As surgeons who care for patients with breast cancer, the possibility of a cancer diagnosis being related to a hereditary predisposition is always a consideration. Not only are we as surgeons always trying to identify these patients and families but also we are often asked about a potential hereditary component by the patients and their family members. It is therefore critical that we accurately assess patients to determine who may benefit from genetic testing. Importantly, the potential benefit for identifying a hereditary breast cancer extends beyond the patient to other family members and the risk may not be only for the development of breast cancers, but for other cancers as well. This review was written from the perspective of a surgeon with additional training in cancer genetics in an effort to provide a unique perspective on the issue and feel that a review of some of the more practical considerations is important. As surgeons who care for patients with breast cancer, the possibility of a cancer diagnosis being related to a hereditary predisposition is always a consideration. Not only are we as surgeons always trying to identify these patients and families but also we are often asked about a potential hereditary component by the patients and their family members. It is therefore critical that we accurately assess patients to determine who may benefit from genetic testing. Importantly, the potential benefit for identifying a hereditary breast cancer extends beyond the patient to other family members and the risk may not be only for the development of breast cancers, but for other cancers as well. This review was written from the perspective of a surgeon with additional training in cancer genetics in an effort to provide a unique perspective on the issue and feel that a review of some of the more practical considerations is important.Hereditary cases of breast cancer have long been recognized to occur, but the ability to test for a marker of risk is a relatively recent occurrence. The first study demonstrating evidence of an autosomal dominant pattern of inheritance for breast cancer was published in 1988 , 3. ShorThere are many instances where a patient’s risk can be assessed. Certainly, when assessing a newly diagnosed breast cancer, the age of onset of the cancer, specific biomarkers, and family history assessment may indicate concern for hereditary risk. Identifying unaffected individuals with hereditary risk is also of critical importance, whether they are identified due to a known genetic mutation in a family member or due to suggestive family history. The US Preventive Services Task Force has published recommendations regarding risk assessment, genetic counseling, and genetic testing for BRCA-related cancers in women .It is well established that when evaluating a new patient, a family history is obtained, but often many of the important features necessary to identify a family at hereditary risk are not assessed. When assessing a family for hereditary risk, a complete history should be obtained to include three generations and note all cancer diagnoses and ages of diagnosis. The comprehensiveness of family cancer history assessments and cancer risk assessment in primary care has been investigated by Murff and colleagues , 7. In tIn an elegant study by Burke and colleagues, primary care physicians were presented three different unannounced standardized patient case scenarios , and at the conclusion, were asked to complete a brief detection questionnaire. Similar deficits in completeness of family history were identified, and again age of onset was often not assessed. Physicians did least well identifying risk due to paternal family history and some physicians’ comments ascribed less significance to the paternal family history of breast cancer than the maternal family history. Many did elicit or recognize the important contribution of ovarian cancer to hereditary breast cancer risk assessment . This stEven in specialized clinics, in more contemporary settings, identification of high-risk patients has been difficult. Ow and colleagues retrospectively reviewed over 6000 pedigrees at their center in Singapore over a 10-year period to identify those with over a 10% risk of having a BRCA mutation. Of the 615 identified, 506 had medical records available for review, and they found that a good family history spanning three generations was documented in only 54% and was taken less frequently than smoking and drinking history. On univariate analysis, they found that taking a good family history and young age of onset of cancer increased the likelihood of physician suspicion .Patient issues also play a role in failure to identify patients at risk for hereditary cancer. Even when the appropriate questions are asked, the quality of family history data is dependent on the accuracy of patient reporting. In a study of over 43,000 women presenting for screening mammogram, Ozzane and colleagues reviewed the reported numbers of breast, colon, prostate, lung, and ovarian cancers and compared those reported in the maternal and paternal relatives and in first- and second-generation relatives. Self-reported family histories were significantly influenced by bloodline and degree of relative affected, with lower reporting of all cancers with the exception of prostate cancer in the paternal lineage and in second degree relatives .Once an accurate personal and family history has been obtained, the issue of genetic testing can be explored, but when should the test be ordered, who should order the test and how should results be communicated? Individuals at risk for hereditary cancer can be identified at multiple different times. They may be identified after diagnosis and definitive treatment of breast cancer, after diagnosis but before definitive treatment, and when unaffected. Each scenario presents its own set of challenges and different approaches to the patients with respect to counseling, testing, and risk management are required . Since tFor unaffected individuals at risk, positive results will help to quantify risk. Generally, options for management of risk include surveillance with regular mammography and clinical examination and in some cases, MRI, chemoprevention, and risk-reducing surgery. Negative genetic test results, however, may not completely eliminate risk. Without testing an affected individual, it is not clear if a negative result indicates population risk, because the mutation in the family was not inherited, or persistent risk, because the genes tested for are not responsible for the cancers in the family. For those with a new diagnosis who have not yet had definitive surgery, genetic testing results may influence surgical decisions. For example, if a patient tests positive for a mutation in one of the BRCA genes, she may choose to have bilateral mastectomy due to increased risk of contralateral cancer compared to non-mutation carriers. Identification of other mutations less well described than BRCA1 and 2 may not influence surgical decisions, since less information regarding contralateral cancer risks is known. The timing necessary for counseling and testing in the perioperative period may lead to delays in care or may cause additional anxiety if results are not obtained prior to surgery. In our institution, surgical decision-making cases are generally seen within 1 week, but extent of testing will impact timing of results availability. If only BRCA testing is performed, results are generally available within 1 week, but if a panel of genes is tested, results can take 4–6 weeks.Regardless of the timing of testing, the growing complexity of genetic testing options, including testing for panels of genes for which less information is available, make pre-test and post-test counseling important components of the process . Pre-tesWhen pre-test counseling is performed, it is generally performed by a genetic counselor, medical geneticist, surgical or medical oncologist, or other health professional with expertise and experience in cancer genetics. Some insurance companies, including Cigna and United Health Care, require that counseling be provided by a certified genetic counselor prior to testing. The American Society of Clinical Oncology recently published a policy statement update on genetic testing for cancer susceptibility. The recognition and management of individuals with an inherited susceptibility to cancer is considered a core element of oncology care, and as such, continued education of oncologists and other health professionals in the area of cancer risk assessment and management of individuals with an inherited predisposition to cancer is recommended . The keyCragun and colleagues assessed the potential differences in genetic counseling and testing services between board-certified genetic health-care providers and non-genetic health care providers by surveying patients who had undergone testing. They evaluated patient recall and content of precounseling for hereditary breast and ovarian cancer and whether BRCA1 and 2 gene sequencing was performed when less expensive single site or Ashkenazi Jewish founder mutation testing may have been sufficient. They found that genetic health-care provider involvement was associated with not only adherence to nationally recommended genetic counseling practices but could also reduce cost of testing . VadaparTesting of affected individuals who have already been treated for their cancer is routinely performed and is historically the way testing was initiated in families with hereditary risk. With the growing public knowledge regarding hereditary forms of breast cancer, newly diagnosed patients who have not yet had treatment often wonder about their hereditary risk and contemplate testing. Testing could be considered once the primary tumor has been treated, but is often being considered prior to definitive therapy. Concern exists regarding the appropriateness of genetic counseling and testing in newly diagnosed patients who have not yet had definitive surgery. This testing may have important implications for surgical decision making, including consideration of contralateral prophylactic mastectomy or the use of certain adjuvant systemic therapies, such as PARP inhibitors and platinum compounds , 20, butFor unaffected women with a strong family history of breast cancer, genetic counseling is also valuable. A Cochrane review of genetic risk assessment for individuals at risk of familial breast cancer was published in 2012, and although this review found favorable outcomes for patients after risk assessment, there were too few papers to make any significant conclusions about how best to deliver services . Fear ofOnce counseling is completed, the genetic testing options should be considered, and risks, benefits, and limitations of the testing options discussed. Once this is complete, testing can be ordered and a plan should be made for disclosure of results. The options for testing have expanded in the last 2 years for two major reasons. The patent held by Myriad Genetics was overturned, enabling other laboratories to offer BRCA testing and newer technology (next generation sequencing) became available, allowing more extensive testing from a single blood sample. Testing options currently include testing only for BRCA1 and 2 mutations, testing for a panel of high-risk breast cancer genes, including BRCA1 and 2, and more extensive panels including moderate risk genes and limited evidence genes. Testing with a panel seems most appropriate when there is concern for more than one hereditary condition. For example, in a family with early onset breast cancer (under the age of 35), ovarian cancer, and early onset colon cancer, panel testing would be ideal, since potential responsible genetic syndromes include Hereditary Breast and Ovarian Cancer Syndrome, Li Fraumeni syndrome, and Lynch Syndrome.Results interpretation is not always straightforward. For the high-risk genes, cancer risks and management options are fairly well established. The moderate risk genes have generally been seen more frequently in individuals with a strong family history of breast cancer in whom BRCA testing was negative, but the exact cancer risks and management options are not clear. It is also not clear whether these gene mutations are sufficient to explain the family history themselves or if, more likely, other modifier genes impact the likelihood of cancer development. It is therefore still debated among genetic professionals whether or not predictive testing of unaffected individuals is appropriate. In addition, a number of different testing results are possible. It is possible that a deleterious mutation in a well-described gene could be identified. This would mean that the affected individual is at increased risk for the development of cancer and that family members are at risk. The result could be negative for a deleterious mutation, but the implications of this negative result will vary based on the individual tested and the family history. An unaffected individual who tests negative for a known mutation in a family can be reassured by these negative results, as their cancer risks will be more closely aligned with population risk. Results can also be uninformative. An affected family member who tests negative for only BRCA1 or BRCA2 may have another hereditary condition not tested for. In an unaffected