “…this breast disease is certainly present in its massive form; I have not experienced this disease on myself; it has hurt many women; and some people say it became “hola” on a woman; some say cancer, and some others say another thing. Even at the moment, there is a woman with breast problem, she gave birth recently, her breast has not produced milk, and she has severe breast ache. The problem is existed among us, however; we don’t know the right time to perform BSE…”In this study, only 110 (13.2%) mothers performed BSE. The main reasons given by the mothers were lack of information [269 (37.1%)], fear of detecting abnormalities [158 (21.7%)], and lack of privacy [138 (16.5%)] Figure 1During an FGD about BSE, six mothers stated that BSE was not practiced among them. For example, a civil servant in her early 20s reported that, “…I don’t need to touch and examine my breast if I don’t suspect the problem. If it develops a problem, symptoms force us to touch and examine the breast. It is important to see some internally felt discomfort or externally observed signs and symptoms of disease. We don’t know of breast cancer. Therefore, we don’t give focus to our breast. If we have a previous problem, we check the improvement of that problem from time to time. Unless and otherwise, we don’t examine our breast…”Those mothers who performed BSE had detected abnormalities in their breasts such as changes in contour [36 (32.7%)] and lump in the breast [29 (26.4%)]. The majority [64 (62.7%)] of the respondents who detected abnormalities in their breasts did consult health workers Table 2.A middle-aged adult housewife also stated, “… for example, when I have pain of the breast, if all things, even children touch me, I feel pain on both breasts specially at the tip of it, before this time I haven’t ever see such things. For this reason, I have started to touch and look at my breast. If somebody has such problem that woman should touch and examine their own breast, otherwise there is no need to touch…”During another FDG, a middle-aged adult health extension worker stated that, “…women know their breasts or their bodies especially during a change. Breast pain is not simple, its pain is more severe than other disease; therefore, it is easy to know breast problem on ourselves. However, women have seen this problem traditionally and some of them mostly seek traditional treatment because, they will not permit to expose their breast to health professionals if the disease is not severe. This makes the disease too fatal among our community…”Women in the age range of young adults and middle age adults were more likely to practice BSE compared to those women in their early 20s, AOR = 3.61  and AOR = 9.35 , respectively. Participants who have finished primary education, AOR = 3.88 , and secondary and above, AOR = 11.14 , were more likely to practice BSE than illiterate mothers. In addition, participants who have ever had breast examination previously by health workers were more likely to practice BSE compared to those mothers who have never had CBE, AOR = 3.62  Table 3.BSE is an important and inexpensive method for the screening and early detection of breast cancer , 25. ThiThis study showed that as a mother’s age increases, especially a female in her early 20s, the practice of BSE increases. This is similar to a study conducted in north Ethiopia  of Bale Zone.As a cross-sectional study, a cause-and-effect relationship cannot be established to identify the actual predictor. The study focused only on reproductive-age women and did not include older women, for whom concerns about breast cancer may be greater.The study revealed low BSE practice among reproductive-age women, compared with the other studies conducted in Ethiopia. The proportion of the respondents who had ever heard of BSE was also low. Present maternal age, mother’s level of education, and previous history of breast examination by a health professional were found to have significant associations with BSE practices. Thus, increasing maternal education and coverage of breast examination by health professionals is crucial to increase BSE practice.The original contributions presented in the study are included in the article/Supplementary Material. Further inquiries can be directed to the corresponding author.Administrative approval was obtained before conducting the study and an ethical clearance letter was obtained from Madda Walabu University ethical clearance committee with approval number MU-14/51/78. An official letter of collaboration was written to Bale Zone administration and Ginnir, Sinana and Madda Walabu district administrations to get formal permission. Informed consent was obtained from each interviewee. Written informed consent for participation was not required for this study in accordance with the national legislation and the institutional requirements.AH, MK, JB, AL and SN developed the concept, developed methods, collected data and analyzed them, and drafted and edited the manuscript. All authors contributed to the article and approved the submitted version.


