Pathogenic Variants In Other Genes Research Articles

Updated European guidelines for clinical management of familial adenomatous polyposis (FAP), MUTYH-associated polyposis (MAP), gastric adenocarcinoma, proximal polyposis of the stomach (GAPPS) and other rare adenomatous polyposis syndromes: a joint EHTG-ESCP revision.

Hereditary adenomatous polyposis syndromes, including familial adenomatous polyposis and other rare adenomatous polyposis syndromes, increase the lifetime risk of colorectal and other cancers. A team of 38 experts convened to update the 2008 European recommendations for the clinical management of patients with adenomatous polyposis syndromes. Additionally, other rare monogenic adenomatous polyposis syndromes were reviewed and added. Eighty-nine clinically relevant questions were answered after a systematic review of the existing literature with grading of the evidence according to Grading of Recommendations, Assessment, Development, and Evaluation methodology. Two levels of consensus were identified: consensus threshold (≥67% of voting guideline committee members voting either 'Strongly agree' or 'Agree' during the Delphi rounds) and high threshold (consensus ≥ 80%). One hundred and forty statements reached a high level of consensus concerning the management of hereditary adenomatous polyposis syndromes. These updated guidelines provide current, comprehensive, and evidence-based practical recommendations for the management of surveillance and treatment of familial adenomatous polyposis patients, encompassing additionally MUTYH-associated polyposis, gastric adenocarcinoma and proximal polyposis of the stomach and other recently identified polyposis syndromes based on pathogenic variants in other genes than APC or MUTYH. Due to the rarity of these diseases, patients should be managed at specialized centres.

Cornelia de Lange Spectrum

Cornelia de Lange syndrome (CdLS) is a rare congenital developmental disorder with multisystemic involvement. The clinical presentation is highly variable, but the classic phenotype, characterized by distinctive craniofacial features, pre- and postnatal growth retardation, extremity reduction defects, hirsutism and intellectual disability can be distinguished from the nonclassic phenotype, which is generally milder and more difficult to diagnose. In addition, the clinical features overlap with those of other neurodevelopmental disorders, so the use of consensus clinical criteria and artificial intelligence tools may be helpful in confirming the diagnosis.Pathogenic variants in NIPBL, which encodes a protein related to the cohesin complex, have been identified in more than 60% of patients, and pathogenic variants in other genes related to this complex in another 15%: SMC1A, SMC3, RAD21, and HDAC8. Technical advances in large-scale sequencing have allowed the description of additional genes (BRD4, ANKRD11, MAU2), but the lack of molecular diagnosis in 15% of individuals and the substantial clinical heterogeneity of the syndrome suggest that other genes and mechanisms may be involved.Although there is no curative treatment, there are symptomatic/palliative treatments that paediatricians should be aware of. The main medical complication in classic SCdL is gastro-esophageal reflux (GER), which should be treated early.

Detection of Germline Mutations in a Cohort of 250 Relatives of Mutation Carriers in Multigene Panel: Impact of Pathogenic Variants in Other Genes beyond BRCA1/2.

Several hereditary-familial syndromes associated with various types of tumors have been identified to date, evidencing that hereditary cancers caused by germline mutations account for 5-10% of all tumors. Advances in genetic technology and the implementation of Next-Generation Sequencing (NGS) have accelerated the discovery of several susceptibility cancer genes, allowing for the detection of cancer-predisposing mutations in a larger number of cases. The aim of this study is to highlight how the application of an NGS-multigene panel to a group of oncological patients subsequently leads to improvement in the identification of carriers of healthy pathogenic variants/likely pathogenic variants (PVs/LPVs) and prevention of the disease in these cases. Starting from a total of 110 cancer patients carrying PVs/LPVs in genes involved in cancer susceptibility detected via a customized NGS panel of 27 cancer-associated genes, we enrolled 250 healthy collateral family members from January 2020 to July 2022. The specific PVs/LPVs identified in each proband were tested in healthy collateral family members via Sanger sequencing. A total of 131 out of the 250 cases (52%) were not carriers of the mutation detected in the affected relative, while 119 were carriers. Of these, 81/250 patients carried PVs/LPVs on BRCA1/2 (33%), 35/250 harbored PVs/LPVs on other genes beyond BRCA1 and BRCA2 (14%), and 3/250 (1%) were PVs/LPVs carriers both on BRCA1/2 and on another susceptibility gene. Our results show that the analysis of BRCA1/2 genes would have only resulted in a missed diagnosis in a number of cases and in the lack of prevention of the disease in a considerable percentage of healthy carriers with a genetic mutation (14%).

