Risk-based breast cancer screening may improve the benefit-harm ratio of screening by tailoring policy to a woman’s personal breast cancer risk. This study aims to explore Dutch women’s preferences regarding the organisation and implementation of a risk-based breast cancer screening and prevention programme, identifying potential barriers and facilitators to uptake.A total of 5110 participants in the Dutch Personalised RISk-based MAmmography screening (PRISMA) study were invited, of whom 942 completed a two-part web-based survey. The first part contained questions about personal characteristics; for the second part, women were randomly assigned to one of four hypothetical breast cancer risk scenarios  with subsequent tailored screening and prevention advice. Descriptive statistics are used to present women’s organisational preferences. Univariable and multivariable logistic regression analyses were performed using seven proxy measures for acceptability of risk-based screening  and risk-based prevention .Interest in breast cancer risk was high (80.3%). Higher assigned risk scenario was most consistently associated with acceptance of tailored screening and prevention recommendations. Increased acceptance of lifestyle changes was additionally associated with higher education. Having a first degree family history of breast cancer decreased women’s motivation to participate in preventative lifestyle measures. Acceptability of medication was associated with a woman’s general beliefs about the (over)use and benefit-harm balance of medication.Dutch women generally appear in favour of receiving their breast cancer risk estimate with subsequent tailored screening and prevention recommendations. However, women’s level of acceptance depends on their assigned risk category. Offering tailored screening and prevention recommendations to low-risk women will be most challenging. Educating women on the benefits and harms of all risk-based screening and prevention strategies is key to acceptability and informed decision-making. Risk-based breast cancer screening may improve the benefit-harm ratio of screening by tailoring policy to a woman’s personal breast cancer risk , 2. ThisAssessing the breast cancer risk of a population sample of women eligible for screening has been shown to be feasible using a modified version of the Tyrer-Cuzick breast cancer risk prediction model, including information on breast density and genetics . MoreoveA first qualitative exploration of the acceptability of risk-based breast cancer screening and prevention among a small group of women eligible for screening in the Netherlands, the United Kingdom, and Sweden confirmed the controversial nature of reduced screening intensity . Women aCross-sectional data were collected between June and October 2018 in the Netherlands using a web-based survey which was designed using qualitative focus group data [Women were selected from the participant database of the PRISMA study. PRISMA participants who met the Dutch screening eligibility criteria at the time of the survey study, i.e. aged 50–75 years and without a breast cancer diagnosis, and who consented to being approached for follow-up studies were invited to take part in this survey study. All participants were unaware of their breast cancer risk at time of participation in the survey study. We invited 5110 women in the Netherlands from three screening regions covering the North, East, and South-West of the country by sending an e-mail containing the survey’s participant information sheet. Women received a second email with a link to the online survey after they professed interest in participating via telephone or email.The survey took 20–30 min to complete and contained two parts. In the first part, all participants answered questions about different aspects of their lives, e.g., demographics, family history, and general health. For the second part, we used a computer-generated randomisation scheme to randomly assign participants to one of four hypothetical breast cancer risk scenarios, i.e., low, average, moderate, or high, with subsequent screening and prevention recommendations Fig. . Each scAcceptability of risk-based screening was assessed using three proxy measures. These measures are based on results from a previous focus group study  which in2, continuous) were acquired.Data on participants’ age (continuous), educational attainment , marital status , and body mass index , personal history of benign breast disease (yes/no), diagnosed with one of the following medical conditions: cardiovascular disease, stroke, high blood pressure, asthma, chronic bronchitis, COPD, diabetes, ulcer, kidney disease, liver disease, anaemia, thyroid disease, depression, arthritis, and backache , and current medication use for one or more of these conditions (yes/no).General health on the day of participation was measured with the general EuroQol visual analogue scale (EQ VAS) . It recoHealth anxiety was measured with the validated 14-item Short Health Anxiety Inventory , 11. EacHealth locus of control refers to a person’s beliefs or expectations about which persons or other factors determine their health . It was Life events experienced in the past year were measured using an abbreviated version of the validated Holmes-Rahe Stress Inventory . Based oGeneral beliefs about medicines were measured with the validated 8-item Beliefs About Medicines Questionnaire . It compPerceived breast cancer risk was assessed using the item ‘How do you perceive your own risk of developing breast cancer in comparison with a woman of your age?’, with five response options: low, below average, average, above average, and high. For analyses we distinguished between below average (low and below average), average, and above average (above average and high) due to limited subgroup sample sizes.Descriptive statistics were presented to establish women’s general characteristics and preferences regarding risk-based screening and prevention. Women’s characteristics and preferences were further stratified by hypothetical risk scenario . Next, we performed exploratory univariable and multivariable regression analyses to assess associations between the determinants and the seven outcome measures which function as proxies for women’s acceptability of risk-based screening and prevention. The selected associations that were explored were based on literature and our previous focus group study . In the n = 236), average risk (n = 228), moderate risk (n = 239), or high risk scenario (n = 239). Table A total of 942 women completed the survey. (response rate 18.4%). At the start of the survey women were randomly assigned to either the hypothetical low risk  would prefer the screening interval to remain at 2 years, and the start age to either remain at 50 years (27.9%) or increase to 55 years (16.2%). Both average-risk and moderate-risk women were presented with a 2-year screening interval, leading to more moderate-risk women perceiving a need for supplemental mammography screening than average-risk women, i.e. 50.0% versus 36.6%. Half of high-risk women found their presented 1-year screening interval acceptable (50.5%), with 43.0% of women opting for supplemental mammography screening. Almost one third of high-risk women would prefer a screening interval of 6 months and a start age of 40 years (59.4%). Of the women who would opt for supplemental mammography screening, 25.4% would not be willing to pay for the extra mammogram, 25.9% of women would pay regardless of the costs, and for 36.2% of women it would depend on the costs. Most women who completed the moderate (87.2%) or high (83.4%) risk scenarios would engage in more frequent breast self-examinations.Women were generally in favour of changing their lifestyle to actively attempt to reduce their risk, although changing their diet was more popular than modifying alcohol intake or exercise habits, respectively. Of the women who would be willing to change their lifestyle, 31.4% would like to receive help from a dietician, whereas 24.6% would prefer to make the changes without any external help. More than half of the women who completed the moderate or high risk scenario were willing to consider taking risk-reducing medication, expressing a preference for a pill rather than a cream. Women who were not in favour of risk-reducing medication indicated that they would only like to take medication once diagnosed with breast cancer (50.9%), they did not think the benefits outweighed the harms (25.2%), or had an aversion to medication (23.9%). Most women who were in favour indicated that they felt the benefits do outweigh the harms (75.5%), but emphasised that interactions with other medications should be avoided (24.5%).adj 1.59, 95% CI 1.09, 2.34) and increased interest in breast self-examination . Higher age was associated with decreased interest in knowing your breast cancer risk . Lower education was associated with increased perceived need for supplemental mammography outside of the national screening programme . Lower perceived breast cancer risk was associated with increased interest in breast self-examination .Table adj 1.95, 95% CI 1.28, 2.97), willingness to limit alcohol intake  and willingness to consider medication . Higher education was associated with increased willingness to increase exercise  and willingness to limit alcohol intake . Higher BMI was associated with an increased willingness to change diet  and a decreased willingness to limit alcohol intake . Having at least one first-degree relative with breast cancer was associated with a decreased willingness to change diet  and increase exercise . Additionally, women who generally believed medicines are not overused and cause more good than harm were more interested in medication than women who did not hold these beliefs .Table Our findings show that Dutch women are generally interested in their breast cancer risk and open to tailored screening and prevention strategies. However, there are some important considerations that need to be addressed to facilitate potential future implementation.We found that around 80% of women wanted to know their breast cancer risk. This is in line with previously reported estimates which vary between 74% and 95% , 16. It Dutch women saw merit in risk-based screening strategies. In line with previous research, annual screening was preferred by women completing the high risk scenario and around half of the women who completed the low risk scenario preferred to maintain the current screening interval of 2 years . ConcernWomen’s preferences regarding the organisation of a potential risk-based screening and prevention programme will likely stretch healthcare resources. With around 90% of women who completed the moderate or high risk scenario wanting a risk consultation from a GP or oncologist, the involvement of other healthcare professionals in the screening process will increase. In many