Analysis of rare disruptive germline mutations in 2135 enriched BRCA-negative breast cancers excludes additional high-impact susceptibility genes.

BACKGROUND: Breast cancer has a significant heritable basis, of which ∼60% remains unexplained. Testing for BRCA1/BRCA2 offers useful discrimination of breast cancer risk within families, and identification of additional breast cancer susceptibility genes could offer clinical utility. PATIENTS AND METHODS: We included 2135 invasive breast cancer cases recruited via the Breast and Ovarian Cancer Susceptibility study, a retrospective UK study of familial breast cancer. ELIGIBILITY CRITERIA: female, BRCA-negative, white European ethnicity, and one of: (i) breast cancer family history, (ii) bilateral disease, (iii) young age of onset (<30 years), and (iv) concomitant ovarian cancer. We undertook exome sequencing of cases and carried out gene-level burden testing of rare damaging variants against those from 51 377 ethnicity-matched population controls from gnomAD. RESULTS: 159/2135 (7.4%) cases had a qualifying variant in an established breast cancer susceptibility gene, with minimal evidence of signal in other cancer susceptibility genes. Known breast cancer susceptibility genes PALB2, CHEK2, and ATM were the only genes to retain statistical significance after correcting for multiple testing. Due to the enrichment of hereditary cases in the series, we had good power (>80%) to detect a gene of BRCA1-like risk [odds ratio (OR) = 10.6] down to a population minor allele frequency of 4.6 × 10-5 (1 in 10 799, less than one-tenth that of BRCA1)and of PALB2-like risk (OR = 5.0) down to a population minor allele frequency of 2.8 × 10-4 (1 in 1779, less than half that of PALB2). Power was lower for identification of novel moderate penetrance genes (OR = 2-3) like CHEK2 and ATM. CONCLUSIONS: This is the largest case-control whole-exome analysis of enriched breast cancer published to date. Whilst additional breast cancer susceptibility genes likely exist, those of high penetrance are likely to be of very low mutational frequency. Contention exists regarding the clinical utility of such genes.

Clinical likelihood ratios and balanced accuracy for 44 in silico tools against multiple large-scale functional assays of cancer susceptibility genes.

PURPOSE: Where multiple in silico tools are concordant, the American College of Medical Genetics and Genomics/Association for Molecular Pathology (ACMG/AMP) framework affords supporting evidence toward pathogenicity or benignity, equivalent to a likelihood ratio of ~2. However, limited availability of "clinical truth sets" and prior use in tool training limits their utility for evaluation of tool performance. METHODS: We created a truth set of 9,436 missense variants classified as deleterious or tolerated in clinically validated high-throughput functional assays for BRCA1, BRCA2, MSH2, PTEN, and TP53 to evaluate predictive performance for 44 recommended/commonly used in silico tools. RESULTS: Over two-thirds of the tool-threshold combinations examined had specificity of <50%, thus substantially overcalling deleteriousness. REVEL scores of 0.8-1.0 had a Positive Likelihood Ratio (PLR) of 6.74 (5.24-8.82) compared to scores <0.7 and scores of 0-0.4 had a Negative Likelihood Ratio (NLR) of 34.3 (31.5-37.3) compared to scores of >0.7. For Meta-SNP, the equivalent PLR = 42.9 (14.4-406) and NLR = 19.4 (15.6-24.9). CONCLUSION: Against these clinically validated "functional truth sets," there was wide variation in the predictive performance of commonly used in silico tools. Overall, REVEL and Meta-SNP had best balanced accuracy and might potentially be used at stronger evidence weighting than current ACMG/AMP prescription, in particular for predictions of benignity.

Genetic testing in a cohort of young patients with HER2-amplified breast cancer.