Genetic and phenotypic characterization of Nexilin (NEXN) related cardiomyopathy

Abstract Background Clinical and genetic features of Nexilin (NEXN) related cardiomyopathies (CMPs) are largely uncharacterized. At present, the strength of the gene-disease association for NEXN is currently moderate for Dilated Cardiomyopathy (DCM), limited for Hypertrophic Cardiomyopathy (HCM), and unevaluated for Arrhythmogenic Cardiomyopathy (ACM). More evidence is needed to establish a definite genotype-phenotype association. Purpose We sought to investigate 1) the causative role of NEXN gene in CMPs, and 2) its phenotypic expression and prognostic profile. Methods All patients carrying NEXN variants, recruited in 11 international referral centres, were classified according to reported phenotype and genetic variant. Variants were considered rare if below GnomAD Popmax Filtering Allele Frequency of 8x10-5. Burden enrichment testing of rare NEXN variants was performed in our case-cohort of patients with DCM/ACM (n=3053), in comparison with the GnomAD population of non-Finnish Europeans (NFE). Clinical phenotypes and outcomes of patients carrying validated variants were described. A previously established cohort of patients with Titin-related CMP was used for prognostic comparison. Results Data from 87 patients were collected. Forty patients were excluded due to being carriers of non-rare NEXN variants (n=22) and/or pathogenic variants in other genes (n=18). Of the remaining patients, only 2 had HCM diagnosis, precluding any further analysis on this phenotype. NEXN truncating variants (tv) resulted significantly enriched in DCM/ACM disease-cohort with a prevalence of 0.39% vs 0.09% in GnomAD NFE (p=0.0001), whereas non-truncating ones were not. Patients carrying NEXNtv (n=16), plus a further patient carrying a homozygous p.G650del variant were thus considered affected by NEXN-related CMP (NEXN-CMP). Out of 17 patients with NEXN-CMP, 82% were probands, 53% were males and median age at diagnosis was 46 (IQR 35-55). At baseline, the most prevalent phenotypes were DCM (53%) and ACM (41%) and NYHA I class was common (71%). Left ventricular (LV) indexed End Diastolic Volume was mildly increased (75mL, IQR 43-87) and ejection fraction mildly reduced (LVEF 41%±15). Myocardial fibrosis was reported for up to 67% of patients, median ventricular ectopic beats were 515/24h (IQR 43-5071). During a median follow of 42 months, 8 patients were implanted with ICD, 1 patient died and 3 (21%) had malignant ventricular arrythmias (MVA). When compared to a cohort of patients with TTN-CMP, patients with NEXN-CMP showed MVA earlier, and with higher LVEF. Conclusions NEXNtv showed significant disease association with DCM/ACM phenotypes. NEXN-CMP seems to be characterized by mild LV dilation and dysfunction, uncommon heart failure, but frequent myocardial fibrosis and ventricular arrythmias. These findings from the largest cohort of NEXN variants carriers strongly contribute to defining the causative role of this rare genotype and its related phenotype.

Determination of Carrier Frequency of Actionable Pathogenic Variants in Autosomal Recessive Genetic Diseases in the Turkish Cypriot Population.