European countries, GPs are an obvious choice due to their accessibility and knowledge of a woman’s personal  history. However, which professional is most suitable will depend on how healthcare is funded and arranged in a country. Previous research has shown that (primary) healthcare professionals generally lack knowledge on breast cancer risk communication and the latest screening and prevention guidelines, making additional training unavoidable , 20. In The potential of increased breast cancer awareness after risk communication was affirmed by women’s perceived need for increased breast self-examination when completing the moderate or high risk scenario. Women have previously reported that breast self-examinations are not part of their routine, yet being aware of their risk would make them feel more inclined to engage in the practice . HoweverAlthough we previously found that women’s acceptability of preventative measures for breast cancer was limited , this suhttps://karmastudy.org/ongoing-research) is investigating the minimal effective dose of tamoxifen, monitoring the accompanying side effects of each dose. Lowering the required dose of tamoxifen without compromising its risk-reducing effects may make the medication more accessible and acceptable to eligible women. Additionally, a considerable number of women professed a preference for topical tamoxifen, making further research into its effectiveness compared with oral tamoxifen worthwhile [Although low uptake of risk-reducing medication for breast cancer has been widely reported , our resrthwhile .This is the first study to comprehensively explore women’s perceptions of an integrated risk-based breast cancer screening and prevention programme. However, some caution in the interpretation of our results is warranted. Firstly, because risk-based breast cancer screening has not yet been implemented and comprehensive data on women’s breast cancer risk is not yet routinely collected, we had to rely on hypothetical risk scenarios and proxy outcome measures. Although it appears that women were able to reason within their assigned risk category, the stakes are inherently lower, and it is unclear how this may have affected the results. Although women expressed positive intent regarding participation in risk-based screening and prevention strategies, previous research has shown that the role of intent is limited in this context . TherefoMoreover, our findings are based on a selection of women who have previously participated in both breast cancer screening and the PRISMA study. This could account for the relatively large number of women with a first-degree family history of breast cancer 21.7%) in our study sample. Previous studies performed in the Dutch breast cancer screening population found that around 4.5–5.6% of women reported a first-degree family history of breast cancer , 28. Add.7% in ouDutch women generally appear in favour of receiving their breast cancer risk estimate with subsequent tailored screening and prevention recommendations. However, educating women on the benefits and harms of all risk-based screening and prevention strategies is key to acceptability and informed decision-making. Women’s preferences for preventative strategies in particular are diverse. Therefore, we need to engage women in decisions about tailored preventative strategies to optimise uptake and adherence.Additional file 1 : Supplement 1. Key characteristics of all survey participants and the proportion of invitees who previously completed the PRISMA study questionnaire


APOE (apolipoprotein E), COMT (Catechol-O-Methyltransferase), MDR1 (multi-drug resistance 1), BDNF (brain derived neurotrophic factor), and GST (glutathione-S-transferase), have been associated with vulnerability to cognitive impairment. In this study, we evaluated the relationship of these genetic variants to measures of brain health in patients with breast cancer, including neurocognitive testing and functional connectome analysis. Women with breast cancer (n = 83) and female healthy controls (n = 53) were evaluated. They underwent resting-state functional MRI scans and neurocognitive testing. Polymerase chain reaction (PCR) was performed on saliva samples to identify single nucleotide polymorphisms (SNPs) in candidate genes: APOE, COMT, MDR1, BDNF, and GST. Breast cancer patients treated with chemotherapy had slower processing speed (p = 0.04) and poorer reported executive function (p < 0.0001) than healthy controls. Those chemotherapy-treated patients that were APOE e4 carriers had significantly slower processing speed. A greater number of risk-related alleles was associated with poorer connectivity in the regions of the left cuneus and left calcarine. While breast cancer patients that are APOE e4 carriers may have a select vulnerability to processing speed impairments, other risk-related alleles were not found to influence cognitive test performance in this population. Conversely, regions of impaired functional connectivity appeared to be related to risk-related genetic polymorphisms in breast cancer patients. This suggests that a cancer patient’s SNPs in candidate genes may influence the risk of neurotoxicity. Further study evaluating the impact of genotype on biomarkers of brain health in cancer survivors is warranted.Genetic polymorphisms in select genes, including  In addition to the direct sequelae of a malignancy itself, cancer has an established impact on cognitive function3. Cancer patients have a 40% greater likelihood of reporting memory complaints than those without cancer, affecting those with active cancer and those in remission4. There are often multiple contributors to cancer related cognitive impairment (CRCI), with surgery, chemotherapy, radiation, psychosocial stress, and the cancer itself all having demonstrated contributions5. The biologic underpinnings of CRCI are likely complex, with both preclinical work and human neuroimaging studies have played central roles in elucidating some of these underlying mechanisms6. Mechanisms, including oxidative stress, mitotic inhibition of neural and glial progenitor cells, and induction of neural apoptosis have supportive contributory data7, and accelerated cortical ageing is now a central biologic construct in CRCI8.With an increasing cancer incidence and decreasing death rate12. Carriers of the epsilon 4 allele of apolipoprotein E (APOE), associated with impaired neuronal membrane repair and synaptic plasticity13, has been associated with increased incidence of cognitive impairment after chemotherapy exposure14. Similarly, polymorphisms of other genes associated with neuronal and glial health and resilience, including COMT (Catechol-O-Methyltransferase), MDR1 (multi-drug resistance 1), BDNF (brain derived neurotrophic factor), and GST (glutathione-S-transferase), also have a proposed role in augmenting neurotoxicity17.More recently, the concept of germline genetic differences imparting increased vulnerability to CRCI has been explored, with genes associated with neurodegenerative diseases and enhanced toxicity from neurologic insults being implicated6. Connectome analysis is the mathematical representation on the brain’s functional and structural networks. It provides insight into brain circuitry, including the integration and segregation of networks21. Functional connectome organization in this cohort of chemotherapy-treated breast cancer survivors compared to healthy female controls was previously evaluated, with decreased clustering as well as alterations of nodal degree in frontal, striatal and temporal areas being observed22. Here we expand upon those results by including a chemotherapy-naïve breast cancer comparison and exploring genetic contributions to chemotherapy-related connectome injury and cognitive deficit. We also focused on evaluation of regional effects using network-based analysis which represents an improvement over our previous approach.Neuroimaging studies have demonstrated changes in brain structure and function in cancer patients, lending insight into the pathophysiology of cognitive impairment in these patientsFor this retrospective study, we used all available cases from our laboratory for breast cancer survivors who had neuroimaging, neuropsychological and genetic data. This included 83 women age 41–74 years with a history of primary breast cancer (stage I-IIIA) who had completed their primary treatment  more than 6 months  prior to study entry and were currently without evidence of active cancer or gross neuropathology. They were compared with 53 healthy female controls (Table 23. Breast cancer and comparison participants were recruited via the Army of Women (http://www.armyofwomen.org/), community flyer postings and advertisements. Breast cancer participants were also recruited via local support groups and physician referrals from the Stanford Women’s Cancer Center.Breast cancer survivors were excluded for history of relapse or prior anti-cancer treatment. All participants were excluded for diagnosed psychiatric, neurologic or comorbid medical conditions that are known to affect cognitive function as well as pregnancy, MRI contraindications or major sensory deficits (e.g. blindness)24 were used to measure processing speed (Symbol Search) and working memory (Digit Span). Executive function was assessed using the Neuropsychological Assessment Battery Categories test25 and verbal fluency was measured using the Delis-Kaplan Executive Function System Letter Fluency subtest26. Verbal memory was assessed using the Hopkins Verbal Learning Test Revised Version27, which assesses both immediate and delayed recall. We also administered the Behavioral Rating Inventory of Executive Function (BRIEF), a patient reported measure of executive function28 and the Clinical Assessment of Depression (CAD) which measures depression, anxiety and fatigue29. Test scores were converted to T scores (mean = 50 + /- 10) based on the published normative data for each test. Some participants had missing data for certain assessments.Subtests from the Wechsler Adult Intelligence Scales Fourth Edition30. To coregister and normalize functional images with a standardized template, a high-resolution, 3 dimension inversion-recovery prepared fast spoiled gradient echo anatomical scan was acquired: relaxation time: minimum, echo time: minimum, flip: 11 degrees, inversion time: 300 ms, bandwidth: ± 31.25 kHz, field of view: 24 cm, phase field of view: 0.75, slice thickness: 1.5 mm, 125 slices, 256 × 256 at 1 excitation, scan time: 4:26. Two task based fMRI scans and a diffusion weighted scan were also acquired during the MRI session that are utilized in other analyses not reported here. However, the resting state scan was acquired prior to any other scans to reduce the effects of specific cognitive processes on the resting state networks.A subset of participants underwent MRI scanning which was performed on a GE Discovery MR750 