BACKGROUND: Young age at diagnosis for breast cancer raises the question of genetic susceptibility. We explored breast cancer susceptibility genes testing on ≤40-year-old patients with HER2-amplified invasive breast cancer. PATIENTS AND METHODS: Patients were selected from a large UK cohort study. The inclusion criterion was age ≤40 at diagnosis with confirmed HER2-amplified breast cancer. The probability of finding a BRCA gene mutation was calculated based on family history. Genetic testing used was either clinical testing for BRCA1 and BRCA2, with a subset also tested for TP53 mutations, or research-based testing using a typical panel comprising 17 breast cancer susceptibility genes (CSGs) including BRCA1, BRCA2 and TP53. RESULTS: Of the 591 eligible patients, clinical testing results were available for 133 cases and an additional 263 cases had panel testing results. BRCA testing across 396 cases found 8 BRCA2 (2%) and 6 BRCA1 (2%) pathogenic mutations. Of the 304 patients tested for TP53 mutations, overall 9 (3%) had deleterious TP53 mutations. Of the 396 patients, 101 (26%) met clinical criteria for BRCA testing (≥10% probability), among whom 11% had pathogenic BRCA mutations (6 BRCA2, 5 BRCA1). Where the probability was calculated to be <10%, only 4 of 295 (1%) patients had BRCA mutations. Among the 59 patients who had TP53 testing meeting the 10% threshold, 7 had mutations (12%). Likely functionally deleterious mutations in 14 lower penetrance CSGs were present in 12 of 263 (5%) panel-tested patients. CONCLUSION: Patients aged <41 at diagnosis with HER2+ breast cancer and no family history of breast cancer can be reassured that they have a low chance of being a high-risk gene carrier. If there is a strong family history, not only BRCA but also TP53 gene testing should be considered. The clinical utility of testing lower penetrance CSGs remains unclear.

An evaluation of the prognostic model PREDICT using the POSH cohort of women aged ⩽40 years at breast cancer diagnosis.

BACKGROUND: Breast cancer is the most common cancer in younger women (aged ⩽40 years) in the United Kingdom. PREDICT (http://www.predict.nhs.uk) is an online prognostic tool developed to help determine the best available treatment and outcome for early breast cancer. This study was conducted to establish how well PREDICT performs in estimating survival in a large cohort of younger women recruited to the UK POSH study. METHODS: The POSH cohort includes data from 3000 women aged ⩽40 years at breast cancer diagnosis. Study end points were overall and breast cancer-specific survival at 5, 8, and 10 years. Evaluation of PREDICT included model discrimination and comparison of the number of predicted versus observed events. RESULTS: PREDICT provided accurate long-term (8- and 10-year) survival estimates for younger women. Five-year estimates were less accurate, with the tool overestimating survival by 25% overall, and by 56% for patients with oestrogen receptor (ER)-positive tumours. PREDICT underestimated survival at 5 years among patients with ER-negative tumours. CONCLUSIONS: PREDICT is a useful tool for providing reliable long-term (10-year) survival estimates for younger patients. However, for more accurate short-term estimates, the model requires further calibration using more data from young onset cases. Short-term prediction may be most relevant for the increasing number of women considering risk-reducing bilateral mastectomy.

BRCA1 and BRCA2 genetic testing-pitfalls and recommendations for managing variants of uncertain clinical significance.

BACKGROUND: Increasing use of BRCA1/2 testing for tailoring cancer treatment and extension of testing to tumour tissue for somatic mutation is moving BRCA1/2 mutation screening from a primarily prevention arena delivered by specialist genetic services into mainstream oncology practice. A considerable number of gene tests will identify rare variants where clinical significance cannot be inferred from sequence information alone. The proportion of variants of uncertain clinical significance (VUS) is likely to grow with lower thresholds for testing and laboratory providers with less experience of BRCA. Most VUS will not be associated with a high risk of cancer but a misinterpreted VUS has the potential to lead to mismanagement of both the patient and their relatives. DESIGN: Members of the Clinical Working Group of ENIGMA (Evidence-based Network for the Interpretation of Germline Mutant Alleles) global consortium (www.enigmaconsortium.org) observed wide variation in practices in reporting, disclosure and clinical management of patients with a VUS. Examples from current clinical practice are presented and discussed to illustrate potential pitfalls, explore factors contributing to misinterpretation, and propose approaches to improving clarity. RESULTS AND CONCLUSION: Clinicians, patients and their relatives would all benefit from an improved level of genetic literacy. Genetic laboratories working with clinical geneticists need to agree on a clinically clear and uniform format for reporting BRCA test results to non-geneticists. An international consortium of experts, collecting and integrating all available lines of evidence and classifying variants according to an internationally recognized system, will facilitate reclassification of variants for clinical use.