The widespread use of screening mammography and breast ultrasound has enhanced early diagnosis and reduced breast cancer mortality. At the same time, this practice has led to an increase in breast biopsies for suspicious lesions, some of them ultimately resulting in breast lesions of uncertain malignant potential (B3 lesions). The correct management of B3 lesions is controversial, but surgical excision is generally recommended because of the considerable risk of an upgrade to cancer diagnosis upon final histology. Very little data exist concerning the role of B3 diagnosis as a risk factor for future development of breast cancer. The current study analyzes the largest series of B3 lesions from a single institution, providing new insights on both the risk of immediate upgrade to cancer and the subsequent risk of cancer development during the follow-up. An upgrade to carcinoma was found in 4.8% to 39.8% of B3 lesions depending on histologic subtype and in 22.7% on average. In the subsequent years, a diagnosis of breast carcinoma occurred in 9.2% of the patients. This information has considerable implications for future management of patients with B3 lesions.p < 0.0001). These data support the idea that B3 lesions should be removed and provide evidence to suggest annual screening mammography for women after a B3 diagnosis because their BC risk is considerably increased.Breast lesions of uncertain malignant potential (B3) are frequently diagnosed in the era of breast cancer (BC) screening and their management is controversial. They are generally removed surgically, but some international organizations and guidelines for breast research suggest follow-up care alone or, more recently, propose vacuum-assisted excision (VAE). The risk of upgrade to BC is known, but very little data exist on its role as risk factor for future BC development. We analyzed 966 B3 lesions diagnosed at our institution, 731 of which had long-term follow-up available. Surgical removal was performed in 91%, VAE in 3.8%, and follow-up in 5.2% of cases. The B3 lesions included flat epithelial atypia (FEA), atypical ductal hyperplasia (ADH), lobular intraepithelial neoplasia (LIN), atypical papillary lesions (PLs), radial scars (RSs), and others. Overall, immediate upgrade to BC (invasive or in situ) was 22.7%. After long-term follow-up, 9.2% of the patients were diagnosed with BC in the same or contralateral breast. The highest risk was associated with ADH diagnosis, with 39.8% of patients upgraded and 13.6% with a future BC diagnosis ( Breast cancer (BC) is the most frequent cancer among women with an increasing trend of incidence; however, mortality rates are slightly decreasing, even though it is still the leading cause of death among women due to oncologic pathology . In receB3 is a group of non-malignant breast lesions with a heterogeneous risk of concurrent or subsequent breast cancer diagnosis over time . B3 lesiAt present, the management of B3 lesions is still controversial since underestimation rates for percutaneous core needle biopsy for the B3 category are high overall , so it iThe aim of our study was to clarify the clinical meaning of breast lesions of uncertain malignant potential and to provide a contribution toward understanding the best approach to treatment and follow-up of these lesions. With this purpose, we assessed the risk associated with the diagnosis of a B3 lesion after core needle biopsy (CNB) or vacuum-assisted breast biopsy (VABB) in our monocentric experience in the last 20 years, both in terms of immediate upgrade to BC and future risk of BC development.From February 2001 to December 2019, we retrospectively reviewed CNBs and VABBs performed in our institution and included those with a histological diagnosis of B3. Out of a total of 29,696 breast biopsies, we identified 1182 patients with the diagnosis of a B3 lesion. We then excluded 216 patients who were referred to other institutions for surgical treatment or who had incomplete information, resulting in a final study population of 966 patients with B3 lesions.Ultrasound (US)-guided CNBs were performed with a 14-gauge semi-automated biopsy gun , with a mean of 3 core samples obtained per lesion (range 3–6).Tomosynthesis-guided VABBs were performed using an 11-gauge or 8-gauge needle  with the patients in a prone position , obtaining a mean of 9 core samples per lesion (range 8–12).A titanium marker clip  was positioned at the end of the VABB procedure to mark the site of the biopsy.The histologic reports after biopsy with a diagnosis of a B3 lesion in our study included flat epithelial atypia (FEA), atypical ductal hyperplasia (ADH), atypical lobular hyperplasia (LIN1) and classic-type lobular carcinoma in situ (LIN2), that in the subsequent text will be collectively referred to as LIN, phyllodes tumors (PTs), papillary lesions (PLs), radial scars (RSs) and a group of rare histologic subtypes  .We retrospectively evaluated the mammograms and US exams performed at the time of the diagnosis and collected information on clinical suspicion and radiological suspicion according to the Breast Imaging Reporting and Data System (BIRADS) .We also collected patient data from medical records, including the type of subsequent management after diagnosis .We used final histology after surgery or VAE, when present, as the gold standard, considering as an upgrade the presence of invasive carcinoma or in situ carcinoma of the breast in the final specimens. We then evaluated the upgrade risk for each