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Purpose: A single maintenance course of a PARP inhibitor (PARPi) improves progression-free survival (PFS) in germline BRCA1/2-mutant high-grade serous ovarian cancer (gBRCAm-HGSOC). The feasibility of a second maintenance course of PARPi was unknown. Patients and Methods: Phase II trial with two entry points (EP1, EP2). Patients were recruited prior to rechallenge platinum. Patients with relapsed, gBRCAm-HGSOC were enrolled at EP1 if they were PARPi-naïve. Patients enrolled at EP2 had received their first course of olaparib prior to trial entry. EP1 patients were retreated with olaparib after RECIST complete/partial response (CR/PR) to platinum. EP2 patients were retreated with olaparib ± cediranib after RECIST CR/PR/stable disease to platinum and according to the platinum-free interval. Co-primary outcomes were the proportion of patients who received a second course of olaparib and the proportion who received olaparib retreatment for ≥6 months. Functional homologous recombination deficiency (HRD), somatic copy-number alteration (SCNA), and BRCAm reversions were investigated in tumor and liquid biopsies. Results: Twenty-seven patients were treated (EP1 = 17, EP2 = 10), and 19 were evaluable. Twelve patients (63%) received a second course of olaparib and 4 received olaparib retreatment for ≥6 months. Common grade ≥2 adverse events during olaparib retreatment were anemia, nausea, and fatigue. No cases of MDS/AML occurred. Mean duration of olaparib treatment and retreatment differed (12.1 months vs. 4.4 months; P < 0.001). Functional HRD and SCNA did not predict PFS. A BRCA2 reversion mutation was detected in a post-olaparib liquid biopsy. Conclusions: A second course of olaparib can be safely administered to women with gBRCAm-HGSOC but is only modestly efficacious.

BRCA2 reversion mutation detected in cell-free circulating DNA from patient 16.

Cell-free circulating DNA yields at each time point.

Purpose:A single maintenance course of a PARP inhibitor (PARPi) improves progression-free survival (PFS) in germline BRCA1/2-mutant high-grade serous ovarian cancer (gBRCAm-HGSOC). The feasibility of a second maintenance course of PARPi was unknown.Patients and Methods:Phase II trial with two entry points (EP1, EP2). Patients were recruited prior to rechallenge platinum. Patients with relapsed, gBRCAm-HGSOC were enrolled at EP1 if they were PARPi-naïve. Patients enrolled at EP2 had received their first course of olaparib prior to trial entry. EP1 patients were retreated with olaparib after RECIST complete/partial response (CR/PR) to platinum. EP2 patients were retreated with olaparib ± cediranib after RECIST CR/PR/stable disease to platinum and according to the platinum-free interval. Co-primary outcomes were the proportion of patients who received a second course of olaparib and the proportion who received olaparib retreatment for ≥6 months. Functional homologous recombination deficiency (HRD), somatic copy-number alteration (SCNA), and BRCAm reversions were investigated in tumor and liquid biopsies.Results:Twenty-seven patients were treated (EP1 = 17, EP2 = 10), and 19 were evaluable. Twelve patients (63%) received a second course of olaparib and 4 received olaparib retreatment for ≥6 months. Common grade ≥2 adverse events during olaparib retreatment were anemia, nausea, and fatigue. No cases of MDS/AML occurred. Mean duration of olaparib treatment and retreatment differed (12.1 months vs. 4.4 months; P < 0.001). Functional HRD and SCNA did not predict PFS. A BRCA2 reversion mutation was detected in a post-olaparib liquid biopsy.Conclusions:A second course of olaparib can be safely administered to women with gBRCAm-HGSOC but is only modestly efficacious.

Supplementary Methodology.

Liquid biopsy time points.

Kaplan Meier curves.

Cell-free circulating DNA copy-number profiles.

Serious adverse events.

Functional homologous recombination repair.

Missense variants in the NF2 gene result in variable NF2 disease presentation. Clinical classification of missense variants often represents a challenge, due to lack of evidence for pathogenicity and function. This study provides a summary of NF2 missense variants, with variant classifications based on currently available evidence. NF2 missense variants were collated from pathology-associated databases and existing literature. Association for Clinical Genomic Sciences Best Practice Guidelines (2020) were followed in the application of evidence for variant interpretation and classification. The majority of NF2 missense variants remain classified as variants of uncertain significance. However, NF2 missense variants identified in gnomAD occurred at a consistent rate across the gene, while variants compiled from pathology-associated databases displayed differing rates of variation by exon of NF2. The highest rate of NF2 disease-associated variants was observed in exon 7, while lower rates were observed toward the C-terminus of the NF2 protein, merlin. Further phenotypic information associated with variants, alongside variant-specific functional analysis, is necessary for more definitive variant interpretation. Our data identified differences in frequency of NF2 missense variants by exon between gnomAD population data and NF2 disease-associated variants, suggesting a potential genotype-phenotype correlation; further work is necessary to substantiate this.

National guidelines recommend testing all cases of non-mucinous epithelial ovarian cancer (NMEOC) for germline (blood) and somatic (tumour) BRCA1/2 pathogenic variants (PVs). We performed paired germline and somatic BRCA1/2 testing in consecutive cases of NMEOC (n = 388) to validate guidelines. Thirty-four somatic BRCA1/2 (sBRCA) PVs (9.7%) were detected in 350 cases with germline BRCA1/2 (gBRCA) wild-type. All sBRCA PVs were detected in non-familial cases. By analysing our regional germline BRCA1/2 database there were 92/1114 (8.3%) gBRCA PVs detected in non-familial cases (only 3% ≥70 years old) and 245/641 (38.2%) in familial cases. Germline non-familial cases were dominated by BRCA2 in older women (8/271 ≥ 70 years old, all BRCA2). The ratio of sBRCA-to-gBRCA was ≤1.0 in women aged

Peer reviewed: True, OBJECTIVE: To describe national patterns of National Health Service (NHS) analysis of mismatch repair (MMR) genes in England using individual-level data submitted to the National Disease Registration Service (NDRS) by the NHS regional molecular genetics laboratories. DESIGN: Laboratories submitted individual-level patient data to NDRS against a prescribed data model, including (1) patient identifiers, (2) test episode data, (3) per-gene results and (4) detected sequence variants. Individualised per-laboratory algorithms were designed and applied in NDRS to extract and map the data to the common data model. Laboratory-level MMR activity audit data from the Clinical Molecular Genetics Society/Association of Clinical Genomic Science were used to assess early years' missing data. RESULTS: Individual-level data from patients undergoing NHS MMR germline genetic testing were submitted from all 13 English laboratories performing MMR analyses, comprising in total 16 722 patients (9649 full-gene, 7073 targeted), with the earliest submission from 2000. The NDRS dataset is estimated to comprise >60% of NHS MMR analyses performed since inception of NHS MMR analysis, with complete national data for full-gene analyses for 2016 onwards. Out of 9649 full-gene tests, 2724 had an abnormal result, approximately 70% of which were (likely) pathogenic. Data linkage to the National Cancer Registry demonstrated colorectal cancer was the most frequent cancer type in which full-gene analysis was performed. CONCLUSION: The NDRS MMR dataset is a unique national pan-laboratory amalgamation of individual-level clinical and genomic patient data with pseudonymised identifiers enabling linkage to other national datasets. This growing resource will enable longitudinal research and can form the basis of a live national genomic disease registry.