3.0 T whole body scanner . Functional MRI (fMRI) data were acquired while participants rested in the scanner with their eyes closed using a T2* weighted gradient echo spiral pulse sequence: relaxation time = 2000 ms, echo time = 30 ms, flip angle = 80° and 1 interleave, field of view = 220, matrix = 64 × 64, in-plane resolution = 3.125. Number of data frames collected was 216, thus total scan time was 7:12. An automated high-order shimming method based on spiral acquisitions was employed to reduce field heterogeneity31 which were used to extract regional time series data. This was performed using the CONN Toolbox32. First, data were band pass filtered to 0.008 Hz—0.09 Hz. Then, the CompCor method33 was used to reduce physiological and other non-neuronal artifacts. This method involves extracting signal from white matter and cerebrospinal fluid regions using principal component analysis and then regressing these signals out of the total fMRI signal. Finally, temporal correlations between all possible pairs of regions were computed based on the corrected fMRI signal and normalized to z-scores, resulting in a 90 × 90 functional connectivity matrix for each participant. Negative functional edges were zeroed given evidence that properties of negative correlation networks are different than those of positive correlation networks35.Image preprocessing was performed using Statistical Parametric Mapping 8 . Briefly, images were realigned to correct for head motion, segmented and coregistered with the anatomic MRI, normalized to a standard space (MNI) template and smoothed with an 8 mm FWHM kernel. We defined 90 cortical and subcortical regions of interest (ROIs) using the Automated Anatomical Labeling atlasAPOE, COMT, MDR1, BDNF, GST.Saliva samples were obtained from all participants using the Oragene DNA OG-250 collection kit . Genotyping was accomplished by polymerase chain reaction (PCR) fragment length polymorphism analysis with restricted fragment length polymorphisms for Test scores were compared between groups using analysis of covariance controlling for age. Significant omnibus tests were supplemented with Tukey’s honestly significant difference test.39. Briefly, a network is defined as a set nodes (regions) and their connections (edges). The clustering coefficient of a node is the proportion of a nodes neighbors that are also neighbors with each other and is a measure of network segregation. Path length describes the minimum number of edges that separate pairs of nodes and is an indication of network integration. Small-worldness is an organization characteristic of brain networks and other large-scale complex biological networks that differentiates them from random networks. Small-worldness was calculated as [CC/CCR]/[PL/PLR] where CC = clustering coefficient and PL = characteristic path length and CCR and PLR are the mean clustering coefficient and characteristic path length of 100 random benchmark networks40.Global connectome properties including clustering, path length, and small-worldness were calculated using graph theoretical analysis as described previously42. Therefore, we compared connectome properties across multiple densities using the area under the curve (AUC)44. Connectomes were first corrected for age via linear regression conducted at every ROI45. We then measured connectome properties at each density from minimum connection density to the last density associated with a small-world organization47 up to a maximum density of 0.548 and calculated the AUC across this entire range. AUCs were compared pairwise between groups using nonparametric permutation testing using 5000 iterations and two-tailed p values as in our previous studies49. Specifically, AUCs from randomized groups with the same number of participants as the original groups were calculated to create a permutation distribution of between-group differences. The actual between group difference was then placed in the corresponding permutation distribution and a two-tailed p value was calculated based on its percentile position45.Thresholding connectomes is necessary for removing false positive edges and facilitating between group comparisons but can remove potentially valid information regarding differences in network topology50 with age as a covariate. Unlike our previous approach which examined connectivity for individual nodes, this method identifies connected substructures, or components, within the larger network50. Permutation testing with 5000 permutations was then used to determine group differences in components controlling for multiple comparisons using false discovery rate (FDR)50.Regional connectome differences were assessed using the Network-Based StatisticAPOE e4, COMT Val, BDNF Met, MDR1 T and GST null variants) within each participant group using t-tests or Wilcoxon rank tests, as appropriate, based on data visualization.Neurocognitive test scores and connectome properties that showed between group differences were further compared between genotypes (e.g. presence vs. absence of p = 0.04). They also rated themselves as having significantly more difficulties with executive functions than the other two groups . Although it differed between groups, CAD score was not significantly associated with any objective cognitive scores but was related to BRIEF score . However, the group comparison remained significant even after adding CAD as a covariate .As shown in Table p = 0.02). Additionally, they showed significantly lower characteristic path length (p = 0.04). The chemotherapy group showed hypo- and hyper-connectivity between several regions  and characteristic path length (p = 0.02). Regionally, there were multiple areas of hypo-connectivity in the chemotherapy group .There were no differences in global connectome metrics between healthy controls and chemotherapy naïve survivors. However, there were some regions of hyper- and hypo-connectivity in the chemotherapy naïve group Fig. . CAD scoAPOE e4 allele  performance was significantly lower in chemotherapy-treated survivors who had the ele Fig. . No otheBDNF Met allele  as well as those who possessed the MDR1 T allele . Additionally, the connectivity between right and left paracentral lobule, which was lower in the chemotherapy group compared to chemotherapy naïve group, was significantly lower in chemotherapy-treated survivors who had the MDR1 T allele . Further, as shown in Fig. p = 0.02) and in the chemotherapy naïve group , but not in healthy controls (p = 0.20). In healthy controls who carried the APOE e4 allele, connectivity between left olfactory and left superior parietal lobe was higher  while connectivity between left hippocampus and left rolandic operculum was lower . No other genotype comparisons or correlations were significant. There was not enough variance to conduct comparisons for the COMT genotype.There were no differences between genotype groups and global connectome properties. Regionally, connectivity between the left calcarine and left cuneus, which was lower in the chemotherapy group compared to both control groups, was significantly lower in chemotherapy-treated survivors who had the 52, have been evaluated in relation to these genetic signatures. With roles identified in neuronal repair and survival, dendritic and axonal growth, long term potentiation, CNS concentrations of cytotoxic therapies57, the genes evaluated in this study have an association with brain ageing, a proposed concept behind CRCI58. The functional connectome, representing key contributions of genetic, epigenetic, and environmental variables61, has been shown to be a viable biomarker of neurologic impact of cancer62. In this study, we identified that patients harboring those genetic signatures associated with accelerate cognitive ageing have associated alterations in brain functional connectivity, supporting genetic relationship to cancer-related neurotoxicity.This innovative study evaluates the relationship of genetic polymorphisms associated with neural ageing and vulnerability to neurologic insults on cerebral connectivity. This is the first study to our knowledge where both cognitive function and the functional connectome, a demonstrated biomarker of CRCIAPOE4 carriers showing impairment. The lack of association with other cognitive functions may be secondary to the low rates of cognitive impairment identified in this population as a whole. A lack of sensitivity of formal neuropsychological evaluation in corroborating subjective cognitive complaints has previously been identified63. The disparity between self-reported and objectively defined cognitive dysfunction is further supported in this study, with patients rating themselves as having significant cognitive issues illustrated by different BRIEF scores between the groups, despite the lack of executive dysfunction by clinical testing. The adult children of Alzheimer’s patients carrying the APOE4 allele Alzheimer’s have been found to have slower processing speed than those that do not harbor this allele, and have lower white matter volumes64. Processing speed is a commonly affected domain in CRCI, and the APOE4 allele could render this domain particularly vulnerable to dysfunction58.Of all cognitive domains evaluated on clinical testing, only processing speed had a significant interrelationship with genotype, with 22, highlighting the general impairments in information processing and network integration that can occur in cancer survivors. Conversely, no global differences were observed in connectome properties between genotype groups. There are several potential explanations for this finding. The selected at-risk alleles may not impact on global connectivity, instead leading to more focused regions of aberrant cerebral connectivity with relative preservation of global brain function. Furthermore, differences in the allele frequency in our test group may have led to these distinct results. While variable amongst ethnicities65, the prevalence of APOE4 carriers in the general population is estimated at 14% globally66, a smaller proportion than in our study population. Similarly, the MDR1 T allele has been identified in 53%67, but was higher in our evaluated population. As such, these could alter our identification of abnormalities.A distinct global connectome organization was demonstrated in the chemotherapy group, including lower clustering, shorter characteristic path length, and hypo- and hyper-connectivity between multiple regions. These distinct alterations in large-scale brain networks have been highlighted previously in chemotherapy-treated patientsAPOE has been found to mediate tissue repair, immune response and regulation, and cell growth and differentiation68, biologic processes that influence individual susceptibility to malignancy. A meta-analysis evaluating the APOE gene in breast cancer patients identified one and a half times increased risk of breast cancer among Asians carrying the e4 