Obesity and the outcome of young breast cancer patients in the UK: the POSH study.

BACKGROUND: Obese breast cancer patients have a poorer prognosis than non-obese patients. We examined data from a large prospective cohort study to explore the associations of obesity with tumour pathology, treatment and outcome in young British breast cancer patients receiving modern oncological treatments. PATIENTS AND METHODS: A total of 2956 patients aged ≤40 at breast cancer diagnosis were recruited from 126 UK hospitals from 2001 to 2007. Height and weight were measured at registration. Tumour pathology and treatment details were collected. Follow-up data were collected at 6, 12 months, and annually. RESULTS: A total of 2843 eligible patients (96.2%) had a body mass index (BMI) recorded: 1526 (53.7%) were under/healthy-weight (U/H, BMI <25 kg/m(2)), 784 (27.6%) were overweight (ov, BMI ≥25 to <30), and 533 (18.7%) were obese (ob, BMI ≥30). The median tumour size was significantly higher in obese and overweight patients than U/H patients (Ob 26 mm versus U/H 20 mm, P < 0.001; Ov 24 mm versus U/H 20 mm, P < 0.001). Obese and overweight patients had significantly more grade 3 tumours (63.9% versus 59.0%, P = 0.048; Ov 63.6% versus U/H 59.0% P = 0.034) and node-positive tumours (Ob 54.6% versus U/H 49.0%, P = 0.027; Ov 54.2% versus U/H 49%, P = 0.019) than U/H patients. Obese patients had more ER/PR/HER2-negative tumours than healthy-weight patients (25.0% versus 18.3%, P = 0.001). Eight-year overall survival (OS) and distant disease-free interval (DDFI) were significantly lower in obese patients than healthy-weight patients [OS: hazard ratio (HR) 1.65, P < 0.001; DDFI: HR 1.44, P < 0.001]. Multivariable analyses adjusting for tumour grade, size, nodal, and HER2 status indicated that obesity was a significant independent predictor of OS and DDFI in patients with ER-positive disease. CONCLUSIONS: Young obese breast cancer patients present with adverse tumour characteristics. Despite adjustment for this, obesity still independently predicts DDFI and OS.

Tamoxifen metabolism predicts drug concentrations and outcome in premenopausal patients with early breast cancer.

Tamoxifen is the standard-of-care treatment for estrogen receptor-positive premenopausal breast cancer. We examined tamoxifen metabolism via blood metabolite concentrations and germline variations of CYP3A5, CYP2C9, CYP2C19 and CYP2D6 in 587 premenopausal patients (Asians, Middle Eastern Arabs, Caucasian-UK; median age 39 years) and clinical outcome in 306 patients. N-desmethyltamoxifen (DM-Tam)/(Z)-endoxifen and CYP2D6 phenotype significantly correlated across ethnicities (R(2): 53%, P<10(-77)). CYP2C19 and CYP2C9 correlated with norendoxifen and (Z)-4-hydroxytamoxifen concentrations, respectively (P<0.001). DM-Tam was influenced by body mass index (P<0.001). Improved distant relapse-free survival (DRFS) was associated with decreasing DM-Tam/(Z)-endoxifen (P=0.036) and increasing CYP2D6 activity score (hazard ratio (HR)=0.62; 95% confidence interval (CI), 0.43-0.91; P=0.013). Low (<14 nM) compared with high (>35 nM) endoxifen concentrations were associated with shorter DRFS (univariate P=0.03; multivariate HR=1.94; 95% CI, 1.04-4.14; P=0.064). Our data indicate that endoxifen formation in premenopausal women depends on CYP2D6 irrespective of ethnicity. Low endoxifen concentration/formation and decreased CYP2D6 activity predict shorter DRFS.

