Evidence-based recommendations for gene-specific ACMG/AMP variant classification from the ClinGen ENIGMA BRCA1 and BRCA2 Variant Curation Expert Panel.

The ENIGMA research consortium develops and applies methods to determine clinical significance of variants in hereditary breast and ovarian cancer genes. An ENIGMA BRCA1/2 classification sub-group, formed in 2015 as a ClinGen external expert panel, evolved into a ClinGen internal Variant Curation Expert Panel (VCEP) to align with Food and Drug Administration recognized processes for ClinVar contributions. The VCEP reviewed American College of Medical Genetics and Genomics/Association of Molecular Pathology (ACMG/AMP) classification criteria for relevance to interpreting BRCA1 and BRCA2 variants. Statistical methods were used to calibrate evidence strength for different data types. Pilot specifications were tested on 40 variants and documentation revised for clarity and ease of use. The original criterion descriptions for 13 evidence codes were considered non-applicable or overlapping with other criteria. Scenario of use was extended or re-purposed for eight codes. Extensive analysis and/or data review informed specification descriptions and weights for all codes. Specifications were applied to pilot variants with pre-existing ClinVar classification as follows: 13 uncertain significance or conflicting, 14 pathogenic and/or likely pathogenic, and 13 benign and/or likely benign. Review resolved classification for 11/13 uncertain significance or conflicting variants and retained or improved confidence in classification for the remaining variants. Alignment of pre-existing ENIGMA research classification processes with ACMG/AMP classification guidelines highlighted several gaps in the research processes and the baseline ACMG/AMP criteria. Calibration of evidence strength was key to justify utility and strength of different data types for gene-specific application. The gene-specific criteria demonstrated value for improving ACMG/AMP-aligned classification of BRCA1 and BRCA2 variants.

Challenges and approaches to calibrating patient phenotype as evidence for cancer gene variant classification under ACMG/AMP guidelines.

Since first publication of the American College of Medical Genetics and Genomics/Association for Medical Pathology (ACMG/AMP) variant classification guidelines, additional recommendations for application of certain criteria have been released (https://clinicalgenome.org/docs/), to improve their application in the diagnostic setting. However, none have addressed use of the PS4 and PP4 criteria, capturing patient presentation as evidence towards pathogenicity. Application of PS4 can be done through traditional case-control studies, or "proband counting" within or across clinical testing cohorts. Review of the existing PS4 and PP4 specifications for Hereditary Cancer Gene Variant Curation Expert Panels revealed substantial differences in the approach to defining specifications. Using BRCA1, BRCA2 and TP53 as exemplar genes, we calibrated different methods proposed for applying the "PS4 proband counting" criterion. For each approach, we considered limitations, non-independence with other ACMG/AMP criteria, broader applicability, and variability in results for different datasets. Our findings highlight inherent overlap of proband-counting methods with ACMG/AMP frequency codes, and the importance of calibration to derive dataset-specific code weights that can account for potential between-dataset differences in ascertainment and other factors. Our work emphasizes the advantages and generalizability of logistic regression analysis over simple proband-counting approaches to empirically determine the relative predictive capacity and weight of various personal clinical features in the context of multigene panel testing, for improved variant interpretation. We also provide a general protocol, including instructions for data formatting and a web-server for analysis of personal history parameters, to facilitate dataset-specific calibration analyses required to use such data for germline variant classification.

Rare Genetic Variants in LDLR, APOB, and PCSK9 are Associated with Aortic Stenosis.