protein isoform of APOE, supporting this allele as a low-penetrant risk factor for development of breast cancer69. As the COMT gene is an estrogen-metabolizing enzyme, polymorphisms have been postulated to impact breast cancer risk, with catechol estrogens having proposed oncogenic influence. Despite this theoretical relationship, population-based studies have failed to demonstrate a compelling relationship between COMT polymorphisms and breast cancer carcinogenesis71. As the MDR1 gene encodes the ATP-dependent cellular efflux pump p-glycoprotein, protecting cells from xenobiotics and metabolites, it has also had a proposed role in carcinogenesis. Multiple cancer types have been associated with the MDR1 C3435T polymorphism in a meta-analysis of case–control studies72. These findings suggest complex interrelationships between these genetic polymorphisms and carcinogenesis and may render individuals vulnerable to both cancer the neurotoxicity of its therapies. These relationships warrant further exploration.The genetic polymorphisms evaluated in this study have may also predispose to cancer itself, adding further complexity to the interrelationship with brain health. BDNF Met allele or MDR1 T allele. Further, lower left calcarine and left cuneus connectivity was also associated with a higher number of at-risk alleles in the breast cancer population, independent of chemotherapy exposure, but not in healthy controls. These regions have had implicated vulnerability in other imaging studies of CRCI. In a study of hippocampal-cortical functional connectivity, increased functional connectivity was demonstrated between the hippocampus and left cuneus in breast cancer survivors, as opposed to healthy controls73. We previously observed significant hyper-activation in left (and right) cuneus among chemotherapy-treated breast cancer survivors compared to healthy controls during a verbal memory task74. Increased perfusion in the left cuneus and left calcarine has been demonstrated in patients with breast cancer exposed to chemotherapy compared to both healthy and chemotherapy-naïve controls, a finding also correlated with dysfunction changes in executive control network scores75. Our analysis suggests dysfunction in these regions may be influenced by the presence of at-risk alleles, a finding that warrants further exploration.In contrast to global connectivity, regional changes were found to be related to at-risk allele status. Regional connectivity involving left calcarine and left cuneus as well as right and left paracentral lobule was significantly lower in chemotherapy-treated patients treated with APOE e4 allele. The olfactory system and hippocampi are of central interest in neurobiology given their roles in neurogenesis. Neurogenesis extends well-beyond the embryonic and post-natal system in select regions of the brain. The subventricular zone generates interneurons for the olfactory bulb, and the olfactory neurons generate new excitatory sensory neurons that send their axons to the olfactory bulb76. Similarly, adult neurogenesis also occurs in the subgranular zone of the hippocampal dentate gyrus77. Alterations of connectivity in these regions among individuals harboring the APOE e4 allele may be associated with increased vulnerability to age-related cognitive decline. For example, APOE e4 is a risk factor for Alzheimer’s disease, where neurogenesis is known to be dysfunctional78. However, post hoc analysis did not indicate significant differences in cognitive performance between healthy controls based on APOE e4 genotype in this sample. It is also unclear why this finding was specific to healthy controls but may suggest an altered relationship between genotype and neurogenesis in patients with breast cancer that requires further study.Another unique finding in our analysis were the differences in connectivity between left olfactory and left superior parietal lobe as well as left hippocampus and left rolandic operculum for healthy controls with the 80, but is not defined in cancer. Recent systematic review has found variable correlations between genetic polymorphisms and CRCI81, precluding the conclusive definition of a relationship. Connectomics provides a novel contribution to elucidating this relationship. The data presented in this study supports at-risk alleles impacting vulnerability to CRCI, with the numbers of alleles expressed in an individual predicting poorer cerebral connectivity. Limitations of this study include the small number of patients studied and the differential frequencies of at-risk alleles compared to the normal population. The latency between initial breast cancer therapy and neurocognitive testing and imaging was greater than 5 years, significantly longer than most studies of CRCI, which generally evaluate patients within a year of treatment completion84. This prolonged interval may have influenced our identification of cognitive impairment on formal testing and our findings on connectome analysis. However, many studies have identified cognitive deficit in cross-sectional cohorts of breast cancer survivors who are decades off-therapy86. Therefore, many factors may contribute such as choice of neuropsychological tests and/or small sample. The focused study of breast cancer patients also suggests these findings should be validated in different oncology patient groups to determine their generalizability. Furthermore, a predictive relationship between these genetic polymorphisms and CRCI or connectome alterations cannot be definitively drawn given the lack of baseline neurocognitive and neuroimaging data. Larger prospective study in this area will allow for more definitive analysis of the influence of genetic polymorphisms on brain network topology and the development of CRCI. Ultimately, a comprehensive understanding of at-risk alleles to the development of cancer-related neurotoxicity may assist in identifying high-risk patients, allowing for appropriate counseling and risk-reduction strategies to be enacted where feasible.The potential for genetic polymorphisms to modulate the impact of neurologic insults has been identified in several diseasesAll procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee  and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.Informed consent was obtained from all individual participants included in the study.


New evidence has found breast and cervical cancer risk factors unique to African American women. Thus, there is a significant need to increase their knowledge and understanding of relevant risk factors and the potential protective benefits associated with breast‐feeding and HPV vaccination. The National Witness Project is a robust, evidence‐ and community‐based lay health advisor programme that uses group education, navigation and survivor narratives to increase cancer screening among diverse underserved women.t tests were used to compare pre‐/post‐test survey scores.A multi‐phase, community‐based participatory research study was conducted across three sites in Buffalo, NY, New York City and Arkansas between October 2016 and January 2017. Pre‐/post‐test surveys were administered during volunteer trainings and community programmes. An evaluation survey was also administered at the Annual Meeting for Education and Networking. Paired sample P ≤ .001).Trainee survey results showed the overall mean per cent correct pre‐/post‐test scores were 47.7% (SD: 21.87) and 79.2% (SD: 16.14). Altogether, 31 educational programmes reached 332 community participants. Participants’ breast and cervical cancer knowledge scores were significantly higher after the education programme (84.4%) than before (55.3%) with a mean change score of 29% (This paper reveals the underlying complexities to update the educational curriculum content of a multi‐site, community‐based outreach organization. The new curriculum significantly improved African American women's knowledge about breast and cervical cancer by 10%‐36%, clearly demonstrating that this information was new to them. The need for education programming in African American communities to disseminate cancer prevention and risk information remains high. The National Witness Project (NWP) model was developed by Erwin in 1991 in collaboration with African American cancer survivors to reduce breast and cervical cancer disparities among African American women through faith‐ and community‐based educational programmes.This paper presents the systematic approach used to update the NWP educational programme curriculum and evaluate its effectiveness among participants who attended educational programmes as part of a pilot study. The first objective in this effort was to update and strengthen the NWP educational curriculum to include new scientific content relevant to breast and cervical cancer prevention in African American women. The second objective was to determine the suitability and effectiveness of the new curriculum among participants attending a NWP educational programme at three pilot sites prior to its dissemination and implementation throughout the NWP network. A unique aspect of this study is the capacity building that occurs at both the community partner level (ie NWP sites) and at the community participant level (ie African American women). Both levels of engagement are central to a community‐based participatory research (CBPR) approach and in line with the mission of the original NWP model. These objectives were accomplished using a progressive, four‐phased approach. What follows is a report of the results and findings from each of the four phases.2Phase I focused on revising the curriculum using a CBPR approach that involved multiple and iterative processes in which the community partners worked alongside the scientists to review the curriculum. The next phases related to a pilot implementation study of the new curriculum at three NWP sites and dissemination of the pilot study results to the national members. More specifically, phase II involved training NWP staff at three sites to deliver the new curriculum as part of the CBPR education programme, and phase III examined its acceptability, relatability and ability to educate participants attending a NWP event. Phase IV involved dissemination of the new education curriculum to NWP members attending the Annual Meeting for Education and Networking (AMEN) and subsequently revising the curriculum in light of their evaluation and feedback.2.1The pilot study of the feasibility and educational effectiveness of the new curriculum was conducted across three established NWP sites located in Buffalo, New York (WNY), New York City (NYC) and Little Rock, Arkansas (AR). These sites were chosen because of established research collaborations with investigators and their ability to conduct the pilot implementations of the new curriculum in less than three months. Also, training requirements were minimal in WNY and NYC as both were versed in the technology