OBJECTIVE: Rearranged ROS1, present in 1%-2% of non-small cell lung cancer (NSCLC) patients, usually young, never or light smokers, is assessed by fluorescence in situ hybridization (FISH) to determine eligibility for tyrosine kinase inhibitors (TKI). Immunohistochemistry (IHC) for the protein product of ROS1 rearrangement, a cost-effective alternative, is validated on cytology and small biopsy samples.METHODS: From 1 March to 31 December 2019, cytology cell blocks and small biopsy samples from a selected cohort of NSCLC patients were concurrently tested for ROS1 gene rearrangement by Vysis 6q22 Break Apart FISH probe and IHC using Cell Signalling D4D6 antibody. Mismatch cases were tested by an RNA fusion next generation sequencing (NGS) panel.RESULTS: In a prospective population of 95 cases, 91 were negative and two were positive by both FISH and IHC. Both dual positive cases were female never smokers and benefited from TKI treatment. Another two cases were positive by FISH but negative by IHC and repeat by NGS showed one to be negative but one failed. Turnaround time for IHC was 0 to 8 days from request to authorisation, whilst that of FISH was 9 to 42 days at a cost of £51 and £159 respectively.CONCLUSION: IHC to assess for the protein product of ROS1 gene rearrangement on cytology cell blocks and small biopsy samples in a routine setting is a promising screening method to assess eligibility for TKI treatment with positive and indeterminate cases confirmed by FISH or NGS as it has good negative predictive value, faster turnaround time and is cost effective, with proven technical and clinical validation.

Schwannomatosis comprises a group of hereditary tumor predisposition syndromes characterized by, usually benign, multiple nerve sheath tumors, which frequently cause severe pain that does not typically respond to drug treatments. The most common schwannomatosis-associated gene is NF2, but SMARCB1 and LZTR1 are also associated. There are still many cases in which no pathogenic variants (PVs) have been identified, suggesting the existence of as yet unidentified genetic risk factors. In this study, we performed extended genetic screening of 75 unrelated schwannomatosis patients without identified germline PVs in NF2, LZTR1, or SMARCB1. Screening of the coding region of DGCR8, COQ6, CDKN2A, and CDKN2B was carried out, based on previous reports that point to these genes as potential candidate genes for schwannomatosis. Deletions or duplications in CDKN2A, CDKN2B, and adjacent chromosome 9 region were assessed by multiplex ligation-dependent probe amplification analysis. Sequencing analysis of a patient with multiple schwannomas and melanomas identified a novel duplication in the coding region of CDKN2A, disrupting both p14ARF and p16INK4a. Our results suggest that none of these genes are major contributors to schwannomatosis risk but the possibility remains that they may have a role in more complex mechanisms for tumor predisposition.

Clinical guidelines recommend testing both germline and tumour DNA forAn observational study that included all tumourOne hundred and thirteen tumourWe predict the likelihood of a tumour

The British Gynecological Cancer Society and the British Association of Gynecological Pathologists established a multidisciplinary consensus group comprising experts in surgical gynecological oncology, medical oncology, genetics, and laboratory science, and clinical nurse specialists to identify the optimal pathways to BRCA germline and tumor testing in patients with ovarian cancer in routine clinical practice. In particular, the group explored models of consent, quality standards identified at pathology laboratories, and experience and data from pioneering cancer centers. The group liaised with representatives from ovarian cancer charities to also identify patient perspectives that would be important to implementation. Recommendations from these consensus group deliberations are presented in this manuscript.

Poly(ADP-ribose) polymerase (PARP) inhibitors improve survival in BRCA-mutant high-grade serous ovarian carcinoma. As a result, germline and somatic BRCA1/2 testing has become standard practice in women diagnosed with ovarian cancer. We outline changes in testing and detection rates of germline BRCA1/2 pathogenic variants (PVs) in cases of non-mucinous epithelial ovarian cancer diagnosed during three eras, spanning 12 years, within the North West of England, and compare the uptake of cascade testing in families identified by oncology-led mainstreaming versus regional genetics clinics. Eras included: Period 1 (20% risk threshold for testing): between January 2007 and May 2013; Period 2 (10% risk threshold for testing): between June 2013 and October 2017 and; Period 3 (mainstream testing): between November 2017 and November 2019. A total of 1081 women underwent germline BRCA1/2 testing between January 2007 and November 2019 and 222 (20.5%) were found to have a PV. The monthly testing rate increased by 3.3-fold and 2.5-fold between Periods 1–2 and Periods 2–3, respectively. A similar incidence of germline BRCA1/2 PVs were detected in Period 2 (17.2%) and Period 3 (18.5%). Uptake of cascade testing from first-degree relatives was significantly lower in those women undergoing mainstream testing compared with those tested in regional genetics clinics (31.6% versus 47.3%, P = 0.038). Mainstream testing allows timely detection of germline BRCA1/2 status to select patients for PARP inhibitors, but shortfalls in the uptake of cascade testing in first-degree relatives requires optimisation to broaden benefits within families.

BackgroundGenetic testing for BRCA1 and BRCA2 pathogenic variants (PVs) has been available in North West England since 1995. We assessed uptake of pre-symptomatic testing in 1564 families with PVs over a 24.5year follow-up (FU) period.MethodsFirst-degree relatives (FDRs) in families with BRCA1 or BRCA2 PVs were eligible from date of index family report if unaffected by a relevant cancer and alive at report date. FDRs were censored as not having undergone a pre-symptomatic test at diagnosis of a relevant cancer, date of death, age 93 or 30/03/2019. Time to uptake of pre-symptomatic testing was assessed by Kaplan–Meier curves, by gender and children.Results2554 male and 3115 female FDRs were eligible. Overall uptake was 775 (30.3%) in men and 1935 (62.1%) in women. This increased at 15 years to 33.6% and 67.9%, and continued to rise until 24 years (p80% by 15 years. Uptake was higher in parous women (pConclusionUptake of BRCA1/2 pre-symptomatic testing is age, gender and time-dependent, and higher in women with children and men with daughters.