Background: Despite a proposed causal role for low-density lipoprotein cholesterol (LDL-C) in aortic stenosis (AS), randomized controlled trials of lipid-lowering therapy failed to prevent severe AS. We aimed to assess the impact on AS and peak velocity across the aortic valve conferred by lifelong alterations in LDL-C levels mediated by protein-disrupting variants in three clinically significant genes for LDL metabolism (LDLR, APOB, PCSK9). Methods: We utilized sequencing data and electronic health records from UK Biobank (UKB) and All of Us and magnetic resonance imaging data from UKB. We identified predicted protein-disrupting variants with the LOFTEE and AlphaMissense algorithms and evaluated their associations with LDL-C and peak velocity across the aortic valve (UK Biobank), as well as diagnosed AS and aortic valve replacement (UK Biobank + All of Us). Results: We included 421,049 unrelated participants (5,621 with AS) in UKB and 195,519 unrelated participants (1,087 with AS) in All of Us. Carriers of protein-disrupting variants in LDLR had higher mean LDL-C (UKB: +42.6 mg/dl, P=4.4e-237) and greater risk of AS (meta-analysis: odds ratio [OR] =3.52 [95% CI 2.39-5.20], P=2.3e-10) and aortic valve replacement (meta-analysis: OR=3.78 [95% CI 2.26-6.32], P=4.0e-7). Carriers of protein-disrupting variants in APOB or PCSK9 had lower mean LDL-C (UKB: -32.3 mg/dl, P<5e-324) and lower risk of AS (meta-analysis: OR=0.49 [0.31-0.75], P=0.001) and aortic valve replacement (meta-analysis: OR=0.54 [0.30-0.97], P=0.04). Among 57,371 UKB imaging substudy participants, peak velocities across the aortic valve were greater in carriers of protein-disrupting variants in LDLR (+12.2cm/s, P=1.6e-5) and lower in carriers of protein-disrupting variants in PCSK9 (-6.9cm/s, P=0.022). Conclusions: Rare genetic variants that confer lifelong higher or lower LDL-C levels are associated with substantially increased and decreased risk of AS, respectively. Early and sustained lipid-lowering therapy may slow or prevent AS development.

D-mannose as a new therapy for fucokinase deficiency-related congenital disorder of glycosylation (FCSK-CDG).

INTRODUCTION: Fucokinase deficiency-related congenital disorder of glycosylation (FCSK-CDG) is a rare autosomal recessive inborn error of metabolism characterized by a decreased flux through the salvage pathway of GDP-fucose biosynthesis due to a block in the recycling of L-fucose that exits the lysosome. FCSK-CDG has been described in 5 individuals to date in the medical literature, with a phenotype comprising global developmental delays/intellectual disability, hypotonia, abnormal myelination, posterior ocular disease, growth and feeding failure, immune deficiency, and chronic diarrhea, without clear therapeutic recommendations. PATIENT AND METHODS: In a so far unreported FCSK-CDG patient, we studied proteomics and glycoproteomics in vitro in patient-derived fibroblasts and also performed in vivo glycomics, before and after treatment with either D-Mannose or L-Fucose. RESULTS: We observed a marked increase in fucosylation after D-mannose supplementation in fibroblasts compared to treatment with L-Fucose. The patient was then treated with D-mannose at 850 mg/kg/d, with resolution of the chronic diarrhea, resolution of oral aversion, improved weight gain, and observed developmental gains. Serum N-glycan profiles showed an improvement in the abundance of fucosylated glycans after treatment. No treatment-attributed adverse effects were observed. CONCLUSION: D-mannose is a promising new treatment for FCSK-CDG.

A stomate by any other name? The open question of hornwort gametophytic pores, their homology, and implications for the evolution of stomates.

Advances in bryophyte genomics and the phylogenetic recovery of hornworts, mosses, and liverworts as a clade have spurred considerable recent interest in character evolution among early embryophytes. Discussion of stomatal evolution, however, has been incomplete; the result of the neglect of certain potential stomate homologues, namely the two-celled epidermal gametophytic pores of hornworts (typically referred to as 'mucilage clefts'). Confusion over the potential homology of these structures is the consequence of a relatively recent consensus that hornwort gametophytic pores ('HGPs' - our term) are not homologous to stomates. We explore the occurrence and diverse functions of stomates throughout the evolutionary history and diversity of extinct and extant embryophytes. We then address arguments for and against homology between known sporophyte- and gametophyte-borne stomates and HGPs and conclude that there is little to no evidence that contradicts the hypothesis of homology. We propose that 'intergenerational heterotopy' might well account for the novel expression of stomates in gametophytes of hornworts, if stomates first evolved in the sporophyte generation of embryophytes. We then explore phylogenetically based hypotheses for the evolution of stomates in both the gametophyte and sporophyte generations of early lineages of embryophytes.