Family history and outcome of young patients with breast cancer in the UK (POSH study).

BACKGROUND: Young patients presenting to surgical clinics with breast cancer are usually aware of their family history and frequently believe that a positive family history may adversely affect their prognosis. Tumour pathology and outcomes were compared in young British patients with breast cancer with and without a family history of breast cancer. METHODS: Prospective Outcomes in Sporadic versus Hereditary breast cancer (POSH) is a large prospective cohort study of women aged less than 41 years with breast cancer diagnosed and treated in the UK using modern oncological management. Personal characteristics, tumour pathology, treatment and family history of breast/ovarian cancer were recorded. Follow-up data were collected annually. RESULTS: Family history data were available for 2850 patients. No family history was reported by 65·9 per cent, and 34·1 per cent reported breast/ovarian cancer in at least one first- or second-degree relative. Patients with a family history were more likely to have grade 3 tumours (63·3 versus 58·9 per cent) and less likely to have human epidermal growth factor receptor 2-positive tumours (24·7 versus 28·8 per cent) than those with no family history. In multivariable analyses, there were no significant differences in distant disease-free intervals for patients with versus those without a family history, either for the whole cohort (hazard ratio (HR) 0·89, 95 per cent c.i. 0·76 to 1·03; P = 0·120) or when stratified by oestrogen receptor (ER) status (ER-negative: HR 0·80, 0·62 to 1·04, P = 0·101; ER-positive: HR 0·95, 0·78 to 1·15, P = 0·589). CONCLUSION: Young British patients presenting to breast surgical clinics with a positive family history can be reassured that this is not a significant independent risk factor for breast cancer outcome.

Understanding of BRCA VUS genetic results by breast cancer specialists.

BACKGROUND: Mainstreaming genetic medicine, increased media coverage and clinical trials for BRCA mutation carriers are leading oncologists into more patient discussions about BRCA genetic testing. BRCA variants of uncertain significance (VUS) occur in 10-20% of tests. VUS detection introduces additional uncertainty for patient and potentially clinician. We aimed to explore the ability of breast cancer specialists (BCS) in the UK to correctly respond to a VUS report. METHODS: A survey sent to 800 UK BCS collected demographics data, VUS general knowledge and interpretation and communication based on two genetics reports. A separate survey of UK clinical geneticists collected demographics data, laboratory reporting practice and methods used to clarify VUS pathogenicity including classification systems. RESULTS: Of the 155 BCS (22.5%) who completed the survey, 12% reported no genetics training. Ninety five percent referred patients for BRCA genetic tests, 71% felt unsure about the clinical implications of the test reports presented here. A VUS report from a patient with a positive family history was interpreted and theoretically communicated correctly by 94% but when presented with a different VUS report with no management guidance and negative family history, 39% did not know how to communicate this result to the patient. Geneticists reported multiple VUS classification systems; the most commonly used was word-based in 32%. CONCLUSIONS: A consistent and standardised format to report particularly VUS results across all diagnostic laboratories plus additional training of UK BCS will be necessary for effective mainstreaming of BRCA testing to the oncology clinic.

Serum bilirubin concentration is modified by UGT1A1 haplotypes and influences risk of type-2 diabetes in the Norfolk Island genetic isolate.