Neurofibromatosis type 2 (NF2) is an autosomal dominant condition caused by pathogenic variants in the NF2 gene. The pathogenic variant is either inherited or obtained by de novo mutation, characterised by the presence of schwannomas, meningiomas and ependymomas. Here we report the presence of NF2 in one twin, with bilateral vestibular schwannomas and a pathogenic variant of the NF2 gene identified in both tumour and lymphocytes, while his monozygous brother remains asymptomatic. Imaging of the unaffected twin showed no tumour load and genetic testing via Sanger sequencing and Amplification Refractory Mutation System assay demonstrated low levels of expression of the NF2 variant in lymphocytes. Further testing on non-haemopoietic tissue showed little expression or absence of the pathogenic variant. Given there is no family history and the low level of the variant, we assume the pathogenic variant is a de novo mutation during embryogenesis. De novo mutations have been described as occurring at three possible time points in the creation of monozygous twins with different genetic make-up; prior to the twinning event, as a cause of the event, or after the twinning event. Of these options, we hypothesise that the discordance in the expression of the NF2 variant between these twins is likely due to a mutational event that occurred as a result of either of the latter two possibilities, between which we cannot determine. The pathogenic variant in lymphocytes was likely transferred between the twins through a shared blood supply in utero, and the non-haemopoietic samples that showed low levels of expression, were likely due to the presence of lymphocytic cells. Therefore, we have a discordance between monozygous twins at the NF2 gene.

PURPOSE: To evaluate the incidence of mosaicism in de novo neurofibromatosis 2 (NF2).METHODS: Patients fulfilling NF2 criteria, but with no known affected family member from a previous generation (n = 1055), were tested for NF2 variants in lymphocyte DNA and where available tumor DNA. The proportion of individuals with a proven or presumed mosaic NF2 variant was assessed and allele frequencies of identified variants evaluated using next-generation sequencing.RESULTS: The rate of proven/presumed mosaicism was 232/1055 (22.0%). However, nonmosaic heterozygous pathogenic variants were only identified in 387/1055 (36.7%). When variant detection rates in second generation nonmosaics were applied to de novo cases, we assessed the overall probable mosaicism rate to be 59.7%. This rate differed by age from 21.7% in those presenting with bilateral vestibular schwannoma CONCLUSION: This study has identified a very high probable mosaicism rate in de novo NF2, probably making NF2 the condition with the highest expressed rate of mosaicism in de novo dominant disease that is nonlethal in heterozygote form. Risks to offspring are small and probably correlate with variant allele frequency detected in blood.

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 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.

BackgroundBilateral vestibular schwannomas (VS) are pathognomonic of Neurofibromatosis type 2 (NF2), but the diagnostic criteria also include unilateral vestibular schwannomas (UVS) in combination with multiple meningiomas (MM) and other schwannomas, as well as MM without VS. ObjectiveTo investigate the diagnostic value of these criteria and establish the presence of other genetic conditions in patients presenting in this manner.Methods The (BLINDED FOR REVIEW) NF2 database was accessed to obtain information on patients presenting with a UVS and MM or ≥2 non-intradermal schwannomas (NIDS). We gathered data on patients diagnosed with NF2 due to MM without VS, and on patients presenting with MM without meeting NF2 criteria. Analysis was performed for pathogenic variants (PV) in NF2, SMARCE1, SMARCB1 and LZTR1.Results 131/131 patients presenting with a UVS and MM had a non-refuted diagnosis of NF2 after molecular studies, in comparison with 85/96 patients presenting with UVS and ≥2 NIDS (p=ConclusionPatients presenting with UVS and MM are significantly more likely to have a non-refuted diagnosis of NF2 than patients presenting with UVS and ≥2 NIDS, but significantly less likely to develop bilateral VS. 7% of those presenting with MM without meeting NF2 criteria had PV in SMARCE1, and 5% had mosaic NF2.

Purpose: We have evaluated deficiencies in existing diagnostic criteria for neurofibromatosis 2 (NF2). Methods: Two large databases of individuals fulfilling NF2 criteria (n=1361) and those tested for NF2 variants with criteria short of diagnosis (n=1416) were interrogated. We assessed the proportions meeting each diagnostic criterion with constitutional or mosaic NF2 variants and the Positive Predictive Value (PPV) with regard to definite diagnosis.Results: There was no evidence for usefulness of old criteria ‘glioma’ or ‘neurofibroma’. ‘Ependymoma’ had 100% PPV and high levels of confirmed NF2 diagnosis (67.7%). Those with bilateral vestibular schwannoma (VS) alone aged ≥60 years had the lowest confirmation rate (6.6%) and reduced PPV(80%). Siblings as a first-degree-relative, without an affected parent, had 0% PPV. All three individuals with unilateral VS and an affected sibling were proven not to have NF2. The biggest overlap was with LZTR1-associated schwannomatosis. In this category, seven individuals with unilateral VS plus ≥2 non-dermal schwannomas reduced PPV to 67%.Conclusion: The present study has confirmed important deficiencies in NF2 diagnostic criteria. The term ‘glioma’ should be dropped and replaced by ‘ependymoma’. Similarly ‘neurofibroma’ should be removed. Dropping ‘sibling’ from first-degree-relatives should be considered and testing of LZTR1 should be recommended for unilateral VS.

A unique cohort of chemo-naive gastrointestinal stromal tumors (GISTs) with double-primary tyrosine kinase mutations was characterized particularly to determine whether coexistent mutations represent a single mutational event. Up to 2013, 4 UK centers reported 9 GISTs with 2 primary tyrosine kinase mutations. In each of 8 cases validated by next generation sequencing, both mutations were present in the same allele of the same exon (KIT exon 11 or 17, or PDGFRA exon 18). One case showed the second mutation only on some of the mutant alleles. Seven cases showed both mutations in all the reads, but in 2 cases, additional variants were found only in some reads. Clinicopathologic features of the 8 cases were similar to GISTs with single-primary mutations. When GIST genotyping rarely uncovers multiple tyrosine kinase variants in an exon, they occur in the same allele but are likely to represent separate mutational events and lack clinical significance.