Microbially induced calcium carbonate precipitation through CO2 sequestration via an engineered Bacillus subtilis.

BACKGROUND: Microbially induced calcium carbonate precipitation has been extensively researched for geoengineering applications as well as diverse uses within the built environment. Bacteria play a crucial role in producing calcium carbonate minerals, via enzymes including carbonic anhydrase-an enzyme with the capability to hydrolyse CO2, commonly employed in carbon capture systems. This study describes previously uncharacterised carbonic anhydrase enzyme sequences capable of sequestering CO2 and subsequentially generating CaCO3 biominerals and suggests a route to produce carbon negative cementitious materials for the construction industry. RESULTS: Here, Bacillus subtilis was engineered to recombinantly express previously uncharacterised carbonic anhydrase enzymes from Bacillus megaterium and used as a whole cell catalyst allowing this novel bacterium to sequester CO2 and convert it to calcium carbonate. A significant decrease in CO2 was observed from 3800 PPM to 820 PPM upon induction of carbonic anhydrase and minerals recovered from these experiments were identified as calcite and vaterite using X-ray diffraction. Further experiments mixed the use of this enzyme (as a cell free extract) with Sporosarcina pasteurii to increase mineral production whilst maintaining a comparable level of CO2 sequestration. CONCLUSION: Recombinantly produced carbonic anhydrase successfully sequestered CO2 and converted it into calcium carbonate minerals using an engineered microbial system. Through this approach, a process to manufacture cementitious materials with carbon sequestration ability could be developed.

CHEK2 is not a Li-Fraumeni syndrome gene: time to update public resources.

The gene-disease relationship for CHEK2 remains listed as 'Li-Fraumeni syndrome 2' in public resources such as OMIM and MONDO, despite published evidence to the contrary, causing frustration among Li-Fraumeni syndrome (LFS) clinical experts. Here, we compared personal cancer characteristics of 2095 CHEK2 and 248 TP53 pathogenic variant carriers undergoing multigene panel testing at Ambry Genetics against 15 135 individuals with no known pathogenic variant. Our results from a within-cohort logistic regression approach highlight obvious differences between clinical presentation of TP53 and CHEK2 pathogenic variant carriers, with no evidence of CHEK2 being associated with any of the TP53-related core LFS cancers. These findings emphasise the need to replace 'Li-Fraumeni syndrome 2' as the CHEK2-associated disease name, thereby limiting potential confusion.

Contribution of large genomic rearrangements in PALB2 to familial breast cancer: implications for genetic testing.

BACKGROUND: PALB2 is the most important contributor to familial breast cancer after BRCA1 and BRCA2. Large genomic rearrangements (LGRs) in BRCA1 and BRCA2 are routinely assessed in clinical testing and are a significant contributor to the yield of actionable findings. In contrast, the contribution of LGRs in PALB2 has not been systematically studied. METHODS: We performed targeted sequencing and real-time qPCR validation to identify LGRs in PALB2 in 5770 unrelated patients with familial breast cancer and 5741 cancer-free control women from the same Australian population. RESULTS: Seven large deletions ranging in size from 0.96 kbp to 18.07 kbp involving PALB2 were identified in seven cases, while no LGRs were identified in any of the controls. Six LGRs were considered pathogenic as they included one or more exons of PALB2 and disrupted the WD40 domain at the C terminal end of the PALB2 protein while one LGR only involved a partial region of intron 10 and was considered a variant of unknown significance. Altogether, pathogenic LGRs identified in this study accounted for 10.3% (6 of 58) of the pathogenic PALB2 variants detected among the 5770 families with familial breast cancer. CONCLUSIONS: Our data show that a clinically important proportion of PALB2 pathogenic mutations in Australian patients with familial breast cancer are LGRs. Such observations have provided strong support for inclusion of PALB2 LGRs in routine clinical genetic testing.