Whole-exome DNA sequencing is a rich source of clinically useful information for specialists, patients, and their families, as well as elucidating the genetic basis of monogenic and complex diseases in clinical diagnosis. However, interpreting and reporting variants encompassing exome and genome sequence analysis outcome data are one of the greatest challenges of the genomic era. In this study, we aimed to investigate the frequency and allele frequency spectrum of single nucleotide variants accepted as recessive disease carrier status in Turkish Cypriot exomes. The same sequencing platform and data processing line were used for the analysis of data from 100 Turkish Cypriot whole-exome sequence analysis. Identified variants were classified according to ACMG guidelines, and pathogenic variants were confirmed in other databases such as ClinVar, HGMD, Varsome, etc. Pathogenic variants were detected in 68 genes out of 100 whole-exome sequence data. The carriage rate was the highest in the CYP21A2 gene, causing 21-hydroxylase deficiency (14.70%), 11.76% in the HBB gene causing β-thalassemia, 10.29% in the BTD gene causing biotinidase deficiency, 8.82% in the CFTR gene causing cystic fibrosis, 8.82% in the RBM8A gene causing thrombocytopenia-absent radius syndrome, which is an ultra-rare disease, and 5.88% in the GAA gene causing glycogen storage disease II. The carriage of pathogenic variants in other genes causing the disease (GJB2, PAH, GALC, CYP11B2, COL4A3, HBA1, etc.) was determined as less than 5.00%. Also, the identified variations in the mentioned gene within the examined population were reported. The most prevalent mutation in North Cyprus was a missense variant (c.1360 C>T, p.Pro454Ser) detected in the CYP21A2 gene (rs6445), and the most frequently seen variant in the HBB gene was c.93-21G>A (rs35004220). We investigated reported pathogenic variants by estimating the lower and upper limits of carrier and population frequencies for autosomal recessive diseases, for which exome sequencing may reveal additional medically relevant information. Determining the lower and upper limits of these frequencies will shed light on preventive medicine practices and governmental actions.

Expanding the genetic and clinical spectrum of osteogenesis imperfecta: identification of novel rare pathogenic variants in type I collagen-encoding genes.

Osteogenesis imperfecta (OI) is a clinically and genetically heterogeneous skeletal disorder. The majority of affected cases are attributed to autosomal dominant pathogenic variants (PVs) found in the COL1A1 and COL1A2 genes, which encode type I collagen. However, PVs in other genes involved in collagen posttranslational modification, processing, crosslinking, osteoblast differentiation, and bone mineralization have also been associated with OI. In this study, we present the results of next-generation sequencing (NGS) analysis using a custom panel of 11 genes known to be associated with OI. This clinical study enrolled a total of 10 patients, comprising 7 male and 3 female patients from 7 families, all from the Puglia Region in South Italy, providing a detailed overview of their age, gender, family history, OI type, and non-skeletal features. The genetic analysis revealed 5 PVs in the COL1A1 gene and 2 PVs in the COL1A2 gene. Importantly, three of these PVs have not been previously reported in the literature. These include two novel heterozygous frameshift PVs in COL1A1 (c.2890_2893del and c.3887del) and one novel heterozygous missense PV in COL1A2 (c.596G>T). The identification of these previously unreported PVs expands the variant spectrum of the COL1A1 and COL1A2 genes and may have implications for accurate diagnosis, genetic counselling, and potential therapeutic interventions in affected individuals and their families.

Shwachman–Diamond syndrome: a hematologist's view

Shwachman–Diamond syndrome is a rare genetic disorder with an autosomal recessive inheritance pattern. Most often (in more than 90% of cases) this disease is caused by biallelic pathogenic variants in the highly conserved SBDS gene located on the long arm of chromosome 7. However, approximately 10% of patients with the clinical phenotype of Shwachman–Diamond syndrome lack mutations in SBDS but have pathogenic variants in other genes, such as DNAJC21 or EFL1. Shwachman–Diamond syndrome is a multisystemic disorder characterized by exocrine pancreatic insufficiency, protein-energy undernutrition, delayed physical development, cognitive disorders, anomalies of the skeletal system, and immunological disorders. In addition to the described symptoms, Shwachman–Diamond syndrome is characterized by the presence of bone marrow failure (most often neutropenia and anemia), as well as an increased risk of cytogenetic abnormalities and a predisposition to myelodysplastic syndromes and acute myeloid leukemia. In this review, the authors summarize the spectrum of hematological disorders observed in Shwachman–Diamond syndrome, as well as describe the molecular mechanisms underlying them.

Guideline-Based, Multi-Gene Panel Germline Genetic Testing for at-Risk Patients with Breast Cancer.