subgroup of B3 lesions.After multidisciplinary team discussion for each of these patients, a mammogram and an ultrasound scan of the breast were performed every 12 months during the follow-up period. All the patients gave written informed consent for the use of their data for study purposes. All patient data were anonymized.We retrospectively reviewed all follow-up exams, mammograms, and US imaging, and when present we reported the diagnosis of a subsequent breast cancer if histologically proven. We then analyzed the risk of developing a subsequent cancer in the homolateral or contralateral breast for each B3 lesion subtype.Along with the occurrence of a diagnosis of breast cancer in the years of follow-up, for each histotype we evaluated the median age of patients, the initial median radiological suspicion according to BIRADS classification, the type of radiological lesion and its size, and the median follow-up time.Data analysis was performed using IBM SPSS Statistics, version 25.0. The frequency distribution was assessed using the chi-square test. The association between histologic diagnosis of B3 lesions, the degree of radiological suspicion, breast density, the type of treatment for the B3 lesion, and the risk of subsequent breast cancer diagnosis was investigated by multiple regression analysis. The interval between the diagnosis of each type of B3 lesion and the subsequent diagnosis of breast cancer was calculated according to the Kaplan–Meier method, and the differences were evaluated by the log-rank test.The study included 966 patients with a histologic diagnosis of a B3 lesion after CNB or VABB, with a median age of 51.9 years (range 18–91) observed over a period of 19 years. Breast density, classified according to the BIRADS lexicon , was disRegarding the type of biopsy, 484 (50.1%) were US-guided CNBs while 482 (49.9%) were stereotactic or tomo-guided VABBs.The distribution by histologic subtype of B3 lesions is indicated in p = 0.004), as shown in Regarding management after biopsy, 879 (91%) B3 lesions underwent surgical OE, 37 (3.8%) VAE, and 50 (5.2%) follow-up. The type of management differed significantly by histotype (p < 0.001). Upgrade rates of B3 lesions to in situ carcinoma and to invasive carcinoma after open excision or VAE are shown in detail in ADH was the histologic type with the highest risk of upgrade (39.8%), followed by papillary lesions (24.4%). Overall, the differences in upgrade rate by histotype were statistically significant .Out of 731 women, a total of 67 (9.2%) developed a subsequent cancer in the homolateral or contralateral breast in the follow-up period. p = 0.814) was found in the risk of developing a subsequent cancer in terms of the type of management of the B3 lesion , as shown in No statistically significant difference (p < 0.01), whereas the degree of radiological suspicion, breast density, and the type of treatment for the B3 lesions were not.We then performed a linear regression analysis using future diagnosis of breast cancer as the dependent variable. Histology of the B3 lesions was the only significant independent predictor of future breast cancer development , and the median age of this group was 48.7 years (range 33.5–80.0 years).The initial radiological findings were 82 microcalcifications, 28 masses, and two architectural distortions; the mean radiological suspicion level was BIRADS 3 (range 3–5). Illustrative radiologic and histologic examples of FEA are shown in Of the 112 FEA patients, 8 (7.3%) developed subsequent breast cancer during follow-up, 2 (25%) in the same breast as the FEA and 6 (75%) in the contralateral breast.The median time interval from first diagnosis of a B3 lesion to subsequent diagnosis of BC was 53.5 months.Of the 731 women, 200 had an initial diagnosis of ADH (27.4%), and the median age of this group was 54.5 years (range 17.5–86.0 years).The initial radiological findings were 114 microcalcifications, 84 masses, and two architectural distortions; the mean radiological suspicion level was BIRADS 4 (range 3–5). Illustrative radiologic and histologic examples of ADH are shown in Of the 200 ADH patients, 27 (13.6%) developed subsequent breast cancer during follow-up, 18 (66.6%) in the same breast as the ADH and 9 (33.3%) in the contralateral breast.The median time interval from first diagnosis of a B3 lesion to subsequent diagnosis of BC was 32 months.In total, 186 women had an initial diagnosis of LIN (25.4%), and the median age of this group was 52.7 years (range 33.7–83.5 years).The initial radiological findings were 111 microcalcifications, 68 masses, and seven architectural distortions; the mean radiological suspicion level was BIRADS 4 (range 2–5). Illustrative radiologic and histologic examples of LIN are shown in Of the 186 LIN patients, 17 (8.8%) developed subsequent breast cancer during follow-up, 11 (64.7%) in the same breast as the LIN and 6 (35.3%) in the contralateral breast.The median time interval from first diagnosis of a B3 lesion to subsequent diagnosis of BC was 50.6 months.In total, 93 women had the initial diagnosis of a PL (12.7%), and the median age of this group was 55.7 years (range 18.0–91.3 years).The initial radiological findings were two microcalcifications, 90 masses, and one architectural distortion; the mean radiological suspicion level was BIRADS 4 (range 2–5). Illustrative radiologic and histologic examples of atypical papillary lesions are shown in Of the 93 PL patients, 9 (9.7%) developed subsequent breast cancer during