used during data collection.The population of African American individuals in each of the study locales is comparatively higher than the national average of 12.2%.2.2Figure The research was conducted by local staff and volunteers at the three site locations with support from the cancer centre. Study procedures were approved by the research ethics board (WNY) and deemed as exempt by IRB (NYC). The AR site is a community‐based organization without affiliations to an academic or cancer centre and was included in the Institutional Review Board approval.2.3In this phase a one‐day collaborative forum was held in May 2016. Investigators, together with four scientific consultants and NWP site staff, met to update the original NWP education curriculum, its presentation to the community, and develop training material for the LHAs. The consultants were selected for their expertise in breast cancer epidemiology, African American cancer disparities, social and behavioural science, vaccine‐related cancer prevention (eg HPV) and cancer genetics. Prior to this undertaking, all had previously contributed to the NWP evolution, dissemination and implementation efforts.Based on contributions from the NWP Board of Directors and staff members, the original narrative communication2.4Training for volunteers to deliver the new education programme was organized and conducted locally at each site by NWP leadership (Erwin and Johnson) over a 2‐month period. A minimum of five LHA volunteers at each site  attended training. This study used the Turning Point Audience Response System (ARS)  polling software which allowed the LHA to ask interactive questions, track participant progress and receive instant feedback. Participants used a clicker to answer questions that were read aloud by the LHA during the education programme. The AR site had not previously used laptops, PowerPoints or embedded videos to deliver their community‐based education programme, nor were they familiar with data collection using the polling software. Thus, AR received training in the use of these technologies to increase their capacity. Additional training time and booster sessions were required to train LHAs at the AR site on using the ARS format to deliver the education programme presentation; however, the uptake of the ARS technology expedited data collection during programme delivery and data transfer for analysis following the presentations. Specific to the delivery of the new curriculum, this training involved didactic presentations, open discussions and role play/programme practices. In order to evaluate trainees’ understanding of the new content and their ability to answer any questions that may arise as they deliver the community education programme, trainees were given a knowledge survey (n = 9 items) about the curriculum before and after training. The polling software shows response data for each item, thereby allowing trainers to spend more time on content that requires further discussion to clarify and ensure understanding of the material to determine readiness of the LHA to deliver the education programme. The survey included items on screening guidelines and modifiable  and non‐modifiable (ie genetic) breast and cervical cancer risk factors. For example, true/false questions such as ‘African American and white women get breast cancer at about the same rate’ and ‘HPV is transmitted only through sexual intercourse’ were used. Information about the trainees’ role was also collected and included items to determine how long they have been with working with the NWP team, as well as their familiarity with and capacity to use technology. Lastly, trainees were also certified in research ethics through the Collaborative Institutional Training Initiative (CITI) programme 20162.5For the CBPR pilot implementation, each site delivered a minimum of nine community educational programmes between October 2016 and January 2017. Programmes were held in churches, community centres or other accommodating facilities and were open to all adults wanting to attend. Inclusion criteria for the pilot study (ie completion of the pre/post programme assessments) limited participants to African American women ages 18 and above. Staff read a study information sheet aloud to eligible participants prior to initiating the education programme and allowed time for any questions about the study. Those who agreed to participate provided verbal consent and were given a copy of the study information sheet.A contact card was distributed to participants to collect their demographic information, recent breast and cervical cancer screening history (ie ≤12 months and ≤3 years) and willingness to be contacted in the future for other programmes and/or research. Subsequently, pre‐ and post‐programme questionnaires were used to assess their breast and cervical cancer‐related knowledge, perceived risk and screening‐related behavioural intentions . Item measures were presented as questions or statements on PowerPoint slides and were also read aloud to address any literacy issues. Sample items included ‘The risks and causes of breast cancer are the same for African American and white women’ and ‘HPV causes cervical cancer’ with ‘True/False/Don't Know’ as response options to choose from. Self‐efficacy items were ‘I can make an appointment for a …mammogram …Pap test’ and ‘I know how to go about getting a …mammogram …Pap test’ with response options on a five‐point Likert scale ‘1—Strongly disagree’ to ‘5—Strongly agree’. Screening behaviour‐related intent items included ‘Do you plan to have a …mammogram …Pap test in the next 12 months?’ (Yes/No/Not sure/Not applicable); and ‘How likely are you to get a …mammogram …Pap test in the next 12 months?’ with response options on a scale ‘1—Not at all likely’ to ‘6—Not applicable’. The post‐programme survey included questions from the pre‐survey in addition to items about the participant's satisfaction and impact of the educational content, for example: ‘Much of the information in this programme was new to me’ with response options on a scale ‘1—Strongly disagree’ to ‘5—Strongly agree’. An item to assess intent to screen as a result of programme attendance was also included: ‘Attending this programme has influenced my thoughts on getting screened’ with response options on a scale ‘1—Very much’ to ‘5—Not at all’. Responses were collected electronically using ARS keypads and then linked to their corresponding contact card. Summary scores were calculated from participant responses to 11 multiple choice knowledge items including six breast cancer and five cervical cancer item. Paper versions of the questionnaire were available if needed.Participants who were eligible for breast and cervical cancer screening were contacted at 2 weeks following the educational programme to assess knowledge, retention, dissemination, risk perception, self‐efficacy and intent for breast and cervical screening. Participants who were non‐adherent to screening guidelines were contacted again at 2 months to assess dissemination of programme content and screening intent/completion.2.6All existing NWP sites (n = 18) were invited to send their Project Directors and up to five LHAs and/or Witness Role Models to the NWP Annual Meeting for Education and Networking (AMEN) to be introduced to the new curriculum, shown the results of the pilot implementation CBPR, and contribute to dissemination plans. AMEN was a three‐day conference hosted by the WNY site. The first day included a two‐hour plenary session where the science and research behind the changes to the education curriculum changes were presented by the investigators and consultants. All presentations were tailored to the lay audience. Day two began with presentations of the new curriculum followed by results from the pilot implementation study. At the end of this session, an evaluation survey was distributed and conference attendees were asked to rate the new curriculum, content and format on various aspects such as presentation configuration, use of technology and length. Ratings were collected on a five‐point Likert scale ranging from ‘1—Not at all acceptable’ to ‘5—Completely acceptable’. Attendees were also asked about their beliefs about the importance of the new curriculum's content. After posing the general question ‘How important are the new messages on breast and cervical health that are included in the program to your health and/or the future health of your family?’, a total of 13 messages that collectively summarized the revised curriculum content were shown separately. Attendees were asked to rate each message on a four‐point Likert scale ranging from ‘1—Not important’ to ‘4—Essential’. Basic demographic information, age  and educational attainment  were collected.The remaining AMEN sessions included various organizational workshops to support the NWP sites on strategies to improve communication and collaboration, grant writing tips, board development activities and volunteer support. A breakout session exercise was also conducted using a small group discussion format led by a facilitator to generate site‐specific strategic implementation plans.2.7The updated curriculum includes a set of PowerPoint presentation slides with embedded videos for LHA‐led educational instruction with community participants. The final education programme was shown from laptops by the trained LHAs. The programme included ‘talking points’ for each slide to support the LHA in their presentation. Pre/post knowledge and screening assessments were included to reflect the changes made to the educational programme. New content on risk and opportunities for prevention regarding breast and cervical cancer relevant to African American women were presented in lay terms. A graphic representation of an African American medical doctor was also inserted throughout the presentation to emphasize and reinforce the importance of consulting one's health‐care provider to further discuss and understand their own personal risk. The two short (7.5 minutes each) educational videos were embedded into the programme presentation as part of the newly revised electronic media format. The first video covered the history and background of the NWP education programme and overview of the African American woman's experience with breast and cervical cancer, and the second focused on research findings from the AMBER study specific to the role of breast‐feeding in reducing breast cancer risk among African American women, including scientists  speaking about the results of their research.2.8t tests for both the training and education programme surveys. Responses from 29 participants from phase 3  were excluded from the final analyses as they did not meet eligibility criteria for race/ethnicity and gender. The final analysis sample had a total of 303 participants including 139 from WNY, 104 from AR and 60 from NYC. Responses from the AMEN survey were summarized