IntroductionPoly(ADP-ribose) polymerase inhibitors significantly improve progression-free survival in platinum-sensitive high-grade serous and endometrioid ovarian carcinoma, with greatest benefits observed in women with a pathogenic BRCA1/2 variant. Consequently, the demand for germline BRCA1/2 testing in ovarian cancer has increased substantially, leading to the screening of unselected populations of patients. We aimed to determine the prevalence of pathogenic germline BRCA1/2 variants in women diagnosed with epithelial ovarian cancer, categorised according to the established risk factors for hereditary breast and ovarian cancer syndrome and the Manchester BRCA Score, to inform risk stratification.MethodsA cohort of sequential epithelial ovarian cancer cases recruited between June 2013 and September 2018 underwent germline BRCA1/2 testing by next-generation sequencing and multiplex ligation-dependent probe amplification.ResultsFive hundred and fifty-seven patients were screened. Of these, 18% had inherited a pathogenic BRCA1/2 variant. The prevalence of pathogenic BRCA1/2 variants was >10% in women diagnosed with ovarian cancer earlier than 60 years of age (21%) and those diagnosed later than 60 years of age with a family history of breast and/or ovarian cancer (17%) or a medical history of breast cancer (34%). The prevalence of pathogenic BRCA1/2 variants was also >10% in women with a Manchester BRCA Score of ≥15 points (14%).DiscussionOur study suggests that age at diagnosis, family history of breast and/or ovarian cancer, medical history of breast cancer or a Manchester BRCA Score of ≥15 points are associated with a >10% prevalence of germline pathogenic BRCA1/2 variants in epithelial ovarian cancer.

Purpose Women at increased familial breast cancer risk have been offered screening starting at an earlier age and increased frequency than national Screening Programmes for over 30 years. There are limited data on longer-term largescale implementation of this approach on cancer diagnosis. Methods Women at our institution at ≥ 17% lifetime breast cancer risk have been offered enhanced screening with annual mammography starting at age 35 or 5-years younger than youngest affected relative, with upper age limit 50 for moderate and 60 for high-risk. Breast cancer pathology, stage and receptor status were assessed as well as survival from cancer diagnosis by Kaplan–Meier analysis. Results Overall 14,311 women were seen and assessed for breast cancer risk, with 649 breast cancers occurring in 129,119 years follow up (post-prevalent annual incidence = 4.55/1000). Of 323/394 invasive breast cancers occurring whilst on enhanced screening, most were lymph-node negative (72.9%), T1 (≤ 20 mm, 73.2%) and stage-1 (61.4%), 126/394 stage2–4 (32%). 10-year breast cancer specific survival was 91.3% (95% CI 87.4–94.0) better than the 75.9% (95% CI 74.9–77.0) published for England in 2013–2017. As expected, survival was significantly better for women with screen detected cancers (p n = 75; 95% CI 84.2–97.6). Women with lobular breast cancers had worse 10-year survival at 85.9% (95% CI 66.7–94.5). Breast cancer specific survival was good for 119 BRCA1/2 carriers with 20-year survival in BRCA1:91.2% (95% CI 77.8–96.6) and 83.8% (62.6–93.5) for BRCA2. Conclusions Targeted breast screening in women aged 30–60 years at increased familial risk is associated with good long-term survival that is substantially better than expected from population data.

Purpose: Lobular breast cancer (LBC) accounts for ~ 15% of breast cancer. Here, we studied the frequency of pathogenic germline variants (PGVs) in an extended panel of genes in women affected with LBC. Methods: 302 women with LBC and 1567 without breast cancer were tested for BRCA1/2 PGVs. A subset of 134 LBC affected women who tested negative for BRCA1/2 PGVs underwent extended screening, including: ATM, CDH1, CHEK2, NBN, PALB2, PTEN, RAD50, RAD51D, and TP53.Results: 35 PGVs were identified in the group with LBC, of which 22 were in BRCA1/2. Ten actionable PGVs were identified in additional genes (ATM(4), CDH1(1), CHEK2(1), PALB2(2) and TP53(2)). Overall, PGVs in three genes conferred a significant increased risk for LBC. Odds ratios (ORs) were: BRCA1: OR = 13.17 (95%CI 2.83–66.38; P = 0.0017), BRCA2: OR = 10.33 (95%CI 4.58–23.95; P ATM: OR = 8.01 (95%CI 2.52–29.92; P = 0.0053). We did not detect an increased risk of LBC for PALB2, CDH1 or CHEK2. Conclusion: The overall PGV detection rate was 11.59%, with similar rates of BRCA1/2 (7.28%) PGVs as for other actionable PGVs (7.46%), indicating a benefit for extended panel genetic testing in LBC. We also report a previously unrecognised association of pathogenic variants in ATM with LBC.

Faiza Gaba, Oleg Blyuss, Jatinderpal Kalsi, Saskia Sanderson, Andrew Wallace, Antonis C. Antoniou, Rosa Legood, Usha Menon, Ian Jacobs, & Ranjit Manchanda, on behalf of the PROMISE-FS team Introduction/Background The current approach to genetic-testing and risk assessment is based on family-history and misses the majority of people at risk. Unselected population-based testing can enable personalised ovarian cancer (OC) risk prediction combining genetic/epidemiology/hormonal data. This permits population risk stratification for risk adapted targeted screening and prevention. Such an intervention study has not previously been undertaken. We aimed to assess the feasibility of OC risk stratification of general population women using a personalised OC risk tool followed by risk management. Methodology Volunteers were recruited through London primary care networks. Inclusion criteria: women ≥18 years. Exclusion criteria: prior ovarian/tubal/peritoneal cancer, previous genetic testing for OC genes. Participants accessed an online/web-based decision aid along with optional telephone helpline use. Consenting individuals completed risk assessment and underwent genetic testing (BRCA1/BRCA2/RAD51C/RAD51D/BRIP1, OC susceptibility single-nucleotide polymorphisms). A validated OC risk prediction algorithm provided a personalised OC risk estimate using genetic/lifestyle/hormonal OC risk factors. Population genetic testing (PGT) for OC-risk stratification uptake/acceptability, satisfaction, decision aid/telephone helpline use, psychological health and quality of life were assessed using validated/customised questionnaires over six months. Linear-mixed models/contrast tests analysed impact on study outcomes. Main outcomes: feasibility/acceptability, uptake, decision aid/telephone helpline use, satisfaction/regret, and impact on psychological health/quality of life. Results In total, 123 volunteers (mean age= 48.5 (SD=15.4) years) used the decision aid, 105 (85%) consented. None fulfilled NHS genetic-testing clinical criteria. OC-risk stratification revealed 1/103 at ≥10% (high), 0/103 at ≥5%– Conclusion Findings suggest population-based personalised OC risk stratification is feasible and acceptable, has high satisfaction, reduces cancer worry/risk perception, and does not negatively impact psychological health or quality-of-life. Larger implementation studies evaluating long-term impact and cost effectiveness of this strategy are needed. Disclosures RM- funding from CRUK & Eve Appeal for thiswork. Funding from Barts Charity, Rosetrees trust outside this work. Honorarium from Astrazeneca & MSD. IJ, UM- Financial interest in Abcodia, company for development of biomarkers for early detection of cancer. Other authors- No disclosures.