follow-up after a median time interval of 51.1 months; in 7 (77.7%) cases the new cancer was in the same breast as the PL and in 2 (22.2%) cases the cancer was found in the contralateral breast.In total, 36 women had the initial diagnosis of a PT (7.2%), and the median age of this group was 42.51 years (range 13.6–79.3 years).All 36 PTs were reported as masses in mammograms and US images; the mean radiological suspicion level was BIRADS 3 (range 2–5). Illustrative radiologic and histologic examples of phyllodes tumors are shown in Of the 36 PT patients, 2 (5.6%) developed a subsequent breast malignancy after a median time interval of 100.7 months; all of them were malignant phyllodes tumors, occurring in the same breast as the PL.In total, 103 women had an initial diagnosis of RSs (14%), and the median age of this group was 47.9 years (range 24.9–81.7 years).The initial radiological findings were three microcalcifications, 58 masses, and 42 architectural distortions; the mean radiological suspicion level was BIRADS 4 (range 2–5). Illustrative radiologic and histologic examples of RSs are shown in Of the 103 RS patients, 4 (3.9%) developed subsequent breast cancer during follow-up after a median time interval of 73.9 months. In three cases (75%) the cancer was found in the same breast as the RS, and in one case (25%) it was found in the contralateral breast.p < 0.001; log-rank test), as shown in We then performed a Kaplan–Meier estimation of the cumulative long-term risk of breast cancer associated with each type of B3 lesion. ADH was associated with the highest risk of BC diagnosis, followed by LIN and FEA. The differences were statistically significant .In our series of 966 B3 lesions histologically diagnosed at our institution via CNBs or VABBs, we found an overall upgrade rate of 21.8% to BC (invasive or in situ) after OE or VAE, and this result is concordant with previous studies ,13.ADH histotype showed the highest upgrade risk (39.8%), followed by PLs (24.4%) and LIN (19.9%); the upgrade rates were concordant with data from the literature for ADH  and LIN The most important finding of our study was that the overall rate of developing subsequent BC during follow-up after diagnosis and excision of a B3 lesion was 9.2%. This result provides evidence that women with a B3 lesion diagnosis should be defined as “high-risk”. In fact, recent studies ,19 evaluTo our knowledge, only one previous study analyzed the benefit of long-term clinical and radiological follow-up of surgically excised B3 lesions . HennessOur results are consistent with the study of Hartmann et al. , in whicIt is known that after a diagnosis of ADH, and especially of LIN, the risk of developing BC is increased in both breasts and not just at the site of the B3 lesion . This trOverall, in our study of 67 BC cases diagnosed during follow-up, 43 (64%) were homolateral to a prior B3 lesion, whereas 24 (36%) were contralateral; thus, our data support the role of uncertain malignant potential lesions of the breast as a risk factor for BC, irrespective of the side of the original lesion. Therefore, our results suggest that after diagnosis of a B3 lesion, bilateral radiological and clinical follow-up should be warranted.In the current series, we found no significant differences in the future risk of developing breast cancer during follow-up independent of management type . However, this result does not mean that surgical removal of B3 lesions may be omitted. In fact, patients submitted to OE were the large majority of our sample (91%), while just 5.2% underwent follow-up and 3.8% VAE; therefore, there is not enough statistical power to detect differences between the groups, as presented in The strength of this study is that it is based on analysis of by far the largest series of B3 lesions ever published, while also including the longest follow-up period. This allowed us to accurately assess the risk of immediate upgrade to BC associated with the different subtypes of B3 lesions, as well as to shed light on the future risk of breast cancer diagnosis among women with a previous B3 lesion after core biopsy. Our study has some limitations. First, as stated above, there were discrepancies in the type of management after diagnosis. Second, the relatively extended period required for collection of data may have partially affected our results, since many improvements in sampling techniques and in diagnosis methods have taken place in recent years.Nonetheless, we believe this is a real-world study that offers a robust contribution to understanding the role of B3 lesions as a risk factor for concurrent or future diagnosis of BC.Diagnosis of a B3 lesion was associated with an overall rate of upgrade to BC of 22.7% after surgical removal. In addition, with a long-term follow-up, our study showed that after histological diagnosis of a B3 lesion, 9.2% of women subsequently developed breast cancer either homolaterally or contralaterally. The highest risk was associated with ADH diagnosis, with up to 39.8% of cases immediately upgraded to BC and 13.6% of women developing BC in subsequent years.Our data support indications for the surgical removal of B3 lesions and provide evidence to suggest that a clinical and radiological follow-up with a yearly bilateral mammogram and US scan of the breast should be proposed for at least 5 years. This approach may possibly translate to earlier diagnosis of BC among patients with previous B3 lesion excision, with clear clinical benefits for these patients.