using descriptive statistics. All analyses were conducted using SPSS 21.0 software .Study site baseline characteristics were compared across all three study sites . Chi‐square was used for categorical values, and one‐way ANOVA was used for all continuous values. Scores from pre/post knowledge tests were computed as the average per cent correct and the mean per cent change scores of responses to each type of question  and across existing vs new educational content. Comparisons of pre‐/post‐test scores were made using paired sample 33.1P < .000). Scores by site are presented in Table Training across the three pilot sites resulted in a total of 24 trained individuals. There were seven LHAs, four role models, seven staff and six additional volunteers. The overall mean for per cent correct pre‐ and post‐training scores were 47.7% SD: 21.87) and 79.2% (SD: 16.14). Pre‐/Post‐test scores indicate that participants scored significantly higher on the post‐test with a mean per cent change score of 31.5% (.87 and 73.2P ≤ .001). The majority of participants were over the age of 50 (56%) across all three sites. Almost 60% of programmes were conducted during the week, and almost half of the programmes were in the afternoon and lasted on average 80 minutes.A total of 31 educational programmes were piloted  and reached a total of 332 African American female participants. Comparisons of baseline characteristics across the three study sites and overall are presented in Table P ≤ .001). For breast cancer items, participants also scored significantly higher on the post‐test (88.7 Mean % vs 57.7 Mean %) with a mean per cent change score of 35.9% (P ≤ .001). Similarly, for cervical cancer items, participants knowledge increased on the post‐test, with an average mean per cent correct score of 94.4% with a mean per cent change score of 36.8% (P ≤ .001). For new items covering content specific to breast‐feeding, parity and HPV virus, participants displayed an increased score in the post‐test (92.4 Mean % vs 43.7 Mean %), with a mean per cent change score of 48.7% (P ≤ .001).The overall mean and mean per cent change scores for pre/post knowledge breast and cervical cancer items including original and new educational content are presented in Table Although the primary aim for the pilot was feasibility and increase of knowledge by participants, telephone follow‐up surveys with a limited sample (20%) of previously non‐adherent women attending the new education programmes demonstrated a 33% screening rate two months later.3.3Evaluation survey results regarding the new curriculum content were collected from participants in attendance during the AMEN and are summarized in Table 4P < .000).We conducted a CBPR approach including four phases to update the NWP educational curriculum and evaluate its effectiveness among participants who attended education programmes as part of a pilot study. The first phase resulted in the production of the updated NWP curriculum. The curriculum includes a set of PowerPoint presentation slides with embedded videos for LHA‐led educational instruction with community participants as well as updated training materials for train the trainer instruction with LHA and WRM volunteers. The second phase involved training the trainers on the new curriculum at each of the three participating pilot sites. A total of 24 individuals were trained across the three pilot sites. Trainees scored significantly higher on the post‐test with a mean per cent change score of 31.5% , it is essential that this critical information be incorporated into this multi‐site programme in order to reach individuals in various communities at a grass roots level. This article reveals the complexity of the process necessary to update educational curriculum content for a multi‐site, community‐based outreach organization such as the NWP, as well as evidence demonstrating the need for this information in these communities. It was essential to receive perspectives, relevant content and evaluation from representatives described in each of the four phases of the research for the final product to be accurate, culturally appropriate and acceptable for dissemination.P‐values all ≤0.001, mean change 10%‐36%) clearly demonstrated that this was new information to participants at these sites—both community members and NWP volunteers/staff. This scientific content provides new opportunities to increase awareness and understanding regarding breast and cervical cancer risk relevant to African American women. The information on the protective benefits of both breast‐feeding and the HPV vaccination relates to behaviours that either occur or are recommended at younger ages in comparison with relevant cancer screening ages for both breast and cervical cancer and therefore apply to a broader age group of women. The new educational content on breast‐feeding and HPV vaccination serves as an opportunity to expand the focus for conducting the NWP programme and outreach with multi‐generational African American audiences.The new NWP educational curriculum was shown to be effective among community participants as the overall change in knowledge scores increased significantly at post‐test. Scores increased consistently across both the breast and cervical cancer items that were assessed. The greatest mean per cent change scores were observed for the new items that covered breast‐feeding, parity and HPV virus content. The significant pre‐ to post‐education knowledge scores  regarding breast and cervical cancer risk and opportunities for risk reduction targeting underserved African American women. Many of the women in these communities were unaware of or very few had been exposed to information on the cancer prevention benefits of either of these primary prevention strategies prior to our pilot study. Despite the important strides that continue to be made in the science of cancer, there are challenges in how to disseminate new information on important advances to achieve health equity at the community level. Part of the challenge in disseminating the new NWP content relates to its acceptability among women in the African American community as was demonstrated in the AMEN evaluation survey results. The information on parity (ie higher parity without breast‐feeding associated with higher risk of TNBC) showed the most variability with respect to importance. This highlights the importance of credible messengers and decades of experience the NWP has amassed through their community‐based LHA‐led cancer prevention efforts targeting underserved African American women.5There are limitations to our study as this was not a randomized control trial at the site or programme level. In phase 2, differences in the trainees’ pre/post knowledge assessment scores across the sites may have been due to the different sizes of training classes at each site. During phase 3, the pilot implementation of the new curriculum, pre/post programme assessments helped to measure changes in knowledge but are not an indicator of participants’ ability to retain the information or their actual behaviours with regard to breast and cervical cancer risk reduction (eg breast‐feeding or HPV vaccination). We encountered missing data for either pre‐ or post‐test or for both the pre‐/post‐test for phase 3 (pilot study) as not all participants answered all questions. Therefore, we have different sample sizes for the comparisons across breast and cervical cancer as well as existing versus new content. We were limited in our collection of breast and cervical cancer screening outcomes as this was not a primary endpoint for this study. Our primary objective was to assess feasibility and test effectiveness of the new curriculum. Cancer screening outcomes were collected for a subsample of participants reached as a component of follow‐up; however, these results were not significant. The collection of participant demographic information was limited due to programme delivery‐related time constraints. Thus, we were unable to look at potential differences by level of education among participants. The addition of multiple novel topics (ie parity and breast‐feeding and HPV vaccination) required additional time to explain and allow for participant questions.6This study demonstrated that updates in scientific content and format changes can be successfully adapted into the community‐based NWP model. Our results also contribute to limited research on the critical importance of community‐based research approaches to both translate and disseminate current research findings out into the community. The NWP model remains a culturally relevant and effective way to reach African American women with breast and cervical cancer information in efforts to address existing cancer health disparities. The NWP model also demonstrates sustainability through its extensive network of sites across the United States which may serve to facilitate advocacy efforts to focus on specific cancer prevention initiatives (eg breast‐feeding) to help reduce barriers and promote the new curriculum on a national level. This network has the potential to disseminate new breast and cervical cancer prevention messages on a much broader scale and provides an opportunity to further study relevant implementation and dissemination factors in adopting the new curriculum on a larger scale.All authors have no financial interest in this study and no conflicts of interests to disclose.Elisa M. Rodriguez; Lina Jandorf; and Deborah O. Erwin conceptualized the data. Frances G. Saad‐Harfouche and Lina Jandorf involved in data curation. Frances G. Saad‐Harfouche and Lina Jandorf formally analysed the data Deborah O. Erwin; Elisa M. Rodriguez; Detric Johnson; and Frances G. Saad‐Harfouche acquired funding. Deborah O. Erwin; Elisa M. Rodriguez; Lina Jandorf; and Detric Johnson investigated the study. Deborah O. Erwin; Elisa M. Rodriguez; and Lina Jandorf involved in methodology. Deborah O. Erwin; Elisa M. Rodriguez; Lina Jandorf; Detric Johnson; Frances G. Saad‐Harfouche; Nikia Clark; and Christy Widman involved in project administration. Deborah O. Erwin; Elisa M. Rodriguez; Lina Jandorf; Detric Johnson; Frances G. Saad‐Harfouche; Nikia Clark; and Christy Widman provided resources. Frances G. Saad‐Harfouche and Lina Jandorf assisted with software. Deborah O. Erwin; Elisa M. Rodriguez; Lina Jandorf; and Detric Johnson supervised the study. Deborah O. Erwin; Elisa M. Rodriguez; and Lina Jandorf validated the data. Deborah O. Erwin; Elisa M. Rodriguez; Lina Jandorf; Detric Johnson; Frances G. Saad‐Harfouche; Nikia Clark; and Christy Widman visualized the data. Elisa M. Rodriguez; Lina Jandorf; Frances G. Saad‐Harfouche; and Deborah O. Erwin involved in writing—original draft. Elisa M. Rodriguez; Lina Jandorf; Frances G. Saad‐Harfouche; Deborah O. Erwin; Julia A. Devonish; Detric Johnson; Anika Stewart; Nikia Clark; and Christy Widman involved in writing—review and editing.