person with strong family history and no prior testing in the family, risk may still be elevated based on family history, but could be reduced to population if another family member was found to carry a mutation. An even more potentially confusing result is a variant of uncertain significance, or VUS. These variants are usually relatively minor changes in the genetic code for which insufficient evidence exists to determine their relevance. Decisions about risk management should not be based on these results, but rather on the family history. These unclear results may be eventually reclassified by tracking their presence with those affected with cancer in the family or with further functional studies, but this process can take years. In addition, since the contribution of these changes to cancer risk are not clear, predictive testing is not recommended for other at risk individuals. This is an area of potential harm to patients, since misinterpretation of these results could lead to unnecessary risk reduction strategies, including unnecessary surgeries. Patients who are found to have a VUS are more likely to have residual cancer distress than patients who have a negative but uninformative result, but genetic counseling has been shown to help with their understanding of their and their family’s risk, helped them make medical decisions to assist in lowering their risk, and decreased worry .Post-test counseling should include an explanation of results, options for managing risk, as well as discussion of the implications for other family members. One of the distinguishing features of genetic medicine is that the discovery of a hereditary predisposition to cancer or other disease has implications not only for the individual tested, but also for their family members. The transmission of this information is generally accomplished by communication between the patient and each of his or her relatives. Although it is well recognized that family history impacts risk, there are barriers to patients disclosing their genetic testing results to their families. These include difficult or distant family relationships, lack of perceived usefulness, the serious nature of the message, and the concern that the message will be rejected , 29. In How can we as providers guide patients and facilitate intrafamilial communication? McClellan and colleagues conducted an exploratory study to assess resources to support patients and health-care providers, specifically addressing intrafamilial communication of cancer risk . ResourcA randomized clinical trial conducted in the Netherlands assessed the use of the Informing Relatives Inventory, a battery of instruments measuring knowledge, motivation, and self-efficacy regarding disclosure of hereditary information to at-risk relatives, and found that it improved insight into patients’ barriers regarding result disclosure . FurtherEnsuring that family members are made aware of the risk is not always easy, as patients are not legally required to disclose medical information to their relatives. In addition, the Health Insurance Portability and Accountability Act (HIPAA) prohibits health-care professionals from disclosing a patient’s information to third parties, including family members. When communication between patient and relatives fails, what is the responsibility of the provider? In some instances, the law mandates release of medical information to third parties , but to date, genetic testing results have not been considered exempt from privacy laws. Even if not subject to HIPAA laws, disclosing genetic test results to all family members of an affected individual places an unrealistic burden on the practitioner. So while it is acknowledged that health-care providers have a duty to inform patients about the implications of test results for the family and should encourage the dissemination of this information, it is currently beyond the scope of practice for most providers. Certain circumstances may provide additional challenges. For example, what if the individual tested is deceased? What about children relinquished to adoption? Further discussion is necessary to determine the best way to assure that those who benefit from learning the family’s results can obtain the information necessary to address their own surveillance needs.There has been tremendous growth in the ability to test for potential hereditary causes of cancer, but the testing to date has largely been guided by family history. As we continue to personalize cancer care, BRCA testing or testing for other genes, will likely be performed for all patients newly diagnosed with breast cancer. There are already some therapeutic implications. Poly ADP-ribose polymerases (PARPs) are important components of the base excision repair pathway for single strand DNA breaks. PARP inhibitors, therefore, block this mechanism of DNA repair. These agents have shown the most promise in patients with breast or ovarian cancer who have germ line BRCA mutation or non-mutational functional defects in BRCA proteins , 35. AloSome argue that waiting until the family meets criteria for genetic testing is too late to identify a mutation in the family. The ideal time to identify risk is prior to a cancer diagnosis. A study published by Manchanda and colleagues evaluated cost-effectiveness of populations screening for BRCA mutations in Ashkenzi Jewish women compared with family history-based testing and found that this was highly cost-effective for women of Ashkenazi Jewish ancestry 30 years and older . Of courFinally, as we learn more about the implications of more moderate risk genes, the use of widespread panels will likely also increase and will likely also expand to routine testing of women. This will allow a better identification of those at risk and will likely allow better predictions of cancer risks and allow for earlier consideration of screening and preventive options.In summary, genetic testing for breast cancer susceptibility has advanced tremendously, but we are only scratching the surface of our understanding of the role heredity plays in carcinogenesis. There is no doubt, however, that our knowledge will continue to grow allowing us to more accurately provide assessment of risk to all individuals, regardless of their family history.The authors jointly conceptualized the article and discussed the aspects of the topic to be addressed. DA performed the primary literature review and first authored the paper. RP provided critical review.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Despite demonstrated efficacy, acceptance of selective estrogen receptor modulators (SERMs), such as tamoxifen, for breast cancer risk reduction remains low. Delivering SERMs via local transdermal therapy (LTT) could significantly reduce systemic effects and therefore may increase acceptance. We aim to assess women’s knowledge of breast cancer prevention medications and views on LTT of SERMs.Focus groups were conducted with healthy women identified through the comprehensive breast center of a large urban cancer institution. Group discussions covered risk perceptions, knowledge of and concerns about risk reducing medications. Participants reported their perceived risk for breast cancer , preference for SERMs in a pill or gel form, risk factors, and prior physician recommendations regarding risk-reducing medicines. Participants’ breast cancer risk was estimated using tools based on the Gail Model. Trained personnel examined all qualitative results systematically; risk perceptions and preferred method of medication delivery were tallied quantitatively.N = 32) were conducted. Most participants had at least a college degree (78.2 %) and were of European (50 %) or African ancestry (31 %). The majority (72 %) were at elevated risk for breast cancer; approximately half of these women perceived themselves to be at elevated risk. Few participants had prior knowledge of preventive medications. The women noted a number of concerns about LTT, including dosage, impact on day-to-day life, and side effects; nonetheless, over 90 % of the women stated they would prefer LTT to a pill.Four focus groups (Awareness of preventive medications was low even in a highly educated sample of high-risk women. If given a choice in the route of administration, most women preferred a gel to a pill, anticipating fewer side effects. Future work should focus on demonstrating equivalent efficacy and reduced toxicity of topical over oral medications and on raising awareness of chemopreventive options for breast cancer. Breast cancer is the most common cancer in women in the United States, with more than 232,000 cases of invasive cancer diagnosed in 2014 [Selective estrogen receptor modulators (SERMs) prescribed as preventive medications hold promise of reducing the breast cancer burden. Tamoxifen, used for both pre- and postmenopausal women, and a second orally administered SERM, raloxifene, used in postmenopausal women, are the only medications currently approved by the FDA for use in the United States in the prevention of breast cancer in women at increased risk for the disease . In the Delivering tamoxifen directly to the breast tissue via the skin, i.e., local transdermal therapy (LTT), is a novel approach with anticipated advantages of reducing or possibly eliminating systemic toxicities associated with oral therapies and avoiding individual variations in drug metabolism that could alter drug effectiveness. A small phase II study demonstrated that an active metabolite of tamoxifen, 4-Hydroxytamoxifen (4-OHT), applied to the skin of the breast as a gel, reduced breast tumor cell proliferation to the same extent as oral tamoxifen , 15. We From this early experience, we conclude that topical application of 4-OHT offers promise as an effective preventive medication that women may find more acceptable than oral medication. As we continue efforts to develop LTT for breast cancer prevention, we must establish that these efforts will actually translate into increased acceptance of this method of drug delivery among at risk women. Therefore, we examined women’s perceptions of preventive medicines and the acceptability of a topical application of this active metabolite of tamoxifen among a sample of high risk and average risk women without a prior breast cancer diagnosis.Because views of preventive medications may vary by a women’s risk of developing breast cancer, we included women at average and elevated risk of breast cancer. For recruitment purposes, elevated risk was defined as risk greater than that of the average-risk peer of the same age or by virtue of possessing one or more of the following conditions: a) a 1st or 2nd degree relative with breast cancer diagnosed at any age; b) multiple prior breast biopsies regardless of histology; c) prior or current evidence of breast epithelial atypia (histologic or cytologic); d) estimated mammographic density on visual inspection of BIRADS 3 or 4; e) known carrier of BRCA 1 or 2 mutation; and/or f) prior history of chest wall radiation. Women were considered average risk if they possessed no more than one of the following characteristics: a) no family history of breast cancer in a first or second degree relative; b) one prior breast biopsy without atypia, and c) BIRADS 1 or 2 estimated mammographic density on visual inspection. All participants were female age 21 or older.Participants were identified through the Bluhm Family Program for Breast Cancer Early Detection and Prevention at Prentice Women’s Hospital, Lynn Sage Comprehensive Breast Center. Eligible women received a phone call from the study coordinator inviting them to participate in a focus group about women’s views of their breast cancer risk and methods to prevent breast cancer.Focus groups were held in Prentice Women’s Hospital, part of the Chicago campus of the Feinberg School of Medicine of Northwestern University. An experienced focus group moderator led consented participants through a discussion of breast cancer risk perceptions and knowledge of and concerns about risk reduction medications. Each woman was asked to characterize her own risk as below average, average, or above average as part of the discussion. After discussing knowledge and concerns of risk reduction medications, the moderator provided a brief overview of LTT; the women then discussed their views of the acceptability of a skin application of risk reduction medication. Focus group sessions lasted approximately 60 min. The discussions were audiotaped and transcribed. At the conclusion of the group, each participant completed a short survey to provide sociodemographic information, breast cancer risk factors , and whether a physician had ever suggested they take medication to reduce their breast cancer risk. At the completion of the group, participants were given a handout providing information on the two FDA approved medications currently in use for breast cancer prevention. The moderator provided information to participants at the end of the focus group discussion, which addressed their questions. These included the instructions for application, odor, and consistency. Participants received $50 compensation for their time and a voucher to cover parking expenses in a Northwestern University parking structure. Following completion of the focus groups, participants’ medical histories  were obtained via medical record review. The Northwestern