Population-based BRCA1/2 testing programmes are highly acceptable in the Jewish community: results of the JeneScreen Study.

BACKGROUND: Ashkenazi Jewish (AJ) people have a higher incidence of BRCA1/2 pathogenic variants (PVs) than unselected populations. Three BRCA-Jewish founder mutations (B-JFMs) comprise >90% of BRCA1/2 PVs in AJ people. Personal/family cancer history-based testing misses ≥50% of people with B-JFM. METHODS: We compared two population-based B-JFM screening programmes in Australia-using (1) an online tool (Sydney) and (2) in-person group sessions (Melbourne). RESULTS: Of 2167 Jewish people tested (Sydney n=594; Melbourne n=1573), 1.3% (n=28) have a B-JFM, only 2 of whom had a significant cancer family history (Manchester score ≥12). Pretest anxiety scores were normal (mean 9.9±3.5 (6-24)), with no significant post-result change (9.5±3.3). Decisional regret (mean 7.4±13.0 (0-100)), test-related distress (mean 0.8+/2.2 (0-30)) and positive experiences (reverse-scored) (mean 3.4±4.5 (1-20)) scores were low, with no significant differences between Sydney and Melbourne participants. Post-education knowledge was good overall (mean 11.8/15 (±2.9)) and significantly higher in Melbourne than Sydney. Post-result knowledge was the same (mean 11.7 (±2.4) vs 11.2 (±2.4)). Participants with a B-JFM had higher post-result anxiety and test-related distress and lower positive experiences, than those without a B-JFM, but scores were within the normal range. Family cancer history did not significantly affect knowledge or anxiety, or pretest perception of B-JFM or cancer risks. Most participants (93%) were satisfied/very satisfied with the programme. CONCLUSION: Both B-JFM screening programmes are highly acceptable to Australian Jewish communities. The programme enabled identification of several individuals who were previously unaware they have a B-JFM, many of whom would have been ineligible for current criteria-based testing in Australia.

Epidemiology and management of oral cavity squamous cell carcinoma in Ireland.

INTRODUCTION: The epidemiology and management of oral cavity cancer have changed considerably in recent decades. This study examines epidemiological and management trends in oral cavity squamous cell carcinoma (OCSCC). METHODS: A retrospective cohort study of data from the National Cancer Registry of Ireland between 1994 and 2014. RESULTS: A total of 2725 patients were identified. The most common subsites were the tongue (34 %, n = 1025), lip (19 %, n = 575), floor of mouth (FOM) (18 %, n = 550), and retromolar trigone (RMT) (6 %, n = 189). The incidence of OCSCC remained largely unchanged (3.14 cases/100000/year) during the study period. 5-year disease-specific survival (DSS) was 58.6 % overall, varying between subsites (lip 85 %, RMT 62.9 %, tongue 54.7 %, and FOM 47.3 %). DSS improved over the study period (p = 0.03), in particular for tongue primaries (p = 0.007). Primary surgery significantly improved DSS versus radiotherapy (HR 0.28, p < 0.0001). Survival of T4 disease managed surgically was superior to that of T1 disease managed with radiotherapy. In node positive patients, chemotherapy improved overall survival (HR 0.8 p = 0.038) but not DSS (HR 0.87 p = 0.215). CONCLUSION: Primary surgery remains the standard of care in the management of OCSCC. Prognosis has improved in line with an increase in the use of primary surgery in the same time frame, though the incidence remains unchanged.