OBJECTIVES: Despite wide excision and post-operative irradiation, loco-regional and/or metastatic recurrence is a significant clinical problem in salivary adenoid cystic carcinoma (SACC). Reliable biomarkers are required to tailor post-treatment surveillance to patients at highest risk of recurrence. We sought to determine the utility of TP53 and PIK3CA mutations as prognostic biomarkers in SACC.MATERIALS AND METHODS: DNA was extracted from archival tumour blocks of 145 SACC patients from 66 UK referral centres and sequenced for TP53 and PIK3CA mutations. Clinical, pathological and outcome data were analysed to determine the impact of the genomic alterations on disease recurrence and overall survival (OS).RESULTS: TP53 and PIK3CA mutations were identified in 8% (10/121 successful analyses) and 2% (3/121) of cases, respectively. There were too few PIK3CA mutations in this cohort for informative further analysis. TP53-mutated SACC had significantly shorter median OS (5.3 vs. 16.3 years, p = 0.019) and lower 10-year survival (48% vs. 81%) compared with TP53 wild-type ACC. Solid-pattern histopathology was more frequent in TP53-mutated SACC (50% vs. 15%, p = 0.27).CONCLUSION: TP53-mutated recurrent and metastatic SACC was associated with shorter OS, which was significant when combined with published genomic data sets. Stratifying by TP53 status, in addition to established clinical, pathological and genomic biomarkers, may usefully inform follow-up strategy.

Where previously, germline genetic testing in deceased affected relatives was not possible due to the absence of lymphocytic DNA, the North-West-Genomic-Laboratory Hub (NWGLH) has developed and validated next-generation sequencing based gene panels utilising formalin-fixed-paraffin-embedded (FFPE) tissue DNA from deceased individuals. This technology has been utilised in the clinical setting for the management of unaffected relatives seen in the Clinical Genetics Service (CGS). Here we assess the clinical impact. At the time of data collection, the NWGLH had analysed 180 FFPE tissue samples from deceased affected individuals: 134 from breast and/or ovarian cancer cases for germline variants in the BRCA1/BRCA2 genes and 46 from colorectal, gastric, ovarian and endometrial cancer cases for germline variants in a panel of 13 genes implicated in inherited colorectal cancer and gastric cancer conditions. Successful analysis was achieved in 140/180 cases (78%). In total, 29 germline pathogenic/likely pathogenic variants were identified in autosomal dominant cancer predisposition genes where the gene was pertinent to the cancer family history (including BRCA1/BRCA2, the mismatch-repair genes and APC). Of the 180 cases, the impact of the result on clinical management of unaffected relatives was known in 143 cases. Of these, the results in 54 cases (38%) directly impacted the clinical management of relatives seen by the CGS. This included changes to risk assessments, screening recommendations and the availability of predictive genetic testing to unaffected relatives. Our data demonstrate how FFPE testing in deceased relatives is an accurate and informative tool in the clinical management of patients referred to the CGS.

BackgroundHereditary causes of ovarian cancer include Lynch syndrome, which is due to inherited pathogenic variants affecting one of the four mismatch repair genes involved in DNA repair. The aim of this study was to evaluate tumour mismatch repair deficiency and prevalence of Lynch syndrome in high-risk women referred to the Manchester Centre for Genomic Medicine with ovarian cancer over the past 20 years.MethodsWomen with ovarian cancer diagnosed before the age of 35 years and/or with a suggestive personal or family history of Lynch syndrome cancers underwent tumour testing with immunohistochemistry for mismatch repair deficiency and, where indicated, MLH1 promoter methylation testing followed by constitutional testing for Lynch syndrome.ResultsIn total, 261 ovarian cancers were tested and 27 (10.3%; 95% CI 6.9% to 14.7%) showed mismatch repair deficiency by immunohistochemistry. Three of 7 with MLH1 loss showed MLH1 promoter hypermethylation, and 18 of the remaining 24 underwent constitutional testing for Lynch syndrome. A further 15 women with mismatch repair proficient tumours underwent constitutional testing because of a strong family history of Lynch syndrome cancers. Pathogenic variants were identified in 9/33 (27%) women who underwent constitutional testing, aged 33–59 years (median 48 years), including one whose tumour was mismatch repair proficient. Most Lynch syndrome tumours were of endometrioid histological subtype.ConclusionsTumour mismatch repair deficiency identified by immunohistochemistry is a useful prescreen for constitutional testing in women with ovarian cancer with personal or family histories suggestive of Lynch syndrome.

Accurate classification of variants in cancer susceptibility genes (CSGs) is key for correct estimation of cancer risk and management of patients. Consistency in the weighting assigned to individual elements of evidence has been much improved by the American College of Medical Genetics (ACMG) 2015 framework for variant classification, UK Association for Clinical Genomic Science (UK-ACGS) Best Practice Guidelines and subsequent Cancer Variant Interpretation Group UK (CanVIG-UK) consensus specification for CSGs. However, considerable inconsistency persists regarding practice in the combination of evidence elements. CanVIG-UK is a national subspecialist multidisciplinary network for cancer susceptibility genomic variant interpretation, comprising clinical scientist and clinical geneticist representation from each of the 25 diagnostic laboratories/clinical genetic units across the UK and Republic of Ireland. Here, we summarise the aggregated evidence elements and combinations possible within different variant classification schemata currently employed for CSGs (ACMG, UK-ACGS, CanVIG-UK and ClinGen gene-specific guidance for PTEN, TP53 and CDH1). We present consensus recommendations from CanVIG-UK regarding (1) consistent scoring for combinations of evidence elements using a validated numerical ‘exponent score’ (2) new combinations of evidence elements constituting likely pathogenic’ and ‘pathogenic’ classification categories, (3) which evidence elements can and cannot be used in combination for specific variant types and (4) classification of variants for which there are evidence elements for both pathogenicity and benignity.