University Internal Review Board approved all procedures.Focus group transcripts were content analyzed by the first and last author. First, the number of women in each group with prior knowledge of risk reduction medications was tallied. Next, thematic analysis identified concerns about preventive medicines and concerns about a topical preventive medication. Concerns were identified and tallied within and across groups. The analysts compared their results and any differences were discussed and resolved. Each participant’s breast cancer risk was calculated using the Breast Cancer Risk Assessment Tool provided by the National Cancer Institute and the Detailed Breast Cancer Risk Calculator created by Halls, both are based on the Gail Model. Participants were classified as below, at, or above the average risk for women of their age , 18. ParN = 4). Twenty-three women were above peer average, 3 were at the peer average, and 6 were below peer average.Four focus groups were conducted with 32 women. A summary of participant sociodemographic characteristics, breast cancer risk factors, and knowledge of preventive medications is shown in Table When comparing women’s perceived breast cancer risk with Gail Model estimates, women correctly assessed their own personal risk 50 % of the time Fig. . Women oWhen asked what drove their risk perceptions, women most often cited family history. For example, one woman who perceived herself as above average yet was calculated to be at average risk stated, “I think I have a greater risk because of a couple of factors, number one is family history of breast cancer.” Other factors that shaped women’s perceived risk included obesity, poor diet, smoking, stress, environmental pollution/toxins, hormone replacement therapy, nulliparity, breast augmentation, abnormal mammogram, dense breast tissue, history of breast biopsy and genetic mutations.Very few of the focus group participants were aware that medications existed to prevent breast cancer. One woman mentioned the drug Evista, stating that she knew “some people use it for osteoporosis and they say it helps reduce your risk for breast cancer, too.” Another woman admitted that she had heard of tamoxifen but when asked what she knew about the medication she stated, “There are some not great side effects…I mean, it sounds a lot like menopause.” Others who knew of tamoxifen were not aware of its application to disease prevention and thought it was only indicated for treatment of disease. “We hear a lot about what’s going on in the research world and tamoxifen…and just that it’s out there, but I don’t know anything, specifics about reduction of rates, or side effects, or anything like that.” Even those familiar with the medication had limited knowledge of its purpose. By comparing women’s focus groups comments about preventive medications with their risk perceptions, we ascertained that participants who perceived themselves to be at high risk for breast cancer tended to be more knowledgeable of preventive medications than those who did not perceive themselves to be at elevated risk, 38 % vs 0 % respectively. Moreover, women with awareness of preventive medications tended to be older and had either a first-degree relative with the disease or had undergone a breast biopsy.“I think the side effects are traumatizing relative to what you’re treating. I mean just like incontinence and like all this other stuff that comes up (from taking various medications) that’s not even related to what it’s treating…so you kind of wonder, why? Why would you want to take this thing that could create these six other problems?”“Anything I have to take for something that I am treating, I can deal with what I have to deal with, but just as far as on the level of prophylactics, I would want it to be pretty simple that way.”“I’d like to know the risk benefit analysis on an individual basis, what is the risk of not taking the medicine versus the risk of taking the medicine. So how elevated would my risk be for breast cancer if I didn’t take it, and what are the benefits if I did take it, and then look at the side effects.”“I think probably the cost as well, I mean, don’t want to send everybody to poor house… or myself trying to prevent something”“I’ve never been a pill person, so I’ve never wanted to, I wouldn’t want to start anything that I’d have to continue”“I think that apart from the side effects, I would also want to know the ease of taking the medicine, because I hate taking pills and I gag and choke even when it’s a tiny pill.”When prompted to list concerns they would have if their doctor recommended that they take preventive medicine, women in every group wanted to know about side effects of the medicine. Further, women stated that they would want their physician to give them an individualized risk/benefit analysis and demonstrate that they would see a significant risk reduction on the drug. A few women voiced reluctance to the idea of taking medicine for any indication and claimed that it would take a lot of convincing and evidence provided to them by their physician before they would consider accepting a drug. Duration of therapy, interactions with other medications or products, cost, and whether medicines used for prevention would be covered by insurance were other major concerns. Also, several women noted fears of taking pills, and in some cases, difficulty swallowing them. The following comments are representative of their concerns:Although women had a number of reservations about preventive medications, they were more open to the idea of a gel application of the medicine. Over 90% of focus group participants stated they would prefer the skin application of preventive medicine over a pill application if a physician advised them to take preventive medication Table . Of the Focus group participants mentioned a number of concerns they would want to discuss with their physician prior to taking a topical preventive medication, including the impact of the gel on day to day life, dosage, characteristics of the gel, side effects, and whether the gel posed a threat to others. Women in every group questioned whether they could continue with normal activities  while using the gel. They were also concerned about the gel staining their clothes, whether they would have to wait to get dressed after applying the gel, and how soon after application they could bathe.The women raised questions about the dosage of the gel: How long would they take it? How much gel would they apply each day? Does it need to be applied at the same time each day? They questioned whether a gel could be dosed properly, particularly given varying breast sizes among women. In addition, they wanted to know more about the characteristics of the gel, including its texture, scent, packaging, and storage requirements.Side effects were also a concern. The women wondered about possible negative effects on hands, impact on organs near the application site , and changes to the texture of the breast skin. One woman remarked, “I would look at those side effects really, really carefully and probably consult other doctors who could either put my mind at rest or convince me not to take it.” A 45-year-old woman who perceived her risk of breast cancer to be above average said, “I mean I don’t know what the risks are, but I would want to know the quality of life I would have after taking that, you know, just for preventative, in the event that there may not even be a chance that I’d… develop breast cancer.” Likewise, women wondered whether the gel posed a threat to others who touched it or touched a woman’s skin after she used the gel. For example, a participant questioned “Do you apply this with a bare hand? Then it’s on my hand and I’m doing dishes or cooking or putting a lunch together, you know, where does, how is that transferring?” They were also concerned about the impact of the gel on intimacy and on a woman’s ability to breastfeed. Women also noted concerns about cost and insurance coverage. One woman commented, “…I don’t know if I’m going to be committed to buying a pharmaceutical daily to prevent something that I may or may not get, you know, but if it’s once a year and it’s included in my health plan, and I don’t have to pay, you know, a whole bunch for it.” A number of other concerns were also mentioned, including interactions with other lotions or foods/beverages a desire to see data to support the gel’s efficacy.Our data, from a highly educated group of healthy women at varying levels of perceived and calculated breast cancer risk, demonstrate surprisingly low awareness of breast cancer preventive medications. This was despite their high educational attainment, adherence to screening and motivation to seek information about breast cancer detection and prevention —100 % of the women over 40 had a screening mammogram and 100 % of those younger than 40 had attended an appointment in the comprehensive breast center to have a clinical breast exam and to learn more about their own risk. We have evaluated, in a rigorous and structured manner, the preference of this group for the mode of delivery of preventive medications for breast cancer. When preventive medications were described to these women, 91 % of women expressed preference for a skin application over an oral form. There was no relationship between calculated or perceived breast cancer risk and preference for the route of medication delivery. The number of women who accurately estimated their own risk, 50 %, was higher than most previously published studies, which report frequent over-estimation of risk, especially among women of European ancestry –21. HoweOur findings also highlight information women would want to know about skin applications of preventive medications, including their impact on day-to-day life, dosage, characteristics of the medications, side effects, and whether the topical medication posed a threat to others. Notably, many of the women’s questions about the gel, such as dosage and application, could be addressed via a brief discussion with a physician or via product packaging. Women also were concerned about the efficacy of a new medication delivery method, cost, insurance coverage, and determination of the population that would benefit most from the medicine. The general consensus of this sample of women was that if these concerns could be adequately addressed then women would be more likely to accept a medication for chemoprevention. Thus, a skin application with limited systemic effects has the potential to gain acceptance among at risk women.Our study has several limitations. The women’s preferences were based on hypothetical situations—women were not actually confronted with the option of taking a pill or gel for breast cancer prevention. Moreover, our results were based on a small sample of mostly well-educated women from one large city. Each focus group expressed many of the same concerns; thus we feel confident that we have captured the most salient concerns for these women. Only four women in our sample were under the age of 35, the group where fertility concerns are an important issue, and could be an influential factor in decisions regarding preventive therapy for breast cancer. Future work should examine how such concerns might impact the willingness to take preventive medications. Additionally, work with different populations of women may reveal other concerns about preventive medications and LTT. Our sample represents women who may be most likely to take a preventive medication because of their high risk status and overall attention to their breast health, as evidenced by their regular mammogram screening and use of clinical breast exams. In addition, our sample, which has a mean age of 48, provides insights into the views of younger women towards preventive medicine; this is particularly important as this is the group least likely to accept preventive medications .Additional studies of the long-term efficacy of LTT are needed and will increase women’s confidence in these novel medications. In addition, increasing public awareness of chemopreventive medications for breast cancer should be a goal of public health initiatives to encourage acceptance of preventive medications. Notably, the women in our study were very interested learning more about the gel, including when it would be available. Future work should also focus on establishing the side effect reduction of topical 4-OHT over oral tamoxifen. Because breast cancer chemoprevention requires a daily, long-term commitment from women, it is not surprising that many women refuse or stop taking the medication due to side effects and daily burden , 24. EveAs additional, larger studies of LTT are designed, it will be important to address issues related to non-compliance that have been identified in the therapeutic setting and may apply to the prevention setting, regardless of the mode of delivery. These include the presence of comorbidities, negative mood prior to the initiation of endocrine therapy, and greater perceived bother associated with multiple symptoms –27. SimiThe study obtained approval from the Institutional Review Board of Northwestern University. The IRB approval number is STU00079604. Although no identifying information is used, the study participants provided informed consent in the form of verbal agreement at the beginning of each focus group interview.