BACKGROUND: Located in the Pacific Ocean between Australia and New Zealand, the unique population isolate of Norfolk Island has been shown to exhibit increased prevalence of metabolic disorders (type-2 diabetes, cardiovascular disease) compared to mainland Australia. We investigated this well-established genetic isolate, utilising its unique genomic structure to increase the ability to detect related genetic markers. A pedigree-based genome-wide association study of 16 routinely collected blood-based clinical traits in 382 Norfolk Island individuals was performed. RESULTS: A striking association peak was located at chromosome 2q37.1 for both total bilirubin and direct bilirubin, with 29 SNPs reaching statistical significance (P < 1.84 × 10(-7)). Strong linkage disequilibrium was observed across a 200 kb region spanning the UDP-glucuronosyltransferase family, including UGT1A1, an enzyme known to metabolise bilirubin. Given the epidemiological literature suggesting negative association between CVD-risk and serum bilirubin we further explored potential associations using stepwise multivariate regression, revealing significant association between direct bilirubin concentration and type-2 diabetes risk. In the Norfolk Island cohort increased direct bilirubin was associated with a 28% reduction in type-2 diabetes risk (OR: 0.72, 95% CI: 0.57-0.91, P = 0.005). When adjusted for genotypic effects the overall model was validated, with the adjusted model predicting a 30% reduction in type-2 diabetes risk with increasing direct bilirubin concentrations (OR: 0.70, 95% CI: 0.53-0.89, P = 0.0001). CONCLUSIONS: In summary, a pedigree-based GWAS of blood-based clinical traits in the Norfolk Island population has identified variants within the UDPGT family directly associated with serum bilirubin levels, which is in turn implicated with reduced risk of developing type-2 diabetes within this population.

BRCA1 testing should be offered to individuals with triple-negative breast cancer diagnosed below 50 years.

BACKGROUND: Triple-negative (TN) tumours are the predominant breast cancer subtype in BRCA1 mutation carriers. Recently, it was proposed that all individuals below 50 years of age with TN breast cancer should be offered BRCA testing. We have evaluated the BRCA1 mutation frequency and the implications for clinical practice of undertaking genetic testing in women with TN breast cancer. METHODS: We undertook BRCA1 mutation analysis in 308 individuals with TN breast cancer, 159 individuals from unselected series of breast cancer and 149 individuals from series ascertained on the basis of young age and/or family history. RESULTS: BRCA1 mutations were present in 45 out of 308 individuals. Individuals with TN cancer <50 years had >10% likelihood of carrying a BRCA1 mutation in both the unselected (11 out of 58, 19%) and selected (26 out of 111, 23%) series. However, over a third would not have been offered testing using existing criteria. We estimate that testing all individuals with TN breast cancer <50 years would generate an extra 1200 tests annually in England. CONCLUSION: Women with TN breast cancer diagnosed below 50 years have >10% likelihood of carrying a BRCA1 mutation and are therefore eligible for testing in most centres. However, implementation may place short-term logistical and financial burdens on genetic services.

Evaluation of the XRCC1 gene as a phenotypic modifier in BRCA1/2 mutation carriers. Results from the consortium of investigators of modifiers of BRCA1/BRCA2.

BACKGROUND: Single-nucleotide polymorphisms (SNPs) in genes involved in DNA repair are good candidates to be tested as phenotypic modifiers for carriers of mutations in the high-risk susceptibility genes BRCA1 and BRCA2. The base excision repair (BER) pathway could be particularly interesting given the relation of synthetic lethality that exists between one of the components of the pathway, PARP1, and both BRCA1 and BRCA2. In this study, we have evaluated the XRCC1 gene that participates in the BER pathway, as phenotypic modifier of BRCA1 and BRCA2. METHODS: Three common SNPs in the gene, c.-77C>T (rs3213245) p.Arg280His (rs25489) and p.Gln399Arg (rs25487) were analysed in a series of 701 BRCA1 and 576 BRCA2 mutation carriers. RESULTS: An association was observed between p.Arg280His-rs25489 and breast cancer risk for BRCA2 mutation carriers, with rare homozygotes at increased risk relative to common homozygotes (hazard ratio: 22.3, 95% confidence interval: 14.3-34, P<0.001). This association was further tested in a second series of 4480 BRCA1 and 3016 BRCA2 mutation carriers from the Consortium of Investigators of Modifiers of BRCA1 and BRCA2. CONCLUSIONS AND INTERPRETATION: No evidence of association was found when the larger series was analysed which lead us to conclude that none of the three SNPs are significant modifiers of breast cancer risk for mutation carriers.

Targeted prostate cancer screening in men with mutations in BRCA1 and BRCA2 detects aggressive prostate cancer: preliminary analysis of the results of the IMPACT study.