[Standardized risk-based antibiotic prophylaxis and adhesive film dressing for the prevention of wound infection following cochlear implantation]. / Standardisierte risikobasierte Antibiotikaprophylaxe und adhäsiver Filmverband zur Prävention von Wundinfektion bei Cochleaimplantation.

BACKGROUND: There is no consensus in the pertinent literature regarding the optimal antibiotic prophylaxis (AP) for cochlear implantation (CI). This study evaluates the implementation of standardized risk-based AP combined with application of an adhesive film dressing. MATERIALS AND METHODS: All CI cases since September 2019 were retrospectively reviewed for postoperative wound complications. While all patients received preoperative AP with ceftriaxone, postoperative AP after CI in patients older than 7 years was no longer routinely performed in our clinic. Exceptions were made according to predefined criteria for an increased risk of infection. The wound was covered with a transparent adhesive polyurethane film. RESULTS: In 72% of the 219 cases, we did not perform postoperative AP. The overall wound complication rate was 2.7% (in the groups with and without postoperative AP, 4.9% and 1.9%, respectively). Wound infection did not occur in any of the patients without postoperative AP older than 70 years (n = 32), with controlled diabetes mellitus (n = 19), or with reimplantation due to technical defect (n = 19). The film did not need to be changed until the suture material was removed. CONCLUSION: Standardized risk-based AP can avoid prolonged administration of antibiotics in selected patients. The film dressing permits continual examination and sufficient wound protection.

Ovarian cancer pathology characteristics as predictors of variant pathogenicity in BRCA1 and BRCA2.

BACKGROUND: The distribution of ovarian tumour characteristics differs between germline BRCA1 and BRCA2 pathogenic variant carriers and non-carriers. In this study, we assessed the utility of ovarian tumour characteristics as predictors of BRCA1 and BRCA2 variant pathogenicity, for application using the American College of Medical Genetics and the Association for Molecular Pathology (ACMG/AMP) variant classification system. METHODS: Data for 10,373 ovarian cancer cases, including carriers and non-carriers of BRCA1 or BRCA2 pathogenic variants, were collected from unpublished international cohorts and consortia and published studies. Likelihood ratios (LR) were calculated for the association of ovarian cancer histology and other characteristics, with BRCA1 and BRCA2 variant pathogenicity. Estimates were aligned to ACMG/AMP code strengths (supporting, moderate, strong). RESULTS: No histological subtype provided informative ACMG/AMP evidence in favour of BRCA1 and BRCA2 variant pathogenicity. Evidence against variant pathogenicity was estimated for the mucinous and clear cell histologies (supporting) and borderline cases (moderate). Refined associations are provided according to tumour grade, invasion and age at diagnosis. CONCLUSIONS: We provide detailed estimates for predicting BRCA1 and BRCA2 variant pathogenicity based on ovarian tumour characteristics. This evidence can be combined with other variant information under the ACMG/AMP classification system, to improve classification and carrier clinical management.

Management of individuals with germline pathogenic/likely pathogenic variants in CHEK2: A clinical practice resource of the American College of Medical Genetics and Genomics (ACMG).

PURPOSE: Although the role of CHEK2 germline pathogenic variants in cancer predisposition is well known, resources for managing CHEK2 heterozygotes in clinical practice are limited. METHODS: An international workgroup developed guidance on clinical management of CHEK2 heterozygotes informed by peer-reviewed publications from PubMed. RESULTS: Although CHEK2 is considered a moderate penetrance gene, cancer risks may be considered as a continuous variable, which are influenced by family history and other modifiers. Consequently, early cancer detection and prevention for CHEK2 heterozygotes should be guided by personalized risk estimates. Such estimates may result in both downgrading lifetime breast cancer risks to those similar to the general population or upgrading lifetime risk to a level at which CHEK2 heterozygotes are offered high-risk breast surveillance according to country-specific guidelines. Risk-reducing mastectomy should be guided by personalized risk estimates and shared decision making. Colorectal and prostate cancer surveillance should be considered based on assessment of family history. For CHEK2 heterozygotes who develop cancer, no specific targeted medical treatment is recommended at this time. CONCLUSION: Systematic prospective data collection is needed to establish the spectrum of CHEK2-associated cancer risks and to determine yet-unanswered questions, such as the outcomes of surveillance, response to cancer treatment, and survival after cancer diagnosis.