This study assessed Australian clinicians’ knowledge, attitudes and referral patterns of patients with suspected Lynch syndrome for genetic services. A total of 144 oncologists, surgeons, gynaecologists, general practitioners and gastroenterologists from the Australian Medical Association and Clinical Oncology Society responded to a web-based survey. Most respondents demonstrated suboptimal knowledge of Lynch syndrome. Male general practitioners who have been practicing for ≥10 years were less likely to offer genetic referral than specialists, and many clinicians did not recognize that immunohistochemistry testing is not a germline test. Half of all general practitioners did not actually refer patients in the past 12 months, and 30% of them did not feel that their role is to identify patients for genetic referral. The majority of clinicians considered everyone to be responsible for making the initial referral to genetic services, but a small preference was given to oncologists (15%) and general practitioners (13%). Patient information brochures, continuing genetic education programs and referral guidelines were favoured as support for practice. Targeted education interventions should be considered to improve referral. An online family history assessment tool with built-in decision support would be helpful in triaging high-risk individuals for pathology analysis and/or genetic assessment in general practice. MLH1, MSH2, MSH6 and PMS2. It accounts for about 5% of all colorectal and endometrial cancers diagnosed [Lynch syndrome, also known as hereditary nonpolyposis colorectal cancer (HNPCC), is an inherited cancer syndrome caused by defect in one of the mismatch repair (MMR) genes—iagnosed ,4. Furthiagnosed ,8,9,10.i.e., genetic consultation and/or testing) have been reported in the literature, including lack of knowledge regarding Lynch syndrome and who should be referred, lack of family history information or referral guidelines, or lack of awareness of clinical genetic services [Numerous barriers to referral for genetic services  or microsatellite instability (MSI) testing directly from a pathology service. Routine IHC for MLH1, MSH2, MSH6 and PMS2 proteins has been recommended and supported by the Australian College of Pathologist for all patients diagnosed with colorectal cancer younger than 50 years of age [In order to facilitate diagnosis of Lynch syndrome, various guidelines such as the Amsterdam II, revised Bethesda or the Society of Gynecologic Oncologists have been developed ,18,19. Canalysis . While tanalysis , only sps of age , althougi.e., preferred timing for referral and who they think might be the most appropriate professional to make the initial genetics referral), perceptions of their role and their desired support for the provision of genetic services.As early identification of mutation carriers allows for more intensive colonoscopic surveillance and consideration of risk-reducing surgeries ,25,26,27i.e., gynaecologists, gastroenterologists, medical oncologists, radiation oncologists, gynaecology oncologists, general and colorectal surgeons), who were identified through membership list of the Australian Medical Association. We purchased the list from AMPCo Data direct, a subsidiary of the Association who owns the list of practicing doctors , who then broadcasted our invitation to their members. An invitation was circulated by email to a randomly stratified sample of 1674 participants. Sample size was calculated to provide 90% power, with a two-sided significant level of α = 0.06. The email described the study and confidentiality, and included a link to the online survey and participant information sheet. Participants were asked to read the information brochure prior to commencing the survey. A reminder email was sent a week apart from the initial broadcast. Responders were allowed to save their responses and return to complete the survey at a later time. Due to the small sample size of oncologists from the Australian Medical Association database, oncologists were also identified from the membership of Clinical Oncological Society of Australia membership directory. An invitation email was sent to gynaecological oncologists, medical oncologists and radiation oncologists by the Society, and was followed-up with a reminder email sent a week after the initial broadcast. Only one reminder email was sent to all invited study participants. Participation was voluntary and no financial incentives were given. Survey response implied consent to participate, and all responses were anonymous. The study was approved by The University of Queensland School of Medicine and Royal Brisbane & Women’s Hospital research ethics committees.We targeted our sample to clinicians who are likely to diagnose, treat and survey Lynch families. Participants therefore included GPs and specialist groups  to assess relevance and face validity of survey items. Changes to the survey were made accordingly using the Delphi method [A web-based survey was developed specifically for this study based on a review of relevant literature ,30,31,32i method  before aThe final version of the survey was created using LimeSurvey and consisted of 19 items see . Items i® SPSS® Statistics, Chicago, IL, USA). Descriptive statistics included frequencies and proportions were calculated. x2 or Fisher’s exact test was used to investigate associations between categorical variables. Throughout the analysis, respondents who selected unsure were designated as missing data. All p-values were two-sided, with a statistical significance level set at p < 0.05.All survey data were collected using LimeSurvey, and data were coded and analysed using SPSS Version 20 (IBMp = 0.005), and were in practice for at least 10 years (p = 0.002). There were no statistically significant differences between gender or state of residence with provider groups.Of 582 email recipients who “opened” the email, 144 (24.7%) responded to the web survey fully  or partially . Demographics of these clinicians are summarized in x2 = 16; p = 0.001; 25% vs. 87%). GPs and gynaecologists were significantly less likely to order IHC testing , whereas oncologists and surgeons were more likely to order such testing . GPs and gynaecologists were also less likely to order MSI testing , but surgeons were more likely than the others to order MSI testing (p = 0.005). Oncologists were more likely than other provider groups to order DNA germline testing (p = 0.007), whereas gynaecologists were less likely to do so. There were no statistically significant differences between age, gender, state of residence or years in practice and referral to genetic services.i.e., no elevated risk for breast or thyroid cancer, somewhat higher risk for ovarian and gastric cancer, and much higher risk for colorectal and endometrial cancer as compared to the general population. Of all respondents, 65% could correctly identify colorectal cancer risk, followed by 63% for thyroid cancer, 60% for endometrial cancer, 45% for gastric cancer and 42% for ovarian cancer. About a third of all respondents considered the fictitious high-risk asymptomatic individual to be at lower risk of developing colorectal and endometrial cancer when compared to the general population, but half of all respondents considered the individual to have a much higher risk for developing ovarian cancer, and another 15% considered the individual to have a much higher risk for developing gastric cancer. A notable 44% considered the individual to have high risk for breast cancer. There were no statistically significant differences in specialist groups, age, gender, state of residence or years in practice with providing the correct answers. Nevertheless, GPs were less likely than other specialists to recognize high risk for colorectal cancer  and endometrial cancer .When participants were presented with a fictitious high-risk Lynch-specific clinical scenario , only 13p < 0.05 for both groups), while oncologists (68%) were more likely to answer the question correctly (p < 0.001) and refer patients for genetic services (p = 0.008).When asked if they consider IHC or MSI testing a germline test, 9% of all respondents thought IHC was a germline test, while another 23% considered MSI a germline test. Forty-nine percent and 35% of all respondents did not consider IHC or MSI a germline test, respectively. Forty-two percent of all respondents were uncertain about both tests. GPs (89%) and gynaecologists (75%) were less likely to answer the question on IHC correctly ; refer to a geneticist (94%); discuss Lynch syndrome cancers with the patient (82%); offer cancer surveillance (77%); order IHC or MSI tumour testing (58%); discuss risk-reducing surgeries (53%); order germline testing (42%); and refer to non-genetics specialists (38%); and no further action (1%). For those who offered cancer surveillance and discuss risk-reducing surgeries, they recommended colonoscopy (87%), gastroscopy (62%), serum CA125 (48%), hysterectomy and bilateral salpingo-oophorectomy (41%), breast ultrasound (36%), subtotal or segmental colectomy (8%), pelvic ultrasound (7%) and mastectomy (4%).p = 0.04); no knowledge of how to make referral (p ≤ 0.001); no access to a genetic health service (p = 0.001); no recommendation and guidelines for referral (p = 0.016).As shown in p = 0.031). Oncologists were preferred as they would most likely recognize at-risk individuals, whereas GPs were preferred because they are the first point of contact for patients, who have the best knowledge about patients’ family history and are the ones responsible for coordinating care. However, compared to the specialist groups, some GPs did not feel that their role is to identify patients for referral to genetic services  or to order IHC or MSI testing . There was no statistically significant difference between provider groups and preferred timing for referral. Clinicians indicated referral should be warranted at any time (51%) and when family history is collected (27%). Although a majority of clinicians felt that everyone is responsible for referring patients to genetic services, a proportion of clinicians believe that oncologists (15%) and GPs (13%) would be best suited for the role  ,51. An iet al.  involvinet al. . It is pIn our previous study, we showed that a research-based self-administered family history questionnaire can be considered for use in specialist clinics to facilitate and improve family history documentation . Indeed,There are several limitations to this study. Firstly, the response rate was low. This may be in part due to unwillingness to complete a web-based survey or lack of interest in hereditary cancer syndromes. A larger sample size and greater response rate would certainly provide a more accurate view on clinicians’ attitudes to genetics referral, however, the response rate in our study is comparable to other web-based studies conducted ,57,58,59MLH1 methylation testing to identify Lynch syndrome patients as a way to prevent second cancers, and identify carrier relatives for predictive testing.Our study suggested that genetics education is necessary for clinicians in order to improve genetic referral. Further studies should be conducted to examine educational topics selected by clinicians , and more behavioural research is needed to investigate barriers to patient uptake of referral as well as patients’ information needs. An online family history tool with built-in decision support for genetics referral would be helpful in triaging high-risk individuals in general practice. Future research should also focus on feasibility and impact assessment of reflex IHC or MSI and