OBJECTIVE: To evaluate the role of targeted prostate cancer screening in men with BRCA1 or BRCA2 mutations, an international study, IMPACT (Identification of Men with a genetic predisposition to ProstAte Cancer: Targeted screening in BRCA1/2 mutation carriers and controls), was established. This is the first multicentre screening study targeted at men with a known genetic predisposition to prostate cancer. A preliminary analysis of the data is reported. PATIENTS AND METHODS: Men aged 40-69 years from families with BRCA1 or BRCA2 mutations were offered annual prostate specific antigen (PSA) testing, and those with PSA > 3 ng/mL, were offered a prostate biopsy. Controls were men age-matched (± 5 years) who were negative for the familial mutation. RESULTS: In total, 300 men were recruited (205 mutation carriers; 89 BRCA1, 116 BRCA2 and 95 controls) over 33 months. At the baseline screen (year 1), 7.0% (21/300) underwent a prostate biopsy. Prostate cancer was diagnosed in ten individuals, a prevalence of 3.3%. The positive predictive value of PSA screening in this cohort was 47·6% (10/21). One prostate cancer was diagnosed at year 2. Of the 11 prostate cancers diagnosed, nine were in mutation carriers, two in controls, and eight were clinically significant. CONCLUSIONS: The present study shows that the positive predictive value of PSA screening in BRCA mutation carriers is high and that screening detects clinically significant prostate cancer. These results support the rationale for continued screening in such men.

A novel HER2-positive breast cancer phenotype arising from germline TP53 mutations.

INTRODUCTION: The Li-Fraumeni Syndrome is caused by a germline TP53 mutation and is associated with a high risk of breast cancer at young ages. Basal (triple negative) breast cancers are now well recognised to be a typical sub-type of breast cancer developing in a large proportion of BRCA1 gene carriers. We considered whether a similar narrow sub-type of breast cancer was found in TP53 gene mutation carriers. OBJECTIVE: A hypothesis generating study to investigate whether there are specific breast tumour characteristics associated with germline TP53 mutations. METHODS: Pathological characteristics in 12 breast cancers arising in nine patients carrying pathogenic TP53 mutations were compared to a reference panel of 231 young onset breast tumours included in the POSH study. RESULTS: Patients carrying a TP53 mutation showed a significantly higher likelihood of developing a breast cancer with Human Epidermal growth factor Receptor (HER2) amplification (83%) when compared to the cohort of young onset breast cancer cases (16%); ER and PR status were equivalent between groups. CONCLUSION: These findings suggest that breast cancer developing on a background of an inherited TP53 mutation is highly likely to present with amplification of HER2.

Risk reducing mastectomy: outcomes in 10 European centres.

BACKGROUND: Increasingly women at high risk of breast cancer are opting for risk reducing surgery. The aim of this study was to assess the effectiveness of this approach in women at high risk in both carriers and non-carriers of BRCA1/2. METHODS: Data from 10 European centres that offer a genetic counselling and screening service to women at risk were obtained prospectively from 1995. Breast cancer risks were estimated from life tables and a control group of women at risk who did not undergo surgery. RESULTS: The combined centres have data on 550 women who have undergone risk reducing mastectomy with greater than 3334 women years of follow-up. Operations were carried out on women with lifetime risks of 25-80%, with an average expected incidence rate of 1% per year. No breast cancers have occurred in this cohort in the "at risk" unaffected breast, whereas >34 would have been expected. A high rate (2-3.6%) of occult disease was identified in the at risk breast at the time of surgery. INTERPRETATION: We conclude that risk reducing surgery is highly effective.

In vitro dual perfusion of human placental lobules as a flow phantom to investigate the relationship between fetoplacental flow and quantitative 3D power doppler angiography.