The relationship between long-chain omega-3 (LCn3), alpha-linolenic acid (ALA), omega-6 and total polyunsaturated fatty acid (PUFA) intakes and cancer risk is unclear.We searched Medline, Embase, CENTRAL and trials registries for RCTs comparing higher with lower LCn3, ALA, omega-6 and/or total PUFA, that assessed cancers over ≥12 months. Random-effects meta-analyses, sensitivity analyses, subgrouping, risk of bias and GRADE were used.We included 47 RCTs . Increasing LCn3 has little or no effect on cancer diagnosis , cancer death  or breast cancer diagnosis ; increasing ALA has little or no effect on cancer death . Increasing LCn3  and ALA  may slightly increase prostate cancer risk; increasing total PUFA may slightly increase risk of cancer diagnosis  and cancer death  but total PUFA doses were very high in some trials.The most extensive systematic review to assess the effects of increasing PUFAs on cancer risk found increasing total PUFA may very slightly increase cancer risk, offset by small protective effects on cardiovascular diseases. The other major health risk worldwide is cardiovascular disease, responsible for 37% of premature deaths due to non-communicable disease in 2012, where cancers were responsible for 27%.4Cancer is a leading cause of morbidity and mortality worldwide with ~17 million new cases and 9.6 million cancer-related deaths in 2018.5 Potential mechanisms for PUFAs in cancer aetiology include their being precursors to lipid mediators regulating metabolic pathways and inflammatory responses,6 oxidative stress, and changes in membrane composition that could affect cell signalling pathways.7 Reducing dietary fat (including PUFAs) appears to result in lower weight in adults,8 so lower PUFA intake  could offer protective effects against those cancers that are associated with overweight. These mechanisms suggest that omega-3 may be protective, and omega-6 and total PUFA may exacerbate cancer risk. However, oily fish and fish oil capsules may contain contaminants such as mercury and dioxins, potential carcinogens.12Dietary polyunsaturated fatty acids (PUFA) have been postulated as a modifiable component of lifestyle that could influence cancer risk. PUFA includes long-chain omega-3 (LCn3 including eicosapentaenoic acid and docosapentaenoic acid), alpha-linolenic acid  and omega-6 fats . Polyunsaturated fats are common healthy eating choices, and fish oil (LCn3) and flaxseed (ALA) supplements commonly consumed.13 Within the Japanese population, whose traditional diet is rich in oily fish, incidence of some cancers has increased with more westernised food consumption and lifestyles.6 One systematic review of cohort studies did not pool data but found some cohorts with positive associations, some with negative associations and more with null associations for omega-3 and a variety of cancers, including breast and prostate cancer – overall there was no trend to suggest that omega-3 fatty acids are associated with total cancer risk.14 A systematic review of 10 RCTs comparing high to low omega-3 intake for at least 6 months found no evidence that increasing omega-3 fats altered cancer incidence.15 Later meta-analysis of RCTs increasing omega 3 intake over at least 6 months found omega-3 supplementation increased the risk of cancer by 10% but this was not statistically significant,16 and this review did not analyse for specific cancer types, provided limited information on dosage and did not stratify by supplementation level.Evidence for effects of polyunsaturated fats on risk of cancer is conflicting. An early RCT, the Lyon Diet Heart Study, suggested that a Mediterranean type diet, supplemented with an experimental canola (rapeseed) oil-based margarine rich in oleic and ALA, reduced cancer diagnoses by 61% compared to those on the American Heart Association diet.18 A cohort study of over 35,000 post-menopausal US women suggested that taking omega-3 supplements was associated with a 32% reduction in breast cancer risk,19 although other cohort studies are not consistent in this relationship.20 A large European cohort study (EPIC) found no association between fatty fish consumption and breast cancer risk.21 Comprehensive systematic reviews of observational studies suggested no relationship between total polyunsaturated fat intake and risk of breast cancer22 or omega-3 intake and breast cancer risk.23The Mediterranean diet, which is high in polyunsaturated fats, has attracted attention because of the historically lower breast cancer rates in Mediterranean countries than in other parts of Europe and the United States.25 but a systematic review found inadequate data to determine whether fish-derived omega-3 fatty acids were associated with prostate cancer incidence and progression.26Two nested case-control studies of men suggested that high serum long-chain omega-3 fatty acids were associated with increased risk of prostate cancer and high-grade prostate cancer,27 Further increasing polyunsaturated fat intake is associated with healthy eating and prevention of cardiovascular disease in the general public, but is still scientifically controversial.28 The use of supplements as additions or replacements to food stuff has gained traction with the general public. It has been estimated that approximately 38% of American adults use complementary medicines and fish oil, omega 3 or DHA supplements are the most commonly used non-vitamin, non-mineral natural product (37.4%) and flaxseed the 4th (15.9%).5 LCn3 is ingested in the form of oily fish or fish oil (often fish liver oil) capsules, however, these may contain contaminants. Heavy metals such as mercury, cadmium, chromium, nickel, lead and cobalt and toxic compounds such as dioxins have been found in fish and fish oils representing a potential risk to health.12 It is therefore important to assess both potential benefits and harms of increasing omega-3, omega-6 and total polyunsaturated fats on cancer risk to better inform members of the public considering dietary change or supplementation.Some polyunsaturated fats are essential in the human diet, and UK dietary reference values suggest we need to eat at least 6.5% of our energy intake in the form of cis-polyunsaturated fats.32 this review assessed the risks and protective effects of increasing omega 3, omega 6 and total polyunsaturated fat (PUFA) intake on total, breast and prostate cancer incidence in adults, gathering a much larger set of randomised trials than has previously been assessed as it included trials where cancer diagnosis was not the primary outcome, but cancer diagnosis or mortality data were available. As this systematic review was conducted as part of a series of systematic reviews assessing a range of health effects of omega-3, omega-6 and total PUFA38  we have been able to compare health benefits and harms across the major causes of mortality and morbidity in developed countries: cancer and cardiovascular disease.As previous systematic reviews of trials and observational studies have been equivocal about effects of omega-3, omega-6 and total PUFA on total, breast and prostate cancer risk,39 This review’s protocol was registered on PROSPERO40 and its specific methods are summarised below.Methods for the series have been reported in detail (including the PRISMA flow diagram and detailed search strategies).41 plus a further 6 months to influence cancer development.We included randomised controlled trials (RCTs) that compared higher versus lower LCn3, ALA, omega-6 and/or total PUFA in adults aged at least 18 years, who were not pregnant or seriously ill. Participants could be free of cancer, at increased risk of cancer or with a previous cancer diagnosis, but were excluded if they were currently undergoing cancer treatment. The minimum study duration was 1 year (≥52 weeks) reflecting metabolic studies suggesting 6 months is the minimum supplementation duration required to equilibrate LCn3 into most body compartments,Interventions could consist of foods, oral supplements  or advice, to increase or decrease omega-3, omega-6 and/or total PUFA intake, or achieve a change of ≥10% of baseline intake, comparing higher versus lower PUFA intake. Studies were excluded if they examined lifestyle or dietary interventions in addition to PUFA unless effects of the PUFA could be separated out.New diagnosis of breast cancerBreast cancer mortalityNew diagnosis of any cancerAny cancer mortalityPrimary outcomes included:Secondary outcomes included prostate cancer diagnosis and mortality (added post-hoc to complement prostate-specific antigen (PSA) data), markers of cancer risk (including breast density and PSA), body weight and measures of adiposity, quality of life, and dropouts.39 From this database, trials with duration of at least 12 months and data collected on any primary outcome were included in this review, even if study objectives were not primarily to assess effects on cancer, or those outcomes were not published.We searched Cochrane CENTRAL, Medline and Embase to 27 April 2017, ClinicalTrials.com and WHO International Clinical Trials Registry Platform to September 2016 and reassessed all ongoing trials in