Despite intensive surveillance, a high rate of interval malignancies is still seen in women at increased breast cancer risk. Therefore, novel screening modalities aiming at early detection remain needed. The intraductal approach offers the possibility to directly sample fluid containing cells, DNA and proteins from the mammary ductal system where, in the majority of cases, breast cancer originates. Fluid from the breast can non-invasively be obtained by oxytocin-assisted vacuum aspiration, called nipple fluid aspiration (NFA). The goal of this feasibility study was to evaluate the potential of repeated NFA, which is a critical and essential step to evaluate its possible value as a breast cancer screening method.In this multicenter, prospective study, we annually collected nipple fluid for up to 5 consecutive years from women at increased breast cancer risk, and performed a questionnaire-based survey regarding discomfort of the aspiration. Endpoints of the current interim analyses were the feasibility and results of 994 NFA procedures in 451 women with total follow-up of 560 person years of observation.In this large group of women at increased risk of breast cancer, repetitive NFA appeared to be feasible and safe. In 66.4% of aspirated breasts, nipple fluid was successfully obtained. Independent predictive factors for successful NFA were premenopausal status, spontaneous nipple discharge, smaller breast size, bilateral oophorectomy and previous use of hormone replacement therapy or anti-hormonal treatment. The procedure was well tolerated with low discomfort. Drop-out rate was 20%, which was mainly due to repeated unsuccessful aspiration attempts. Only 1.6% of women prematurely declined further participation because of side effects.Repeated NFA in women at increased breast cancer risk is feasible and safe. Therefore, NFA is a promising method to non-invasively obtain a valuable source of potential breast cancer specific biomarkers. TP53, PTEN, LKB1, CDH1, or, most frequently, BRCA1 or BRCA2 [BRCA1 mutation have a 57–65% chance of developing breast cancer before the age of 70 years, which for BRCA2 mutation carriers is 45–55% [BRCA1/2 mutation carriers are also at increased risk of subsequent contralateral breast cancer. This risk is estimated to be 20–60% or even higher and is influenced by various factors such as age at diagnosis and adjuvant systemic therapy [Breast cancer causes the highest cancer related mortality in women with 458,000 deaths worldwide in 2008 . MoreoveBRCA1/2 mutation carriers [BRCA1/2 mutation carriers without prior breast cancer (HR 0.36–0.63) [To date, the most effective preventive options in women at increased breast cancer risk are bilateral or contralateral prophylactic mastectomy (PM) and/or prophylactic bilateral salpingo-oophorectomy (PBSO). PM yields a risk reducing effect of more than 95% in healthy carriers . PBSO be36–0.63) . Data ab36–0.63) ,9. An al36–0.63) .BRCA1/2 mutation carriers [BRCA1 mutation carriers compared to BRCA2 mutation carriers or women with a moderate to high familial breast cancer risk [BRCA1 mutation carriers have more interval cancers (32%) and an unfavourable tumor size at diagnosis [Another option to prevent breast cancer related mortality in high risk women is intensive surveillance including e.g. mammography and MRI. In follow-up studies, annual MRI significantly reduced the incidence of advanced-stage breast cancers in carriers  and detecarriers . MRI is cer risk . Moreoveiagnosis . Other diagnosis –16.The intraductal approach offers a way to directly access or sample fluid from the mammary ductal system, where in the majority of patients breast cancer develops. Nipple fluid contains cells, DNA and proteins directly derived from the breast ducts and can thereby be a rich source of breast cancer biomarkers ,18. FluiIn the present analyses, we investigated the feasibility of and variables affecting a successful NFA procedure in a prospective, multicenter study where nipple fluid was obtained annually in women at increased breast cancer risk adhering to a surveillance program. Also, compliance and discomfort associated with the procedure was studied. The goal of this clinical feasibility study was to evaluate the potential of repeated nipple fluid aspiration, which is a critical and essential step to evaluate its possibility as a breast cancer screening method.BRCA1/2 mutation after genetic testing. Person years of observation were calculated from baseline to the last NFA procedure or last moment of contact with the participant.Women at increased risk of breast cancer adhering to a regular surveillance program were included in this prospective clinical study aiming to establish methylation profiles in nipple fluid. According to the Dutch guidelines, the surveillance program provides imaging based screening as standard of care for women at increased breast cancer risk based on inheritance or a history of breast cancer. The study design is observational: a cohort of high-risk women is being followed from baseline to the end of follow-up or until development of breast cancer or (preventive) breast surgery. Nipple fluid was aspirated annually, for a follow-up period of five years. Besides ending the follow-up period of five years, participation could be discontinued because of the development of breast cancer, breast surgery making NFA impossible, or the exclusion of a Apart from the NFA, participants had their regular follow-up consultations with the physician or nurse practitioner and their imaging examinations. The indication for breast surgery was based on usual clinical and radiological findings, and some patients opted for risk reducing mastectomy at which time the follow-up into the study stopped. Physicians and patients remained blinded to the results of the nipple aspirate analyses. Enrolment of participants started at August 7, 2008 in the University Medical Center Utrecht (UMCU) and at April 22, 2011 in the Erasmus Medical Center Rotterdam (EMC).The study has been approved by the Ethics Committees of the UMCU and the EMC, The Netherlands . Written informed consent was obtained from all participants.The objective of the present analysis was to assess the feasibility and compliance of repeated NFA, the variables predictive for a successful aspiration of nipple fluid, and discomfort experienced by participants in comparison with other surveillance procedures and breast feeding.BRCA1 or BRCA2 gene mutation, women with a pedigree-based increased lifetime breast cancer risk, or a history of DCIS or invasive breast cancer. Exclusion criteria were age below 18 years, bilateral mastectomy, pregnancy or lactation, active breast infection, and disseminated breast cancer.Women at increased risk of developing (a new) breast cancer adhering to a surveillance program were eligible for the study, including: carriers of a The technique of NFA has been previously described ,21. In sTo test the feasibility of obtaining nipple fluid with an electric pump, we also performed a pilot study in 43 women in which vacuum was applied simultaneously to both breasts using an electric device  for approximately 15–20 minutes. If after 10 minutes no droplets were seen, a second dose of oxytocin was given and the aspiration procedure was repeated.Participants received oxytocin nasal spray (Syntocinon) in a dose of 4 IU per spray in order to stimulate the production of nipple fluid. One spray of nasal oxytocin contains 4 IU and is the standard dose to induce lactation in breastfeeding women. In the mammary gland, oxytocin induces contraction of the myoepithelial cells which surround the milk-storing alveoli and in this way facilitates the release of milk from the breast during lactation . MoreoveReported adverse events in the Summary of Product Characteristics of nasal oxytocin include headache , nausea , allergic dermatitis , and uterine contractions (<1/100). Syntocinon is quickly absorbed by the nasal mucosa and effective 5 minutes after administration. In case of supratherapeutic dosing, the oxytocin will be swallowed and degraded quickly by proteolytic enzymes in the gastro-intestinal tract. Oxytocin has a half-life that varies from 3 to 20 minutes and it is mainly eliminated by the liver and the kidneys .et al. [A questionnaire addressing age, phase in menstrual cycle, menarche, menopause, use of oral anticonceptives or hormonal replacement therapy, parity, breast feeding, spontaneous nipple discharge, prior mammography, prior palpable masses in the breast, biopsy or breast surgery, oophorectomy and chemo- or radiotherapy, was filled out before the first NFA procedure, and updated at every subsequent visit. Spontaneous nipple discharge is defined as physiologic discharge which is spontaneous, usually bilateral, involves multiple ducts and does not contain blood. Around 50%-80% of women in their reproductive years can express fluid from their breasts. Spontaneous nipple discharge is different from pathological nipple discharge, which is unilateral, and can be bloody, serous, clear, or associated with a mass . A discoet al. . In ordeP-value <0.05 was considered statistically significant. To account for the clustered data, we analysed the nipple fluid aspiration procedure results using repeated measurement analysis by General Estimation Equations (GEE) with participant as the subject level and breast*visit as within-subject levels, using robust standard error estimation and accounting for within-subject dependencies assuming an autoregressive relationship. For categorical outcomes  we used a cumulative logit multinomial approach, and for success rate a negative binomial approach with a log link . The reported overall frequency of the various outcomes and the frequency of success in patient subgroups are based on the GEE-estimated values. We used a stepwise-backward multivariable selection approach for determinants of aspiration success (through P<0.1). To compare electric and manual aspiration, we made use of paired analyses from the electric aspiration compared with a manual aspiration the visit before, or after if there was no earlier visit. Cases were excluded if only one visit with electronic aspiration was performed. To analyse discomfort rates, only the first visit was taken into account.For statistical analyses IBM SPSS Statistics Version 20 was used. A two-sided At the time of analysis, a total of 994 NFA procedures had been performed in 451 women . Since the Erasmus Medical Center started including participants from April 2011, only first and second year participants of this center were yet analysed. 