Development and evaluation of a novel educational program for providers on the use of polygenic risk scores.

PURPOSE: This study aimed to develop an online educational program for using polygenic risk score (PRS) for breast and ovarian cancer risk assessments and to evaluate the impact on the attitudes, confidence, knowledge, and preparedness of genetic health care providers (GHPs). METHODS: The educational program comprises an online module that covers the theoretical aspects of PRS and a facilitated virtual workshop with prerecorded role-plays and case discussions. Data were collected in pre- and posteducation surveys. Eligible participants were GHPs working in Australian familial cancer clinics registered to recruit patients for a breast and ovarian cancer PRS clinical trial (n = 12). RESULTS: A total of 124 GHPs completed the PRS education, of whom 80 (64%) and 67 (41%) completed the pre- and posteducation surveys, respectively. Before education, GHPs reported limited experience, confidence, and preparedness using PRS, but they recognized its potential benefits. After education, GHPs indicated improved attitudes (P ≤ .001), confidence (P ≤ .001), knowledge (P ≤ .001), and preparedness (P ≤ .001) to use PRS. Most GHPs thought that the program entirely met their learning needs (73%) and was completely relevant to their clinical practice (88%). GHPs identified PRS implementation barriers, including limited funding models, diversity issues, and need for clinical guidelines. CONCLUSION: Our education program improved GHP attitudes, confidence, knowledge, and preparedness for using PRS/personalized risk and provides a framework for the development of future programs.

StomaAI: an efficient and user-friendly tool for measurement of stomatal pores and density using deep computer vision.

Using microscopy to investigate stomatal behaviour is common in plant physiology research. Manual inspection and measurement of stomatal pore features is low throughput, relies upon expert knowledge to record stomatal features accurately, requires significant researcher time and investment, and can represent a significant bottleneck to research pipelines. To alleviate this, we introduce StomaAI (SAI): a reliable, user-friendly and adaptable tool for stomatal pore and density measurements via the application of deep computer vision, which has been initially calibrated and deployed for the model plant Arabidopsis (dicot) and the crop plant barley (monocot grass). SAI is capable of producing measurements consistent with human experts and successfully reproduced conclusions of published datasets. SAI boosts the number of images that can be evaluated in a fraction of the time, so can obtain a more accurate representation of stomatal traits than is routine through manual measurement. An online demonstration of SAI is hosted at https://sai.aiml.team, and the full local application is publicly available for free on GitHub through https://github.com/xdynames/sai-app.

Nonpharmaceutical interventions reduce the incidence and mortality of COVID-19: A study based on the survey from the International COVID-19 Research Network (ICRN).