December 2018. We checked included trials of relevant systematic reviews, and wrote to authors of included studies for additional trial data, creating a database of trials that randomised participants to increased omega-3, omega-6 or total PUFA compared to lower omega-3, omega-6 or total PUFA.42 plus risk from compliance problems and attention bias.39 We considered supplementation trials to be at low summary risk of bias where randomisation, allocation concealment, blinding of participants, personnel and outcome assessors were judged adequate . Dietary advice trials were at low summary risk of bias where randomisation, allocation concealment and blinding of outcome assessors were assessed adequate.39Study inclusion, data extraction and risk of bias assessment  were conducted independently in duplicate. We assessed Cochrane risk of bias domains43) in Review Manager 5.3.44 Treatment/control differences in outcomes were combined across studies using risk ratios (RR) or mean differences (MD), the at-risk population included only men for prostate cancer and women for breast cancer. Change from baseline in each arm with standard deviations were used for continuous outcomes where available, otherwise endpoint data were used.43 Pre-specified sensitivity analyses included fixed effects analysis, limiting analysis to studies at low summary risk of bias, and limiting to trials randomising ≥100 participants. At the request of our funders, we added sensitivity analyses limiting to studies at low risk for compliance issues. At the request of referees, we added sensitivity analyses using Peto fixed-effects analysis (creating odds ratios), to ensure that our findings are robust to analysis methods despite the inclusion of trials with rare events. Pre-specified subgroup analyses were conducted for outcomes with ≥8 studies by intervention type, replacement, dose, duration, age, sex and cancer risk .39 We planned to sub-group also by medications used, baseline omega-3, omega-6 or total PUFA intake, pre- or post-menopausal, BMI, ethnicity and omega-3/omega-6 ratio, however, this information was not available in most trials, so subgrouping was not attempted. Heterogeneity was assessed using I2 and considered important where >50%.45 Small study bias was assessed using funnel plots where at least 10 trials were included in a meta-analysis.46 Data from individual participants were only counted once in any meta-analytical pooling.Primary analyses assessed effects of total PUFA, omega-6, LCn3 and ALA using random-effects Mantel–Haenszel meta-analysis , while a mean difference between arms of at least 10% of baseline was required for a relevant clinical effect for markers. Where we found a suggested effect we quantified the effect using number needed to treat for an additional benefit (NNTB) or number needed to treat to cause an additional harm (NNTH).47 Outcome data were interpreted using GRADE assessment, drafted by LH then discussed and agreed with WHO NUGAG.39 Where sensitivity analyses using Mantel–Haenszel or Peto fixed-effects analyses were not consistent with the main random-effects analysis we downgraded (for inconsistency), and where sensitivity analyses including only trials at low summary risk of bias, or only trials with good compliance differed from the main analysis we downgraded (for risk of bias). Where GRADE suggested data of very low-quality we did not interpret effect sizes. Where data were of low-quality, we used the term “may”, moderate-quality evidence warranted “probably” in describing effects. Summary of findings’ (GRADE) tables show effects on all cancers, breast cancer and prostate cancer diagnoses and deaths (marker evidence strengthened or weakened findings for relevant cancers).Effect sizes were interpreted as agreed with the WHO Nutrition Guidance Expert Advisory Group (NUGAG) Subgroup on Diet and Health who commissioned this research.We included 47 RCTs (49 comparisons). Thirty-four trials  assessed effects of LCn3, three (3179 participants) assessed ALA, eight (4976 participants) assessed omega-6 and 9 trials  assessed total PUFA  and probably has little or no effect on risk of cancer death . We meta-analysed 27 trials  assessing effects of LCn3 on cancer diagnosis , but effects on breast cancer deaths are unclear as the evidence is of very low-quality (two deaths). We meta-analysed 12 trials  assessing effects of LCn3 on breast cancer diagnosis , but effects on prostate cancer death were unclear . Seven trials  reported on 1021 prostate cancer diagnoses, finding higher risk of prostate cancer in men with increased LCn3  and may slightly increase the risk of prostate cancer diagnosis . Data on any cancer diagnoses, breast cancer diagnoses, breast or prostate cancer deaths and breast density were too limited to provide useful information, so effects were unclear.2 0%), which did not alter in sensitivity analyses. The same two trials reported 46 prostate cancer diagnoses in 4010 male participants . This increased risk was consistent across all sensitivity analyses and supported by a rise in PSA in those taking more ALA in the single large trial .Two trials  provided data on 123 cancer deaths and meta-analysis suggested little or no effect . No trials reported breast cancer deaths or breast density, prostate cancer deaths or PSA and effects on breast and prostate cancer diagnoses were unclear .2 0%), consistent across sensitivity analysis. While the funnel plot suggested small trials with higher risk ratios may be missing , consistent across all sensitivity analyses.Eight trials  assessed effects of increasing total PUFA on cancer diagnosis . Increasing ALA probably has little or no effect on risk of cancer death but may slightly increase prostate cancer risk (NNTH 334). Effects of omega-6 were unclear. Increasing total PUFA may slightly increase risk of diagnosis of any cancer (NNTH 125) and cancer death (NNTH 500).51 This ensures that review results are robust to analysis methods.Strengths of this systematic review include its large size . Creation of a dataset of RCTs randomising to higher or lower PUFA intakes, regardless of primary and reported outcomes, allowed the inclusion of trials and data that would otherwise have been missed or remained unpublished. This allowed us to include many large and long-term RCTs of PUFAs in populations recruited for health problems other than cancer risk, so allowing us to assess effects of increasing PUFA on diagnosis of cancers in low-risk populations. As meta-analysis of trials with rare events can produce different effect sizes when using different analytical methods we ran sensitivity analyses using Mantel–Haenszel and Peto fixed-effects meta-analyses and compared the results with the main random-effects Mantel–Haenszel analysis.52 and lack of blinding with additional bias54 caution is needed in interpreting small effects in weaker trials. As prostate cancer was not a primary outcome in this review, we did not ask trialists for additional prostate cancer data, which means that more information on prostate cancer may be available from existing trials.Review limitations include limited available data on effects of increasing ALA, omega-6 and total PUFA. It was notable that doses of total PUFA were highly variable , but the small number of trials made subgrouping by dose uninformative  suggesting LCn3 intake is not associated with total cancer risk.16 Two previous systematic reviews of trials and observational data suggested there were inadequate data to determine whether LCn3 intake was associated with prostate cancer incidence or progression.29 A systematic review of cohort studies assessing relationships between omega-3 and eleven types of cancer found mixed results, including cohorts suggesting both statistically significantly increased and decreased risk for prostate cancer.23 This review is new in suggesting that actively increasing dietary total PUFA may slightly increase the risk of both cancer diagnosis and cancer mortality. A recent systematic review of observational studies suggested no association between total polyunsaturated fat intake and breast cancer risk,22 but as higher PUFA intake is associated with healthier lifestyles small harms may be difficult to spot in observational studies due to confounding. RCT data are insufficient to corroborate or contradict two nested case-control trials suggesting that higher PUFA intake correlates to higher prostate cancer risk.25Our review concurs with a systematic review of observational data55 The balance appears similar for ALA—for every 1000 people increasing their ALA intake two will avoid a CVD event, eleven will avoid arrhythmia but three will be diagnosed with prostate cancer who would not otherwise have been diagnosed , trial evidence suggests that increasing omega-3 may slightly increase prostate cancer risk, and increasing total PUFA may slightly increase cancer risk , although this could result from very high intakes of PUFA in some trials. Considering both cancer and cardiovascular outcomes, overall health effects of increasing LCn3, ALA, omega-6 and total PUFA appear small.Supplementary file Ca & PUFA