295 women underwent a second NFA, 139 a third, and 84 a fourth. A total of 25 women completed the follow-up period of 5 years and 88% of them were willing to continue follow-up. Taken together there were 560 person years of observation at time of analysis.Baseline characteristics and reasons for inclusion of participants are shown in The aspect of the obtained fluid was clear in 56.0%, cloudy in 31.0%, different colours from several ducts in 12.7%, and bloody in 0.3%. The estimated volume of the fluid aspirated was less than 5 μl in 37.1%, 5–50 μl in 59.6%, and more than 50 μl in 3.3% of women. In 74.8% of women 1 or 2 droplets were aspirated, in 21.2% 3 or 4 droplets, and in 4.0% 5 droplets or more.To analyse the influence of clinical characteristics on successful aspiration, analyses were done considering NFA per breast taking into account within-participant dependency between observations from each breast. NFA was performed in 1824 breasts and aspiration was successful in 66.4%. P = 0.115), and the procedure takes on average 15 minutes less than manual aspiration.In the 43 participants from whom nipple fluid was obtained with an electric breast pump, the procedure was successful in 62.3%. The success of obtaining fluid did not significantly differ between the manual or electric procedure in the same participant . In In BRCA1/2 mutation (0.7%), women underwent preventive mastectomy (2.7%), or breast cancer developed. Twelve women developed breast cancer during follow up (2.7%) and three women died from disseminated breast cancer that developed during follow-up (0.7%). True drop-out was 20.0% and mainly caused by repeated unsuccessful aspiration attempts (10.9%), or adverse events (1.6%).During follow-up, 141 women (31.3%) left the study , either In this multicenter, prospective study, we annually collected nipple fluid for up to 5 consecutive years from women at increased breast cancer risk adhering to a surveillance program. We were able to obtain nipple fluid by vacuum aspiration and using oxytocin nasal spray in 66.4% of aspirated breasts. Annually repeated aspiration was feasible and very well tolerated with the occurrence of very few and self-limiting adverse events. Success rates were higher in premenopausal women, in women with a history of spontaneous nipple discharge, bilateral oophorectomy, previous hormonal replacement therapy or anti-hormonal therapy, and in women with smaller breast size. True drop-out was mainly due to repetitively unsuccessful aspiration.Our first experience with NFA was obtained in healthy volunteers, where aspiration was successful in 94% of patients and in 84% per breast. Having spontaneous nipple discharge showed to be the only predictive factor in successfully obtaining nipple fluid. The procedure was well tolerated and no side effects from using oxytocin nasal spray were reported . After tIn order to optimize adherence to nipple fluid aspiration as a new screening tool, we tested the feasibility of an electric breast pump, allowing women to obtain nipple fluid at home, in a subgroup of women. As the success rate was comparable to manual aspiration, we are currently optimizing the procedure for at home use, which may facilitate using NFA in a wider screening setting.et al. reported that fluid yielding ducts could be identified in 36% of high risk women (N = 33), in contrast to in 84% of women without an increased risk. Reduced yield of nipple fluid was associated with postmenopausal state, BRCA germline mutation and a history of risk reducing strategies such as PBSO or use of selective estrogen receptor modulator inhibitors. The authors hypothesized that endocrine mechanisms associated with risk-reducing therapies could explain the diminished production of nipple fluid [et al. identified fluid yielding ducts in 60% of BRCA mutation carriers (N = 52) and again postmenopausal status was associated with less fluid yielding [et al. also studied women at increased breast cancer risk (N = 67), but did not include women with a known BRCA mutation. Nipple fluid was produced in 83% in at least one duct. Following NFA, 77% of ducts were cannulated for ductal lavage of which 83% produced samples with adequate cellularity. In 40% women experienced mild discomfort after ductal lavage. One women developed mild breast inflammation, resolving after antibiotics. Total drop-out rate was 21%. Withdrawal occurred in 3 women because of intolerance of the procedure and in another 3 by anxiety and pain [Other publications on obtaining breast fluid in high-risk women focused mainly on ductal lavage. In these studies NFA is solely used to identify fluid yielding ducts which can be cannulated by ductal lavage. Higgins le fluid . Mitchelyielding . Twelvesand pain . These set al., 9 healthy women were given one spray of oxytocin in both nostrils  before NFA and no adverse events were reported [An important difference between the present and earlier studies is the use of oxytocin nasal spray, which may explain higher success rates. Oxytocin is a hormone which plays a key role in the contraction of the uterus during parturition. Moreover, oxytocin is important in the ejection of milk from the mammary gland during breast feeding. In the present study the incidence of reported side effects was low. In 1.7% side effects were probably related to the aspiration procedure and in 0.5% possibly. The adverse events were mild and self-limiting in all cases. Abdominal discomfort was reported in 0.5% of aspirations after the procedure, which could be specifically related to the use of oxytocin. Long-term effects were not observed, which is in accordance with the findings of the use of long-term oxytocin intranasally in male children with autism (8–24 IU/dose) . In the reported . This maA limitation of NFA applicability in screening programs is that, at this point, fluid can be obtained in 66% of the breasts aspirated. To increase nipple fluid yielding, it is important to get more insight into the determinants affecting successful aspiration. It has been shown that the yielding of nipple fluid is associated with higher prolactin, regardless of parity and menopausal status . TogetheNipple fluid contains cells and free DNA, which makes it suitable for the detection of different biomarkers such as methylation –37, protIn conclusion, sampling of nipple fluid as breast-derived material with oxytocin-assisted aspiration is a feasible and promising approach yielding a valuable source of breast cancer specific biomarkers. Since sampling of nipple fluid is feasible in a screening population, the samples could be used for many different purposes like methylation, protein, or hormone biomarkers analysis.


P = 1.49 x 10-21). The association for rs11249433 was limited to ER-positive breast cancers (test for heterogeneity P≤8.41 x 10-5). Additional analyses by other tumor characteristics showed stronger associations with moderately/well differentiated tumors and tumors of lobular histology. Although no significant eQTL associations were observed, in silico analyses showed that rs11249433 was located in a region that is likely a weak enhancer/promoter. Fine-mapping analysis of the 1p11.2 breast cancer susceptibility locus confirms this region to be limited to risk to cancers that are ER-positive.The Cancer Genetic Markers of Susceptibility genome-wide association study (GWAS) originally identified a single nucleotide polymorphism (SNP) rs11249433 at 1p11.2 associated with breast cancer risk. To fine-map this locus, we genotyped 92 SNPs in a 900kb region  flanking rs11249433 in 45,276 breast cancer cases and 48,998 controls of European, Asian and African ancestry from 50 studies in the Breast Cancer Association Consortium. Genotyping was done using iCOGS, a custom-built array. Due to the complicated nature of the region on chr1p11.2: 120,300,000–120,505,798, that lies near the centromere and contains seven duplicated genomic segments, we restricted analyses to 429 SNPs excluding the duplicated regions (42 genotyped and 387 imputed). Per-allelic associations with breast cancer risk were estimated using logistic regression models adjusting for study and ancestry-specific principal components. The strongest association observed was with the original identified index SNP rs11249433  0.402; per-allele odds ratio (OR) = 1.10, 95% confidence interval (CI) 1.08–1.13,  NOTCH2 and FCGR1B genes [Genome-wide association studies (GWAS) have identified over 90 common genetic variants associated with breast cancer risk –19. A mu1B genes . Further1B genes –22, and 1B genes . Two ind1B genes , 25.Fine-scale mapping of the susceptibility regions identified by GWAS has the potential to further narrow down the relevant area of interest, identifying additional risk SNPs, and predicting potential functional mechanisms. Fine-mapping of the 1p11.2 locus among Chinese women (878 cases and 900 controls) identified a novel SNP rs2580520 as a variant significantly associated with breast cancer risk, which was not identified in European women . HoweverFifty breast cancer studies participating in the Breast Cancer Association Consortium (BCAC) were included in this analysis. The majority of the included studies were population-based or hospital-based case-control studies that included participants of European ancestry (41 studies), Asian ancestry (9 studies), and African ancestry (2 studies), totaling 45,276 breast cancer cases and 48,998 controls. Study participants were recruited under protocols approved by the Institutional Review Board at each institution, and all subjects provided written informed consent, as previously described [P < 1 x 10-7 and those with more than 2% of discrepant genotypes in duplicate samples across all COGS consortia. The 900 