Genetic testing for at-risk patients with breast cancer should be routinely offered. Knowledge generated may influence both treatment decisions and cancer prevention strategies among the patients themselves and their relatives. In this study, we report on the prevalence and patterns of germline mutations, using commercially available next-generation sequencing (NGS)-based multi-gene panels (MGP). Consecutive at-risk breast cancer patients, as determined by international guidelines, were offered germline genetic testing using a 20-gene NGS-based panel at a reference lab. Samples of peripheral blood were obtained for DNA extraction and genetic variants were classified as benign/likely benign (negative), pathogenic/likely pathogenic (positive) or variants of uncertain significance (VUS). A total of 1310 patients, median age (range) 43 (19-82) years, were enrolled. Age ≤45 years (n = 800, 61.1%) was the most common indication for testing. Positive family history of breast, ovarian, pancreatic or prostate cancers, and triple-negative disease were among the common indications. Among the whole group, 184 (14.0%) patients had pathogenic/likely pathogenic variants; only 90 (48.9%) were in BRCA1 or BRCA2, while 94 (51.9%) others had pathogenic variants in other genes; mostly in APC, TP53, CHEK2 and PALB2. Mutation rates were significantly higher among patients with positive family history (p = 0.009); especially if they were 50 years or younger at the time of breast cancer diagnosis (p < 0.001). Patients with triple-negative disease had relatively higher rate (17.5%), and mostly in BRCA1/2 genes (71.4%). Variants of uncertain significance (VUS) were reported in 559 (42.7%) patients; majority (90.7%) were in genes other than BRCA1 or BRCA2. Pathogenic mutations in genes other than BRCA1/2 are relatively common and could have been missed if genetic testing was restricted to BRCA1/2. The significantly high rate of VUS associated with multi-gene panel testing can be disturbing.

Haemochromatosis revisited.

Haemochromatosis is a genetic disease caused by hepcidin deficiency, responsible for an increase in intestinal iron absorption. Haemochromatosis is associated with homozygosity for the HFE p.Cys282Tyr mutation. However, rare cases of haemochromatosis (non-HFE haemochromatosis) can also be caused by pathogenic variants in other genes (such as HJV, HAMP, TFR2 and SLC40A1). A working group of the International Society for the Study of Iron in Biology and Medicine (BIOIRON Society) has concluded that the classification based in different molecular subtypes is difficult to be adopted in clinical practice and has proposed a new classification approaching clinical questions and molecular complexity. The aim of the present review is to provide an update on classification, pathophysiology and therapeutic recommendations.

Splice-site variant in the RPS7 5'-UTR leads to a decrease in the mRNA level and development of Diamond-Blackfan anemia.

Diamond-Blackfan anemia (DBA) is an inherited bone marrow failure syndrome characterized by erythroid aplasia. Pathogenic variants in ribosomal protein (RP) genes, GATA1, TSR2, and EPO, are considered to be the etiology of DBA. Variants in 5'-untranslated regions (UTRs) of these genes are poorly studied and can complicate the variant interpretation. We investigated the functional consequences NM_001011.4:c.-19 + 1G > T variant in the donor splice-site of the RPS7 5'-UTR. This variant was found in a family where two sons with DBA were carriers. Father, who also had this variant, developed myelodysplastic syndrome, which caused his death. Search for candidate causal variants and copy number variations in DBA-associated genes left RPS7 variant as the best candidate. Trio whole exome sequencing analysis revealed no pathogenic variants in other genes. Functional analysis using luciferase expression system revealed that this variant leads to disruption of splicing. Also, a decrease in the levels of mRNA and protein expression was detected. In conclusion, the established consequences of 5'-UTR splice-site variant c.-19 + 1G > T in the RPS7 gene provide evidence that it is likely pathogenic.

Molecular and clinical profile of patients referred as Noonan or Noonan-like syndrome in Greece: a cohort of 86 patients.

Craniofacial dysmorphism and PS prevail among pathogenic variant positive compared to pathogenic variant negative NS and NS-like patients while neurological defects are less common in PTPN11-affected NS patients compared to patients harbouring pathogenic variants in other genes. The significant prevalence of the Ras/MAPK pathogenic variants (17.4%), other than PTPN11, in Greek NS patients, highlights the necessity of a wider spectrum of molecular diagnosis. •Noonan syndrome (NS) has been associated with pathogenic variants in molecules-components of the Ras/MAPK pathway. • Clinical and genetic description of NS patients worldwide helps establishing personalized monitoring. •NS and NS-like mutational rate in Greece reaches 50% with pathogenic variants identified mostly in PTPN11 (32.5%), RIT1 (6%) and SOS1 (4.7%) genes. • The risk for pulmonary stenosis increases 6.71-fold in NS patients with a pathogenic variant compared to patients without genetic alterations.