Unselected population-based personalised ovarian cancer (OC) risk assessment combining genetic/epidemiology/hormonal data has not previously been undertaken. We aimed to perform a feasibility study of OC risk stratification of general population women using a personalised OC risk tool followed by risk management. Volunteers were recruited through London primary care networks. Inclusion criteria: women &ge, 18 years. Exclusion criteria: prior ovarian/tubal/peritoneal cancer, previous genetic testing for OC genes. Participants accessed an online/web-based decision aid along with optional telephone helpline use. Consenting individuals completed risk assessment and underwent genetic testing (BRCA1/BRCA2/RAD51C/RAD51D/BRIP1, OC susceptibility single-nucleotide polymorphisms). A validated OC risk prediction algorithm provided a personalised OC risk estimate using genetic/lifestyle/hormonal OC risk factors. Population genetic testing (PGT)/OC risk stratification uptake/acceptability, satisfaction, decision aid/telephone helpline use, psychological health and quality of life were assessed using validated/customised questionnaires over six months. Linear-mixed models/contrast tests analysed impact on study outcomes. Main outcomes: feasibility/acceptability, uptake, decision aid/telephone helpline use, satisfaction/regret, and impact on psychological health/quality of life. In total, 123 volunteers (mean age = 48.5 (SD = 15.4) years) used the decision aid, 105 (85%) consented. None fulfilled NHS genetic testing clinical criteria. OC risk stratification revealed 1/103 at &ge, 10% (high), 0/103 at &ge, 5%&ndash, &lt, 10% (intermediate), and 100/103 at &lt, 5% (low) lifetime OC risk. Decision aid satisfaction was 92.2%. The telephone helpline use rate was 13% and the questionnaire response rate at six months was 75%. Contrast tests indicated that overall depression (p = 0.30), anxiety (p = 0.10), quality-of-life (p = 0.99), and distress (p = 0.25) levels did not jointly change, while OC worry (p = 0.021) and general cancer risk perception (p = 0.015) decreased over six months. In total, 85.5&ndash, 98.7% were satisfied with their decision. Findings suggest population-based personalised OC risk stratification is feasible and acceptable, has high satisfaction, reduces cancer worry/risk perception, and does not negatively impact psychological health/quality of life.

ObjectivesCases of sporadic vestibular schwannoma (sVS) have a low rate of association with germline pathogenic variants. However, some individuals with sVS can represent undetected cases of neurofibromatosis type 2 (NF2) or schwannomatosis. Earlier identification of patients with these syndromes can facilitate more accurate familial risk prediction and prognosis.MethodsCases of sVS were ascertained from a local register at the Manchester Centre for Genomic Medicine. Genetic analysis was conducted in NF2 on blood samples for all patients, and tumour DNA samples when available. LZTR1 and SMARCB1 screening was also performed in patient subgroups.ResultsAge at genetic testing for vestibular schwannoma (VS) presentation was younger in comparison with previous literature, a bias resulting from updated genetic testing recommendations. Mosaic or constitutional germline NF2 variants were confirmed in 2% of patients. Pathogenic germline variants in LZTR1 were found in 3% of all tested patients, with a higher rate of 5% in patients SMARCB1 variants were identified within the cohort. Considering all individuals who received tumour DNA analysis, 69% of patients were found to possess two somatic pathogenic NF2 variants, including those with germline LZTR1 pathogenic variants.ConclusionsUndiagnosed schwannoma predisposition may account for a significant minority of apparently sVS cases, especially at lower presentation ages. Loss of NF2 function is a common event in VS tumours and may represent a targetable common pathway in VS tumourigenesis. These data also support the multi-hit mechanism of LZTR1-associated VS tumourigenesis.

ObjectivesSchwannomatosis is a dominantly inherited condition predisposing to schwannomas of mainly spinal and peripheral nerves with some diagnostic overlap with neurofibromatosis-2 (NF2), but the underlying epidemiology is poorly understood. We present the birth incidence and prevalence allowing for overlap with NF2.MethodsSchwannomatosis and NF2 cases were ascertained from the Manchester region of England (population=4.8 million) and from across the UK. Point prevalence and birth incidence were calculated from regional birth statistics. Genetic analysis was also performed on NF2, LZTR1 and SMARCB1 on blood and tumour DNA samples when available.ResultsRegional prevalence for schwannomatosis and NF2 were 1 in 126 315 and 50 500, respectively, with calculated birth incidences of 1 in 68 956 and 1 in 27 956. Mosaic NF2 causes a substantial overlap with schwannomatosis resulting in the misdiagnosis of at least 9% of schwannomatosis cases. LZTR1-associated schwannomatosis also causes a small number of cases that are misdiagnosed with NF2 (1%–2%), due to the occurrence of a unilateral vestibular schwannoma. Patients with schwannomatosis had lower numbers of non-vestibular cranial schwannomas, but more peripheral and spinal nerve schwannomas with pain as a predominant presenting symptom. Life expectancy was significantly better in schwannomatosis (mean age at death 76.9) compared with NF2 (mean age at death 66.2; p=0.004).ConclusionsWithin the highly ascertained North-West England population, schwannomatosis has less than half the birth incidence and prevalence of NF2.

In families with multiple affected individuals with early-onset breast/ovarian cancer pathogenic variants in BRCA1 or BRCA2 are identified in approximately 20% of the cases. Extensive efforts have been made to identify additional highly penetrant breast cancer genes or alternative mutational mechanisms affecting BRCA1 and BRCA2 to explain the missing heritability. It has been proposed that part of the missing heritability may be explained by gene silencing due to promoter methylation of cancer associated genes, as described in colorectal cancer (MLH1 and MSH2). Here, for the first time, we report two independent families with multiple individuals affected by breast and ovarian cancer with transgenerational promoter methylation of BRCA1. RNA analysis of BRCA1 in the germline of breast/ovarian families, previously found to be BRCA1/2 mutation negative by Sanger sequencing and copy number analysis, identified two families with allelic loss of expression. To investigate the mechanism of transcriptional silencing, a total of 14 affected and unaffected family members from these two families were tested for BRCA1 promoter methylation by pyrosequencing in blood, buccal, and hair follicle cells. Allele specific methylation was determined by clonal bisulphite sequencing. BRCA1 promoter methylation in all three germ layers was present in 11 of 14 family members. Of the 7 women with promoter methylation five were affected with grade 3 breast/high grade serous ovarian cancer. The four males with BRCA1 promoter methylation had no history of cancer. Methylation levels were ˜50%, consistent with the silencing of one allele detected in RNA in these family members. Clonal bisulphite sequencing of an affected family member of each family confirmed that the alternative allele was specifically methylated. Interestingly, in both families the methylation pattern of the BRCA1 promoter segregated with the same novel heterozygous variant in the 5'UTR of BRCA1. These results indicate a novel mechanism for familial breast/ovarian cancer, caused by epigenetic silencing of one allele by transgenerational hypermethylation of the BRCA1 promoter, secondary to a variant in cis of BRCA1. We propose that methylation analyses are indicated in all families affected by early onset breast/ovarian cancer where standard mutation screening of BRCA1/2 has not identified a causative variant. Citation Format: van Veen EM, Smith MJ, Byers HJ, Wallace AJ, Lalloo FI, Newman WG, Evans DGR. Transgenerational epigenetic silencing of BRCA1 due to a germline variant unmasks a new mechanism for familial breast and ovarian cancer [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr PD1-05.