Flow phantoms have been used to investigate and quantify three-dimensional power Doppler data but this is the first study to use the in vitro, dual perfused, placental perfusion model. We used this model to investigate and quantify the effect of variation in fetal-side flow rates and attenuation on 3D power Doppler angiography. Perfusion of a placental lobule was commenced within 30 min of delivery and experimentation was successful in 8 of the 18 placenta obtained. Fetal and maternal perfusate was modified Earle's bicarbonate buffer which, following equilibration, was supplemented on the fetal side with whole heparinised cord blood. Imaging was performed with a Voluson-i ultrasound machine. A 'vascular biopsy' the thickness of the placental lobule was defined and signal quantified within using VOCAL (GE Medical Systems, Zipf, Austria). Three vascular indices are generated: vascularisation index (VI) defined as the percentage of power Doppler data within a volume of interest; flow index (FI), the mean signal intensity of the power Doppler information; and vascularisation flow index (VFI), a combination of both factors derived through their multiplication. Attenuation was investigated in this model with the addition of tissue mimic blocks. Our results showed a predictable relationship between flow rates and the vascular indices VI and VFI. However the FI was a less reliable predictor of flow; thus it should be interpreted with caution. The power Doppler signal was markedly affected by attenuation leading to a complete loss of information at a depth of 6 cm in the model used. In conclusion this model can be adapted to provide a phantom to analyse and quantify 3D power Doppler signals and demonstrates that vascular indices within a tissue remain related to volume flow. This model provides further evidence that depth dependent attenuation of signal needs to be accounted for in any in vivo work where the probe is not in direct contact with the tissue of interest.

Predicting the likelihood of carrying a BRCA1 or BRCA2 mutation: validation of BOADICEA, BRCAPRO, IBIS, Myriad and the Manchester scoring system using data from UK genetics clinics.

OBJECTIVES: Genetic testing for the breast and ovarian cancer susceptibility genes BRCA1 and BRCA2 has important implications for the clinical management of people found to carry a mutation. However, genetic testing is expensive and may be associated with adverse psychosocial effects. To provide a cost-efficient and clinically appropriate genetic counselling service, genetic testing should be targeted at those individuals most likely to carry pathogenic mutations. Several algorithms that predict the likelihood of carrying a BRCA1 or a BRCA2 mutation are currently used in clinical practice to identify such individuals. DESIGN: We evaluated the performance of the carrier prediction algorithms BOADICEA, BRCAPRO, IBIS, the Manchester scoring system and Myriad tables, using 1934 families seen in cancer genetics clinics in the UK in whom an index patient had been screened for BRCA1 and/or BRCA2 mutations. The models were evaluated for calibration, discrimination and accuracy of the predictions. RESULTS: Of the five algorithms, only BOADICEA predicted the overall observed number of mutations detected accurately (ie, was well calibrated). BOADICEA also provided the best discrimination, being significantly better (p<0.05) than all models except BRCAPRO (area under the receiver operating characteristic curve statistics: BOADICEA = 0.77, BRCAPRO = 0.76, IBIS = 0.74, Manchester = 0.75, Myriad = 0.72). All models underpredicted the number of BRCA1 and BRCA2 mutations in the low estimated risk category. CONCLUSIONS: Carrier prediction algorithms provide a rational basis for counselling individuals likely to carry BRCA1 or BRCA2 mutations. Their widespread use would improve equity of access and the cost-effectiveness of genetic testing.

Inherited predisposition to colorectal adenomas caused by multiple rare alleles of MUTYH but not OGG1, NUDT1, NTH1 or NEIL 1, 2 or 3.

BACKGROUND: MUTYH-associated polyposis (MAP) is a recessive trait characterised by multiple colorectal adenomas and a high risk of colorectal cancer. MUTYH functions in the DNA base excision repair pathway and has a key role in the repair of oxidative DNA damage. OBJECTIVES: To assess the contribution of inherited variants in genes involved in base excision repair and oxidative DNA damage including MUTYH, OGG1, NEIL1, NEIL2, NEIL3, NUDT1 and NTH1 to the multiple colorectal adenoma phenotype. METHODS: Inherited variants of MUTYH, OGG1, NEIL1, NEIL2, NEIL3, NUDT1 and NTH1 were sought in 167 unrelated patients with multiple colorectal adenomas whose family histories were consistent with recessive inheritance. These variants were also characterised in approximately 300 population controls. RESULTS: Thirty-three patients (20%) and no controls were MUTYH homozygotes or compound heterozygotes (ie, carried two mutations) and therefore had MAP. Eight different pathogenic MUTYH mutations were identified, of which four were novel. MAP cases had significantly more adenomas than non-MAP cases (p = 0.0009; exact test for trends in proportions) and presented earlier (p = 0.013; analysis of variance). Twenty-four protein-altering variants were identified upon screening of OGG1, NEIL1, NEIL2, NEIL3, NUDT1 and NTH1. However, all combinations of two (or more) variants that were identified at an individual locus in patients were also seen in controls, and no variants were significantly over-represented (or under-represented) in cases. CONCLUSION: Multiple rare alleles of MUTYH are associated with autosomal recessive MAP, while OGG1, NEIL1, NEIL2, NEIL3, NUDT1 and NTH1 do not contribute significantly to autosomal recessive polyposis.