Kb genomic region for fine-mapping of the 1p11.2 locus  included all known SNPs correlated (r2> 0.1) with the index variant rs11249433. In total, 92 genotyped SNPs from the iCOGS array satisfied the initial QC metrics above.Genotyping and quality control (QC) measures used in COGs have been described elsewhere . In brieWe used imputation in order to estimate the genotypes at variants in the region not typed on the iCOGS array. Imputation was performed using IMPUTE2 , separatwww.1000genomes.org/data [After reviewing the 1p11.2 genomic region using the UCSC genome browser, we observed that there were several SNPs which mapped to multiple genomic regions due to duplication of genomic segments on both sides of the centromere on chromosome 1 , 31. Theorg/data ) and falThe LD structure based on the 1000 Genomes CEU data was visualized using the R package snp.plotter . A line P < 7x10-5, corrected for 4,371 SNPs.Standard logistic regression models were used where the common allele was the referent to assess the association of all genotyped and imputed SNPs with breast cancer risk and all analyses  used a 1-degree of freedom test (additive model) to estimate the per-allele odds ratio (OR) for the variant allele and corresponding 95% confidence interval (CI) for each SNP. Association analyses were adjusted for study and eight eigenvectors to capture population structure, obtained from principal component analyses . P-value2 statistic and p-value for heterogeneity by study.To determine if there were differences in the associated effects of the independent signals on different subtypes of breast cancer among women of European ancestry, we conducted stratified analyses according to subtypes defined by: 1) tumor histology , 2) tumor grade , and 3) ER status (ER-positive or ER-negative) subtypes. To determine if SNP associations varied significantly between defined subtypes of breast cancer, we performed polytomous logistic regression models, and P-values for heterogeneity were obtained from case-case analysis for tumor subtypes . Meta-analyses were performed using the random effects model to estimate the Iin silico functional data and expression quantitative trait loci (eQTL). Utilizing the UCSC Genome Browser and HaploReg v3 we reviewed ENCODE data to determine potentially altered regulatory motifs. RegulomeDB v1.1 was used to query publicly available eQTL data from multiple cell types associated with the identified SNPs and select SNPs significantly correlated to the tag SNP rs11249433.To evaluate any possible functional implications of our top-associated SNPs, we assessed NOTCH2 gene, and the degree of linkage disequilibrium between the SNPs, are also shown in Following quality control and genomic restrictions, a total of 429 SNPs (42 genotyped and 387 imputed) were examined for their association with breast cancer risk. -8 , and is located 57 kb telomeric from rs11249433, and closer to the NOTCH2 gene. Stepwise regression analyses conditioning on both rs146784183 and rs11249433 did not result in the identification of any additional independent signals at this locus  and rs146784183 .Of the 429 SNPs, 136 SNPs were associated with breast cancer risk overall in European women at P < 5x10P-het = 9.88 x 10-9; rs146784183: per-A-allele OR = 0.86, 95% CI 0.82–0.89, P-het = 8.41 x 10-5; P = 0.90; rs146784183: per-A-allele OR = 0.99, 95% CI 0.92–1.06, P = 0.68; 2 = 0%, P-hetstudy = 0.846) and rs146784183 .We next determined whether risk associations at the 1p11.2 locus varied by estrogen receptor (ER) status; associations observed were limited to ER-positive  and moderately-differentiated tumors , but not poorly-differentiated tumors , and less so for ductal/mixed or other tumor histology (P -heterogeneity = 0.11), though the risk reduction associated with this SNP was strongest for lobular tumors  were also associated with ductal/mixed tumors, and only 30 (19%) of those also associated with tumors of other histology . SNP rs115775083 is not correlated with the rs11249433 and rs146784183 SNPs identified to be associated with breast cancer risk in European women (r2 < 0.01). Conditioning on the top SNP identified in the women of Asian did not identify any novel signals within the 1p11.2 locus, but did reaffirm SNP rs115775083 as the most significant signal among Asian women , which showed a similar association with risk . All other SNPs were less strongly associated with risk (likelihood ratio < 1:1000 relative to rs11249433), suggesting that one or both SNPs rs11249433 and rs12134101 are likely to be causally implicated in breast cancer risk.SNP rs11249433, was strongly correlated with one other SNP, rs12134101 , we found index SNP rs11249433 to be the strongest signal within the 1p11.2 locus associated with breast cancer risk in European women. An additional association signal was identified, rs146784183, that was independent of the index SNP for overall breast cancer risk. Neither signal was found to be significantly associated with breast cancer risk among women with Asian or African ancestry, after adjusting for multiple comparisons. Notably, rs11249433 and rs146784183 displayed significant heterogeneity in risk associations by important tumor characteristics including ER status, tumor grade and histology. Our findings highlight the value of fine-mapping analyses to identify novel risk associations, and the utility of performing large-scale genotyping projects within varied ethnic populations to aid in narrowing down the genomic area relevant to future functional analyses.r2 = 0.086), and this newly identified SNP is located about 57 kb telomeric from the index SNP, and closer to the NOTCH2 gene.Fine-mapping the 1p11.2 locus was complex due to the proximity to the centromere and the presence of duplicate genomic segments. As such, we employed strict quality control measures to increase our likelihood for finding true association signals. In this study we have identified SNP rs146784183 as a novel independent signal within the 1p11.2 locus among European women. SNP rs146784183 and the index SNP were not correlated  are influencing different genes, however, the patterns of association with breast cancer sub-types suggest that they may affect similar biological and/or signaling processes.In the current study we did not perform functional analyses, however, in a study of 180 breast tumors Fu and colleagues found that carriers of the risk genotypes of rs11249433 (AG/GG) were associated with increased mRNA expression of the CH2 gene –22. Furtd tumors . If, as d tumors , rs11249Our analyses in a diverse population of women showed that the top association signals found in European women showed similar associations in women of Asian ancestry, although associations were weaker. However, no significant signals were observed among women with African ancestry. These findings support what has been previously shown in multi-ethnic studies of the 1p11.2 locus , 25, 43.The strengths of our study are in analysis of a very large data set, which includes subjects of European, Asian and African ancestry; and availability of detailed genetic and tumor pathology data, which allowed us to refine these risk associations by pathologic subtypes of breast cancer. Moreover, the findings observed in this pooled analysis did not differ significantly by study. Our study was limited by the available genomic information of the 1p11.2 region. However, the genomic map of the peri-centromeric region that harbors our region of interest was significantly improved in the latest build of the reference human genome. Due to this improvement, some genomic gaps were filled and some new pseudogene transcripts were mapped in the region; this could potentially increase SNP coverage and improve fine-mapping quality.In summary, we showed the 1p11.2 locus is specific for ER-positive breast cancers and provided data to narrow the relevant area of interests for future functional studies, which should provide further insights into the underlying causal SNPs responsible for its association with breast cancer.S1 FigPvalues  of the SNPs are shown according to their chromosomal positions (x axis). Physical locations are based on hg19.Regional plot of association results for the 1p12-11.2:120,505,799–121,481,132 breast cancer susceptibility loci from women of European (top panel), Asian (middle panel) and African (lower panel) ancestry. Association result from a trend test in—log10(DOC)Click here for additional data file.S2 FigLinkage disequilibrium (LD) plots for (A) women with ancestry from northern and western Europe (CEU), (B) Asian ancestry and (C) Yoruba (YRI) women with West African ancestry based on HapMAP version 3, chromosome 1: 120505–121481 kb. Index SNP rs11249433 is circled in red.(DOCX)Click here for additional data file.S1 FileTable A: List of studies and ethics approvals. Table B: Genotyped and imputed SNPs at the 1p11.2 locus associated with overall breast cancer risk at genome-wide significance (p < 5x10-8) among European women in BCAC. Table C: Genotyped and imputed SNPs at the 1p11.2 locus associated breast cancer risk at genome-wide significance (p < 5x10-8) after conditioning on SNP rs146784183 among European women in BCAC. Table D: Genotyped and imputed SNPs at the 1p11.2 locus associated with ER-positive breast cancer risk at Bonferroni adjusted significance (p < 7x10-5) and the corresponding association with ER-negative breast cancer risk among European women in BCAC. Table E: Genotyped and imputed SNPs at the 1p11.2 locus associated with well differentiated breast cancer risk at Bonferroni adjusted significance (p < 7x10-5) and the corresponding association with moderately differentiated and poorly differentiated breast cancer risk among European women in BCAC. Table F: Two independent association signals at the 1p11.2 locus: Association results for breast cancer risk among European women in BCAC, by tumor characteristic. Table G: Genotyped and imputed SNPs at the 1p11.2 locus associated with lobular breast cancer risk at Bonferroni adjusted significance (p < 7x10-5) and the corresponding association with ductal or mixed and other breast cancer risk among European women in BCAC. Table H: Top 5 SNPs at the 1p11.2 locus and their association with breast cancer risk among women with Asian ancestry in BCAC. Table I: Results of association analyses after conditioning on the top SNP (rs115775083) identified among women with Asian Ancestry. Table J: Two independent association signals at the 1p11.2 locus, lack of association with breast cancer risk among women with African Ancestry. Table K: Acknowledgements and funding.(XLSX)Click here for additional data file.