Ovarian cancer patients with germline or somatic pathogenic variants benefit from treatment with poly ADP ribose polymerase (PARP) inhibitors. Tumor BRCA1/2 testing is more challenging than germline testing as the majority of samples are formalin-fixed paraffin embedded (FFPE), the tumor genome is complex, and the allelic fraction of somatic variants can be low. We collaborated with 10 laboratories testing BRCA1/2 in tumors to compare different approaches to identify clinically important variants within FFPE tumor DNA samples. This was not a proficiency study but an inter-laboratory comparison to identify common issues. Each laboratory received the same tumor DNA samples ranging in genotype, quantity, quality, and variant allele frequency (VAF). Each laboratory performed their preferred next-generation sequencing method to report on the variants. No false positive results were reported in this small study and the majority of methods detected the low VAF variants. A number of variants were not detected due to the bioinformatics analysis, variant classification, or insufficient DNA. The use of hybridization capture or short amplicon methods are recommended based on a bioinformatic assessment of the data. The study highlights the importance of establishing standards and standardization for tBRCA testing particularly when the test results dictate clinical decisions regarding life extending therapies.

We present findings of a cancer multidisciplinary-team (MDT) coordinated mainstreaming pathway of unselected 5-panel germline BRCA1/BRCA2/RAD51C/RAD51D/BRIP1 and parallel somatic BRCA1/BRCA2 testing in all women with epithelial-OC and highlight the discordance between germline and somatic testing strategies across two cancer centres. Patients were counselled and consented by a cancer MDT member. The uptake of parallel multi-gene germline and somatic testing was 97.7%. Counselling by clinical-nurse-specialist more frequently needed &gt, 1 consultation (53.6% (30/56)) compared to a medical (15.0% (21/137)) or surgical oncologist (15.3% (17/110)) (p &lt, 0.001). The median age was 54 (IQR = 51–62) years in germline pathogenic-variant (PV) versus 61 (IQR = 51–71) in BRCA wild-type (p = 0.001). There was no significant difference in distribution of PVs by ethnicity, stage, surgery timing or resection status. A total of 15.5% germline and 7.8% somatic BRCA1/BRCA2 PVs were identified. A total of 2.3% patients had RAD51C/RAD51D/BRIP1 PVs. A total of 11% germline PVs were large-genomic-rearrangements and missed by somatic testing. A total of 20% germline PVs are missed by somatic first BRCA-testing approach and 55.6% germline PVs missed by family history ascertainment. The somatic testing failure rate is higher (23%) for patients undergoing diagnostic biopsies. Our findings favour a prospective parallel somatic and germline panel testing approach as a clinically efficient strategy to maximise variant identification. UK Genomics test-directory criteria should be expanded to include a panel of OC genes.

Background: Whilst the requirement for thresholds for testing for mutations in BRCA1/2 is being questioned, they are likely to remain for individuals unaffected by a relevant cancer. It is still useful to provide pre-testing likelihoods, but models need to take into account tumour pathology.Methods: The Manchester scoring system (MSS) is a well utilised, simple, paper based model for assessing carrier probability that already incorporates pathology data. We have used mutation testing data from 4115 unrelated samples from affected non-Jewish individuals alongside tumour pathology to further refine the scoring system.Results: Adding additional points for high grade serous ovarian cancer

Multireference electronic structure methods are used to assign ground state electronic configurations for a series of metallophthalocyanines. Ligand orbital occupancies remain constant across the period and are consistent with a formal 2– charge on the ligand. The d electron configurations of some metallophthalocyanines are straightforward and can be unambiguously assigned, (dxy)2(dxz,dyz)2,2( [Formula: see text])2([Formula: see text])n, with n = 2, 1, 0, respectively, for ZnPc, CuPc, and NiPc. Controversies over ground state electronic structure assignments for other metallophthalocyanines arise due to multiple complicating factors: accidental near-degeneracies, environmental effects, and different ligand field models used in interpreting experimental spectra. We demonstrate that explicit ligand field models provide more reliable and consistent interpretations of experimental data than implicit, parameterized alternatives. On this basis, we assign gas-phase electronic ground states for MnPc, (dxy)2(dxz,dyz)1,1([Formula: see text])1 and CoPc, (dxy)2(dxz,dyz)2,2([Formula: see text])1, and show that the ground state of FePc cannot be resolved to a single state, with two near-degenerate states that are likely spin-orbit coupled: (dxy)2(dxz,dyz)1,1( [Formula: see text])2 and (dxy)2(dxz,dyz)2,1([Formula: see text])1. Remaining differences between computational predictions and experimental observations are small and may be ascribed primarily to environmental effects but are also partly due to incomplete modelling of electron correlation.