The recently emerged novel coronavirus, "severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2)," caused a highly contagious disease called coronavirus disease 2019 (COVID-19). It has severely damaged the world's most developed countries and has turned into a major threat for low- and middle-income countries. Since its emergence in late 2019, medical interventions have been substantial, and most countries relied on public health measures collectively known as nonpharmaceutical interventions (NPIs). We aimed to centralize the accumulative knowledge of NPIs against COVID-19 for each country under one worldwide consortium. International COVID-19 Research Network collaborators developed a cross-sectional online survey to assess the implications of NPIs and sanitary supply on the incidence and mortality of COVID-19. The survey was conducted between January 1 and February 1, 2021, and participants from 92 countries/territories completed it. The association between NPIs, sanitation supplies, and incidence and mortality were examined by multivariate regression, with the log-transformed value of population as an offset value. The majority of countries/territories applied several preventive strategies, including social distancing (100.0%), quarantine (100.0%), isolation (98.9%), and school closure (97.8%). Individual-level preventive measures such as personal hygiene (100.0%) and wearing facial masks (94.6% at hospitals; 93.5% at mass transportation; 91.3% in mass gathering facilities) were also frequently applied. Quarantine at a designated place was negatively associated with incidence and mortality compared to home quarantine. Isolation at a designated place was also associated with reduced mortality compared to home isolation. Recommendations to use sanitizer for personal hygiene reduced incidence compared to the recommendation to use soap. Deprivation of masks was associated with increased incidence. Higher incidence and mortality were found in countries/territories with higher economic levels. Mask deprivation was pervasive regardless of economic level. NPIs against COVID-19 such as using sanitizer, quarantine, and isolation can decrease the incidence and mortality of COVID-19.

Validity evidence for observational ERCP competency assessment tools: a systematic review.

BACKGROUND : Assessment of competence in endoscopic retrograde cholangiopancreatography (ERCP) is critical for supporting learning and documenting attainment of skill. Validity evidence supporting ERCP observational assessment tools has not been systematically evaluated. METHODS : We conducted a systematic search using electronic databases and hand-searching from inception until August 2021 for studies evaluating observational assessment tools of ERCP performance. We used a unified validity framework to characterize validity evidence from five sources: content, response process, internal structure, relations to other variables, and consequences. Each domain was assigned a score of 0-3 (maximum score 15). We assessed educational utility and methodological quality using the Accreditation Council for Graduate Medical Education framework and the Medical Education Research Quality Instrument, respectively. RESULTS : From 2769 records, we included 17 studies evaluating 7 assessment tools. Five tools were studied for clinical ERCP, one for simulated ERCP, and one for simulated and clinical ERCP. Validity evidence scores ranged from 2 to 12. The Bethesda ERCP Skills Assessment Tool (BESAT), ERCP Direct Observation of Procedural Skills Tool (ERCP DOPS), and The Endoscopic Ultrasound (EUS) and ERCP Skills Assessment Tool (TEESAT) had the strongest validity evidence, with scores of 10, 12, and 11, respectively. Regarding educational utility, most tools were easy to use and interpret, and required minimal additional resources. Overall methodological quality (maximum score 13.5) was strong, with scores ranging from 10 to 12.5. CONCLUSIONS : The BESAT, ERCP DOPS, and TEESAT had strong validity evidence compared with other assessments. Integrating tools into training may help drive learners' development and support competency decision making.

DDOST-CDG: Clinical and molecular characterization of a third patient with a milder and a predominantly movement disorder phenotype.

Congenital disorders of glycosylation (CDG) are a group of heterogeneous inherited metabolic disorders affecting posttranslational protein modification. DDOST-CDG, caused by biallelic pathogenic variants in DDOST which encodes dolichyl-diphospho-oligosaccharide-protein glycosyltransferase, a subunit of N-glycosylation oligosaccharyltransferase (OST) complex, is an ultra-rare condition that has been described in two patients only. The main clinical features in the two reported patients include profound developmental delay, failure to thrive, and hypotonia. In addition, both patients had abnormal transferrin glycosylation. Here, we report an 18-year-old male who presented with moderate developmental delay, progressive opsoclonus, myoclonus, ataxia, tremor, and dystonia. Biochemical studies by carbohydrate deficient transferrin analysis showed a type I CDG pattern. Exome sequencing identified compound heterozygous variants in DDOST: a maternally inherited variant, c.1142dupT (p.Leu381Phefs*11), and a paternally inherited variant, c.661 T > C (p.Ser221Pro). Plasma N-glycan profiling showed mildly increased small high mannose glycans including Man0-5 GlcNAc2, a pattern consistent with what was previously reported in DDOST-CDG or defects in other subunits of OST complex. Western blot analysis on patient's fibroblasts revealed decreased expression of DDOST and reduced intracellular N-glycosylation, as evident by the biomarkers ICAM-1 and LAMP2. Our study highlights the clinical variability, expands the clinical and biochemical phenotypes, and describes new genotype, which all are essential for diagnosing and managing patients with DDOST-CDG.