Background: The impact of different dietary flavonoid subclasses on risk of epithelial ovarian cancer is unclear, with limited previous studies that have focused on only a few compounds.Objective: We prospectively examined associations between habitual flavonoid subclass intake and risk of ovarian cancer.Design: We followed 171,940 Nurses’ Health Study and Nurses’ Health Study II participants to examine associations between intakes of total flavonoids and their subclasses  and risk of ovarian cancer by using Cox proportional hazards models. Intake was calculated from validated food-frequency questionnaires collected every 4 y.Results: During 16–22 y of follow-up, 723 cases of ovarian cancer were confirmed through medical records. In pooled multivariate-adjusted analyses, total flavonoids were not statistically significantly associated with ovarian cancer risk . However, participants in the highest quintiles of flavonol and flavanone intakes had modestly lower risk of ovarian cancer than did participants in the lowest quintile, although the P-trend was not significant . The association for flavanone intake was stronger for serous invasive and poorly differentiated tumors  compared with nonserous and less-aggressive tumors. Intakes of other subclasses were not significantly associated with risk. In food-based analyses used to compare subjects who consumed >1 and ≤1 cup black tea/d, the HR was 0.68 .Conclusions: Higher intakes of flavonols and flavanones as well as black tea consumption may be associated with lower risk of ovarian cancer. Additional prospective studies are required to confirm these findings. WithFlavonoids are present in many foods and beverages including fruit, vegetables, tea, and wine. Flavonoid subclasses commonly consumed in the United States include flavanones, flavonols, anthocyanins, flavan-3-ols, flavones, and polymeric flavonoids. Relatively few studies have examined the association between these compounds and ovarian cancer risk. Case-control studies generally have reported inverse associations between flavonols and flavones and risk of ovarian cancer –17. Our The NHS commenced in 1976 when 121,700 US women aged 30–55 y completed a mailed questionnaire about known and suspected risk factors for cancer and cardiovascular disease. In 1989, a younger cohort of 116,430 women aged 25–42 y were enrolled in the NHSII by using similar questionnaires. Participants have completed follow-up questionnaires biennially, providing updated information on lifestyle factors and disease diagnoses. Every 2–4 y, women receive semiquantitative food-frequency questionnaires (FFQs) , 21. To The outcome was incident epithelial ovarian cancer. We collected information about new ovarian cancer diagnoses on each biennial questionnaire. For 90% of reported cases as well as deaths that were ovarian cancer identified through family members, the US National Death Index, or the US Postal Service, we obtained medical records related to the diagnosis. When we were unable to obtain medical records, we linked to the appropriate cancer registry to confirm the diagnosis. An estimated 98% of all deaths in the NHS and NHSII are captured through the National Death Index , 23. A gDietary intake data were assessed in 1984, 1986, and every 4 y thereafter in the NHS. Similar FFQs were initially administered in 1991 and subsequently every 4 y in the NHSII. A database for the assessment of intakes of different flavonoid subclasses was constructed as previously described  by usingBriefly, intakes of individual compounds were calculated as the sum of the consumption frequency of each food multiplied by the content of the specific flavonoid for the specified portion size. We derived intakes of the subclasses commonly consumed in the US diet, specifically flavanones , anthocyanins , flavan-3-ols (catechins and epicatachin), flavonols , flavones (luteolin and apigenin), flavonoid polymers , and proanthocyanidins alone. Cumulative average intakes (energy adjusted) were calculated for a given questionnaire cycle by averaging the intake for current and preceding FFQs to assess long-term flavonoid intake and minimize the within-person variation; for questionnaire cycles without an FFQ, we used the cumulative average exposure assessed through the previous cycle. The validity and reproducibility of the FFQs have been reported previously, and correlations between major dietary sources of flavonoids, including fruit, vegetables, tea, and wine, measured by diet records and the FFQs were 0.70, 0.50, 0.77, and 0.83, respectively –29. We aParticipants contributed person-time of follow-up from the date of return of the first available FFQ to the date of any cancer diagnosis, bilateral oophorectomy, menopause as a result of pelvic irradiation, death, or end of follow-up (June 2010 for the NHS and June 2011 for the NHSII). We used Cox proportional hazard regression for time-varying covariates to estimate the HR and 95% CI for flavonoid intake in relation to risk of ovarian cancer with the lowest quintile of intake as the referent group. We used a 4-y lag between exposure assessment and disease incidence to address the issue that early symptoms of undiagnosed ovarian cancer may alter habitual food intake. For example, we used cumulative average intake from 1984 to 1990 as the exposure for the follow-up period from 1994 to 1998 and cumulative average intake from 1984 to 1994 for follow-up from 1998 to 2002. Because of relatively low within-individual correlations over time in flavonoid intake, if a woman did not answer a particular FFQ, she did not contribute person-time until she answered a subsequent FFQ.Analyses were conducted within each cohort separately and pooled (stratifying by cohort) because no heterogeneity in associations was observed by using a random-effects meta-analysis . To faciP values were 2 sided.In addition, we examined associations by histologic subtypes (serous invasive and poorly differentiated cancers compared with nonserous) and tumor aggressiveness [defined by rapidly fatal (death ≤3 y of an ovarian cancer diagnosis) compared with less aggressive ]  by usingDuring 1,139,897 person-years of follow-up in the NHS and 1,128,526 person-years of follow-up in the NHSII, 723 cases of incident ovarian cancer were confirmed. At the midpoint of follow-up, women with higher compared with lower total flavonoid intakes were slightly older and had lower total energy and lactose intakes (P-trend = 0.17) (P-trend = 0.11) and 0.79 , respectively]. Intakes of other subclasses were not significantly associated with ovarian cancer risk. We also examined the contribution of individual constituents of flavonol and flavanone subclasses , but none reached statistical significance. In stratified analysis, we did not observe an interaction with menopausal status . We generally did not observe heterogeneity by cohort (data not shown).In pooled multivariate-adjusted analyses, total flavonoids were not significantly associated with ovarian cancer risk  with nonserous tumors (n = 274). There was a suggestion that flavanones were more-strongly associated with lower risk of serous invasive and poorly differentiated tumors  than risk of nonserous tumors . In general, we did not observe different associations by tumor aggressiveness, although higher proanthocyanidin intake was associated with suggestively lower risk of rapidly fatal tumors, but not less-aggressive tumors , than risk of less-aggressive tumors .We examined associations by histologic subtype by comparing serous and poorly differentiated tumors (P-trend = 0.03) (P < 0.01). For other main foods that contributed to intake , we did not observe significant associations.We conducted food-based analyses for main dietary sources of flavonols and flavonols. Main dietary sources of flavonols were black tea (31%), onions (20%), and apples (10%), whereas citrus fruit  and juices  were the main dietary sources for flavanones ( = 0.03) . We alsoIn this large prospective cohort study we observed that participants who had the highest habitual intakes of flavonols and flavanones had significantly lower risk of epithelial ovarian cancer. Associations for flavanones were more strongly associated with lower risk of serous and poorly differentiated tumors, a more-aggressive disease phenotype. Main sources of these compounds include tea and citrus fruits or juice, foods and drinks that are readily incorporated into the habitual diet, suggesting that simple changes in food intake could have an impact on ovarian cancer risk. To our knowledge, this is the only prospective study to comprehensively examine the 6 major flavonoid subclasses present in the diet with ovarian cancer risk and the first to investigate the impact of polymers and anthocyanins.In the current study, we observed significantly lower risk of high intakes of flavonols and flavanones only for women in the top quintile (compared with the bottom quintile) of intakes. This results may have been because the absolute range in intake across the 3 middle quintiles compared with the lowest quintile was relatively small ; the measurement error in these middle quintiles may have obscured an association. Another possibility is that there was a threshold effect. Previous data suggested that there may be a threshold dose required for biological effects and low intakes are unlikely to be bioactive (n = 346 cases), we examined the only 5 flavonoid constituents available in food-composition tables at the time [3 flavonols  and the flavones ]. We observed significant inverse trends for all 3 flavonols and luteolin, although after multivariate adjustment, only kaempferol and luteolin remained associated with risk (n = 141). However, this study did observe a small increase in risk of endometrial and breast cancer when multivariable HRs between highest and lowest quintiles of intake were compared  for total flavonoid intake . To our knowledge, this is the first population-based study to suggest a chemopreventive effect for flavanones on ovarian cancer risk. Although potential anticancer effects for several flavanones, including hesperetin (and its glycoside hesperidin) and naringenin, have been described, including effects on inhibiting cell growth via cell cycle arrest and p53-dependent apoptosis , the focAvailable literature from both a molecular and epidemiologic perspective has provided some evidence for a potential role for tea in ovarian cancer prevention –42. In oSubtypes of ovarian carcinomas have different molecular, pathologic, and clinical characteristics, which support the notion that this histologic diversity may constitute several distinctive entities rather than one single disease –49, and Strengths of our study included its prospective design, large sample size with long-term follow-up, histologic subtype characterization, repeated measures of dietary intake, comprehensive assessment of the range of flavonoid subclasses present in the habitual diet, and detailed data on ovarian cancer risk factors. Our study also had some limitations. Although we adjusted for possible confounders that are strongly associated with risk of ovarian cancer, there was still the possibility of residual confounding from unmeasured factors. However, because of our detailed and updated adjustment for potential confounders, it was unlikely that these would have accounted fully for the observed results. Also, mean cumulative dietary flavonoid intakes were calculated from a database developed from recent USDA databases with additional input from other sources ; howeverIn conclusion, our data suggest that specific bioactive compounds, including flavonols and flavanones, present in plant-based foods may be associated with lower risk of ovarian cancer. In particular, black tea intake was associated with lower risk, which is already commonly consumed in many countries. Additional prospective studies and mechanistic studies are required to confirm these findings and determine whether the effect of diet varies with different subtypes of ovarian cancer because of their different molecular, pathological, and clinical characteristics.