Genetic testing forFirst-degree relatives (FDRs) in families with2554 male and 3115 female FDRs were eligible. Overall uptake was 775 (30.3%) in men and 1935 (62.1%) in women. This increased at 15 years to 33.6% and 67.9%, and continued to rise until 24 years (p0.001). For women, the 29-year to 39-year age group had the highest uptake at 10 years FU (72.5%; p0.01), whereas the 50-year to 59-year age group was highest in men (37.2%; p0.01). Women18 years at the time of familial variant identification had lower initial uptake, but this rose to80% by 15 years. Uptake was higher in parous women (p0.001) and in men with daughters (p0.0001).Uptake of

To evaluate the incidence of mosaicism in de novo neurofibromatosis 2 (NF2).Patients fulfilling NF2 criteria, but with no known affected family member from a previous generation (n = 1055), were tested for NF2 variants in lymphocyte DNA and where available tumor DNA. The proportion of individuals with a proven or presumed mosaic NF2 variant was assessed and allele frequencies of identified variants evaluated using next-generation sequencing.The rate of proven/presumed mosaicism was 232/1055 (22.0%). However, nonmosaic heterozygous pathogenic variants were only identified in 387/1055 (36.7%). When variant detection rates in second generation nonmosaics were applied to de novo cases, we assessed the overall probable mosaicism rate to be 59.7%. This rate differed by age from 21.7% in those presenting with bilateral vestibular schwannoma20 years to 80.7% in those aged ≥60 years. A mosaic variant was detected in all parents of affected children with a single-nucleotide pathogenic NF2 variant.This study has identified a very high probable mosaicism rate in de novo NF2, probably making NF2 the condition with the highest expressed rate of mosaicism in de novo dominant disease that is nonlethal in heterozygote form. Risks to offspring are small and probably correlate with variant allele frequency detected in blood.

Poly(ADP-ribose) polymerase inhibitors significantly improve progression-free survival in platinum-sensitive high-grade serous and endometrioid ovarian carcinoma, with greatest benefits observed in women with a pathogenicA cohort of sequential epithelial ovarian cancer cases recruited between June 2013 and September 2018 underwent germlineFive hundred and fifty-seven patients were screened. Of these, 18% had inherited a pathogenicOur study suggests that age at diagnosis, family history of breast and/or ovarian cancer, medical history of breast cancer or a Manchester BRCA Score of ≥15 points are associated with a10% prevalence of germline pathogenic

Heterozygous whole gene deletions (WGDs), and intragenic microdeletions, account for a significant proportion of mutations underlying cancer predisposition syndromes. We analyzed the frequency and genotype-phenotype correlations of microdeletions in 12 genes (BRCA1, BRCA2, TP53, MSH2, MLH1, MSH6, PMS2, NF1, NF2, APC, PTCH1, and VHL) representing seven tumor predisposition syndromes in 5,897 individuals (2,611 families) from our center. Overall, microdeletions accounted for 14% of identified mutations. As expected, smaller deletions or duplications were more common (12%) than WGDs (2.2%). Where a WGD was identified in the germline in NF2, the mechanism of somatic second hit was not deletion, as previously described for NF1. For neurofibromatosis type 1 and 2, we compared the mechanism of germline deletion. Unlike NF1, where three specific deletion sizes account for most germline WGDs, NF2 deletion breakpoints were different across seven samples tested. One of these deletions was 3.93 Mb and conferred a severe phenotype, thus refining the region for a potential NF2 modifier gene to a 2.04-Mb region on chromosome 22. The milder phenotype of NF2 WGDs may be due to the apparent absence of chromosome 22 loss as the second hit. These observations of WGD phenotypes will be helpful for interpreting incidental findings from microarray analysis and next-generation sequencing.

The crystal structures of tris(ethylmethylamino)-cyclopropenium chloride and tris(diethylamino)-cyclopropenium iodide reveal the presence of closely bound dicationic dimers formed from two closed-shell monomer units. The distances between the C3 centroids of the staggered monomers are at the short end of those normally found in π-stacked neutral arenes, let alone charged aromatic rings. Computational analysis reveals that short-range interactions are dominated by strong dispersion forces, enabling metastable dicationic dimers to form without covalent intermolecular bonding. Surrounding counterions then provide a background source of charge balance, imparting strong thermodynamic stability to the system. Additionally, these counterions form a weak but attractive electrostatic bridge between the monomer units, contributing to the surprisingly short observed intermolecular C3-C3 centroid distance.

Background Lynch syndrome (LS) predisposes to endometrial cancer (EC), colorectal cancer, and other cancers through inherited pathogenic variants affecting mismatch-repair (MMR) genes. Diagnosing LS in women with EC can reduce subsequent cancer mortality through colonoscopic surveillance and aspirin chemoprevention; it also enables cascade testing of relatives. A growing consensus supports LS screening in EC; however, the expected proportion of test positives, and optimal testing strategy is uncertain. Previous studies from insurance-based healthcare systems were limited by narrow selection criteria, failure to apply reference standard tests consistently, and poor conversion to definitive testing. The aim of this study was to establish the prevalence of LS and the diagnostic accuracy of LS testing strategies in an unselected EC population. Methods and findings This was a prospective cross-sectional study carried out at a large United Kingdom gynaecological cancer centre between October 2015 and January 2017. Women diagnosed with EC or atypical hyperplasia (AH) were offered LS testing. Tumours underwent MMR immunohistochemistry (IHC), microsatellite instability (MSI), and targeted MLH1-methylation testing. Women, Emma Crosbie and colleagues investigate the prevalence of and strategies for identifying Lynch syndrome among women with endometrial cancer., Author summary Why was this study done? Endometrial (womb) cancer (EC) is the most common gynaecological cancer in the developed world, and its incidence is rising. A significant minority (around 3%) of EC are caused by an inherited genetic predisposition called Lynch syndrome (LS). EC may be the first sign that a woman has LS. She is likely to survive this cancer but develop other preventable cancers related to LS later in life. Her family members are also at risk. Identifying women with LS can enable them to reduce their risk of new cancers, for example, through bowel (colorectal) cancer surveillance (colonoscopy). We do not know how many women with EC have LS or how best to identify them; current practice is based on experience in bowel cancer and may not be accurate in EC. What did this research do and find? We tested 500 women with EC treated in a large tertiary referral centre in the North West of England for LS. We did not preselect women to test based on clinical or tumour characteristics. We tested tumours for features of LS called mismatch repair (MMR) deficiency and microsatellite instability (MSI). Women with strongly suggestive clinical or tumour characteristics underwent germline LS testing. In total, 16 of 500 women (3%) had LS, and these women could not always be predicted by their age or family history. MMR deficiency was more accurate than MSI at identifying LS-EC, picking up 16/16 (100%) versus 9/16 (56%). What do these findings mean? In our study, we found that 3% of women with endometrial cancer have LS and can benefit from strategies to reduce their future cancer risk. Our results suggest that it may be best to test everyone because preselecting women to test based on clinical or tumour characteristics misses cases of LS. In this population, tumour MMR deficiency was more accurate than MSI at identifying LS in EC. Our results should be interpreted with caution because we did not do germline testing on all women, and the number of women we tested was relatively small.