Influence of germline test results on surgical decision making in women with invasive breast cancer

BackgroundWhile therapeutic mastectomy with contralateral prophylactic mastectomy (TM+CPM) and/or bilateral salpingo-oophorectomy (BSO) are recommended for women with pathogenic variants (PV) in some cancer predisposition genes, evidence for the utility of these surgeries for women with PV in other genes currently is insufficient. In conjunction, current guidelines recommend that clinical management should not be influenced by a return of a variant of uncertain significance (VUS). Return of germline test results may, however, influence surgical decision making regardless of current guidelines. We thus evaluated surgical choices amongst a cohort of women with invasive breast cancer who underwent clinical genetic testing. MethodsGermline test results and all surgical procedures were extracted for women who had unilateral invasive breast cancer and had clinical testing before definitive surgery (n = 591). Results were classified as pathogenic/likely pathogenic (PV, 17.1%), VUS (19.5%) or benign/likely benign (63.4%). Data were analyzed using chi-square tests with p<0.05 defining significance. ResultsRates of TM+CPM and BSO were not significantly different for women with VUS compared to those with benign findings. Rates of TM+CPM were significantly higher for women with PV in BRCA1 and BRCA2, PALB2, PTEN and TP53, as well in genes with insufficient data to recommend risk-reducing mastectomy. Rates of BSO were significantly higher in women with PV in BRCA1 and BRCA2, PALB2, PTEN and TP53 and BRIP1, RAD51C and RAD51D compared to those with benign findings. ConclusionOverall, surgical choices for women with a VUS were more similar to those from women with benign variants than to those with PV, however, in the group with PV in genes for which insufficient evidence exists for the benefit of risk-reducing mastectomy, rates of TM+CPM were high. Thus, while the management of women with VUS is in agreement with ACMG guidelines, patients with mutations in other cancer genes demonstrate a preference for more aggressive breast surgeries.

Further delineation of SLC9A3‐related congenital sodium diarrhea

BackgroundCongenital sodium diarrhea (CSD) is a rare enteropathy displaying both broad variability in clinical severity and genetic locus and allelic heterogeneity. Eleven CSD patients were reported so far with SLC9A3 variants that impair the function of the encoded intestinal sodium‐proton exchanger 3 (NHE3).MethodsWe report a 4‐year‐old patient, born prematurely in the 35th week of gestation, with antenatal polyhydramnios and dilated intestinal loops, and with diarrhea of congenital onset, 2–6 times a day, and with polydipsia. She thrived age‐appropriately under a normal family diet. Serum sodium levels were repeatedly normal but urinary sodium excretion was low. Exome sequencing revealed compound heterozygous variants in SLC9A3 as the likely cause of the congenital diarrhea.ResultsWhile exome sequencing did not reveal pathogenic or likely pathogenic variants in other genes that cause syndromic or non‐syndromic forms of congenital and intractable diarrheas, we identified novel compound heterozygous variants in SLC9A3, a complex allele with two missense changes, NP_004165.2:p.[Ser331Leu;Val449Ile] and in‐trans the missense variant p.(Phe451Ser).ConclusionThe clinical phenotype here appears to localize to the milder end of the known CSD spectrum, and the identified variants suggest that this is the twelfth patient reported to date with CSD due to mutations in SLC9A3.

148P Descriptive study of mutation status and clinical phenotype in breast cancer patients of South Catalonia

The role of BRCA1 and BRCA2 germline pathogenic or likely pathogenic variants (PV) in hereditary breast cancer (HBC) is well established. PV in other genes can also confer an increased risk of cancer development; however, the evidence in their clinical phenotype characteristics remains scarce.

Pathogenic variants among Mexican patients with colorectal cancer referred for genetic cancer risk assessment.