Fractionated plasma N-glycan profiling of novel cohort of ATP6AP1-CDG subjects identifies phenotypic association.

ATP6AP1-CDG is an X-linked disorder typically characterized by hepatopathy, immunodeficiency, and an abnormal type II transferrin glycosylation pattern. Here, we present 11 new patients and clinical updates with biochemical characterization on one previously reported patient. We also document intrafamilial phenotypic variability and atypical presentations, expanding the symptomatology of ATP6AP1-CDG to include dystonia, hepatocellular carcinoma, and lysosomal abnormalities on hepatic histology. Three of our subjects received successful liver transplantation. We performed N-glycan profiling of total and fractionated plasma proteins for six patients and show associations with varying phenotypes, demonstrating potential diagnostic and prognostic value of fractionated N-glycan profiles. The aberrant N-linked glycosylation in purified transferrin and remaining plasma glycoprotein fractions normalized in one patient post hepatic transplant, while the increases of Man4GlcNAc2 and Man5GlcNAc2 in purified immunoglobulins persisted. Interestingly, in the single patient with isolated immune deficiency phenotype, elevated high-mannose glycans were detected on purified immunoglobulins without glycosylation abnormalities on transferrin or the remaining plasma glycoprotein fractions. Given the diverse and often tissue specific clinical presentations and the need of clinical management post hepatic transplant in ATP6AP1-CDG patients, these results demonstrate that fractionated plasma N-glycan profiling could be a valuable tool in diagnosis and disease monitoring.

Universal genetic testing for women with newly diagnosed breast cancer in the context of multidisciplinary team care.

OBJECTIVE: To determine the feasibility of universal genetic testing of women with newly diagnosed breast cancer, to estimate the incidence of pathogenic gene variants and their impact on patient management, and to evaluate patient and clinician acceptance of universal testing. DESIGN, SETTING, PARTICIPANTS: Prospective study of women with invasive or high grade in situ breast cancer and unknown germline status discussed at the Parkville Breast Service (Melbourne) multidisciplinary team meeting. Women were recruited to the pilot (12 June 2020 - 22 March 2021) and expansion phases (17 October 2021 - 8 November 2022) of the Mutational Assessment of newly diagnosed breast cancer using Germline and tumour genomICs (MAGIC) study. MAIN OUTCOME MEASURES: Germline testing by DNA sequencing, filtered for nineteen hereditary breast and ovarian cancer genes that could be classified as actionable; only pathogenic variants were reported. Surveys before and after genetic testing assessed pilot phase participants' perceptions of genetic testing, and psychological distress and cancer-specific worry. A separate survey assessed clinicians' views on universal testing. RESULTS: Pathogenic germline variants were identified in 31 of 474 expanded study phase participants (6.5%), including 28 of 429 women with invasive breast cancer (6.5%). Eighteen of the 31 did not meet current genetic testing eligibility guidelines (probability of a germline pathogenic variant ≥ 10%, based on CanRisk, or Manchester score ≥ 15). Clinical management was changed for 24 of 31 women after identification of a pathogenic variant. Including 68 further women who underwent genetic testing outside the study, 44 of 542 women carried pathogenic variants (8.1%). Acceptance of universal testing was high among both patients (90 of 103, 87%) and clinicians; no decision regret or adverse impact on psychological distress or cancer-specific worry were reported. CONCLUSION: Universal genetic testing following the diagnosis of breast cancer detects clinically significant germline pathogenic variants that might otherwise be missed because of testing guidelines. Routine testing and reporting of pathogenic variants is feasible and acceptable for both patients and clinicians.