Ion torrent high throughput mitochondrial genome sequencing (HTMGS).

The implementation and popularity of next generation sequencing (NGS) has led to the development of various rapid whole mitochondrial genome sequencing techniques. We summarise an efficient and cost-effective NGS approach for mitochondrial genomic DNA in humans using the Ion Torrent platform, and further discuss our bioinformatics pipeline for streamlined variant calling. Ion 316 chips were utilised with the Ion Torrent semi-conductor platform Personal Genome Machine (PGM) to perform tandem sequencing of mitochondrial genomes from the core pedigree (n = 315) of the Norfolk Island Health Study. Key improvements from commercial methods focus on the initial PCR step, which currently requires extensive optimisation to ensure the accurate and reproducible elongation of each section of the complete mitochondrial genome. Dual-platform barcodes were incorporated into our protocol thereby extending its potential application onto Illumina-based systems. Our bioinformatics pipeline consists of a modified version of GATK best practices tailored for mitochondrial genomic data. When compared with current commercial methods, our method, termed high throughput mitochondrial genome sequencing (HTMGS), allows high multiplexing of samples and the use of alternate library preparation reagents at a lower cost per sample (~1.7 times) when compared to current commercial methodologies. Our HTMGS methodology also provides robust mitochondrial sequencing data (>450X average coverage) that can be applied and modified to suit various study designs. On average, we were able to identify ~30 variants per sample with 572 variants observed across 315 samples. We have developed a high throughput sequencing and analysis method targeting complete mitochondrial genomes; with the potential to be platform agnostic with analysis options that adhere to current best practices.

Final Results of the Prospective FH02 Mammographic Surveillance Study of Women Aged 35-39 at Increased Familial Risk of Breast Cancer.

BACKGROUND: Many women who are at increased risk of breast cancer due to a mother or sister diagnosed with breast cancer aged under 40 do not currently qualify for surveillance before 40 years of age. There are almost no available data to assess whether mammography screening aged 35-39 years would be effective in this group, in terms of detection of breast cancer at an early stage or cost effective. METHODS: A cohort screening study (FH02) with annual mammography was devised for women aged 35-39 to assess the sensitivity and screening performance and potential survival of women with identified tumours. FINDINGS: 2899 women were recruited from 12/2006-12/2015. These women underwent 12,086 annual screening mammograms and were followed for 13,365.8 years. A total of 55 breast cancers in 54 women occurred during the study period (one bilateral) with 50 cancers (49 women) (15 CIS) adherent to the screening. Eighty percent (28/35) of invasive cancers were ≤ 2 cm and 80% also lymph node negative. Invasive cancers diagnosed in FH02 were significantly smaller than the comparable (POSH-unscreened prospective) study group (45% (131/293) ≤ 2 cm in POSH vs 80% (28/35) in FH02 p < 0.0001), and were less likely to be lymph-node positive (54% (158/290, 3 unknown) in POSH vs 20% (7/35) in FH02: p = 0.0002. Projected and actual survival were also better than POSH. Overall radiation dose was not higher than in an older screened population at mean dose on study per standard sized breast of 1.5 mGy. INTERPRETATION: Mammography screening aged 35-39 years detects breast cancer at an early stage and is likely to be as effective in reducing mortality as in women at increased breast cancer risk aged 40-49 years.