69 Background: Lynch syndrome (LS) is the most frequent hereditary cancer syndrome among patients with colorectal cancer. Screening tests such as immunohistochemistry (IHC) for mismatch repair (MMR) proteins and PREMM5 model help to identify patients at risk of germline pathogenic variants (PVs). However, there has been a disparity in that evaluation of these screening tools and their correlation with pathogenic variants (PVs) has been limited in Hispanic populations. Methods: Patients with colorectal cancer referred for genetic cancer risk assessment were enrolled in the Clinical Cancer Genomics Community Research Network (CCGCRN) registry from October 2017 to February 2021 at the Instituto Nacional de Ciencias Médicas y Nutrición Salvador Zubirán. Genetic testing was performed by full sequencing of the MMR genes ( MLH1, PMS2, MSH2, MSH6 and EPCAM) and other cancer-associated genes ( APC, BRCA1, BRCA2, TP53, NF1, ATM, CHEK2, PALB2, BRIP1) and multiplex ligation dependent probe amplification to detect copy number variants (CNV) was performed for selected genes. Demographic, clinical characteristics and IHC results were obtained from clinical records. MMR PV probability was calculated using PREMM5. Results: Sixty-nine patients with colorectal cancer were included; mean age at diagnosis was 50 (26-82) years and 39/69 (56%) were women. A MMR gene PV was identified in 23/69 patients (33%); most frequently in MLH1 n = 14, followed by MSH2 n = 2, MSH6 n = 2 and PMS2 n = 1. Four recurrent PVs in MLH1 and MSH2 represented 22% of PVs. CNVs were identified in 4/23 (17.4%) patients with LS. PVs in other genes were identified in 8.6% of the cases: 2 ATM, 1 APC, 1 PALB2, 1 BRIP1 and 1 BRCA1. IHC results were available in 52/69 cases (75.4%) and MMR protein deficiency was found in 16/17 (94%) carriers and in 14/31 (45%) non-carriers (sensitivity 94.1% and specificity 54.8%). The area under the ROC curve for PREMM5 score was 0.94 (95% CI 0.88-0.99) with a mean score 31.6 (2.4-50) in patients with LS and 4.1 (0.9-50) in non-carriers. The diagnosis of a second primary colon cancer was more frequent among LS (30% vs 2.5%; p &lt; 0.01). Conclusions: We found a high frequency of MMR gene PVs among patients referred for GCRA with personal history of colon cancer, and only a small proportion with PVs in other genes. Our results showed a good performance of PREMM5 model and a high sensitivity of MMR IHQ in a Mexican population, indicating that these are tools that can be used to prioritize patient selection for germline testing.

Comprehensive Genetic Analysis Reveals Complexity of Monogenic Urinary Stone Disease

IntroductionBecause of phenotypic overlap between monogenic urinary stone diseases (USD), gene-specific analyses can result in missed diagnoses. We used targeted next generation sequencing (tNGS), including known and candidate monogenic USD genes, to analyze suspected primary hyperoxaluria (PH) or Dent disease (DD) patients genetically unresolved (negative; N) after Sanger analysis of the known genes. Cohorts consisted of 285 PH (PHN) and 59 DD (DDN) families.MethodsVariants were assessed using disease-specific and population databases plus variant assessment tools and categorized using the American College of Medical Genetics (ACMG) guidelines. Prior Sanger analysis identified 47 novel PH or DD gene pathogenic variants.ResultsScreening by tNGS revealed pathogenic variants in 14 known monogenic USD genes, accounting for 45 families (13.1%), 27 biallelic and 18 monoallelic, including 1 family with a copy number variant (CNV). Recurrent genes included the following: SLC34A3 (n = 13), CLDN16 (n = 8), CYP24A1 (n = 4), SLC34A1 (n = 3), SLC4A1 (n = 3), APRT (n = 2), CLDN19 (n = 2), HNF4A1 (n = 2), and KCNJ1 (n = 2), whereas ATP6V1B1, CASR, and SLC12A1 and missed CNVs in the PH genes AGXT and GRHPR accounted for 1 pedigree each. Of the 48 defined pathogenic variants, 27.1% were truncating and 39.6% were novel. Most patients were diagnosed before 18 years of age (76.1%), and 70.3% of biallelic patients were homozygous, mainly from consanguineous families.ConclusionOverall, in patients suspected of DD or PH, 23.9% and 7.3% of cases, respectively, were caused by pathogenic variants in other genes. This study shows the value of a tNGS screening approach to increase the diagnosis of monogenic USD, which can optimize therapies and facilitate enrollment in clinical trials.

Common molecular mechanisms of SLC6A1 variant-mediated neurodevelopmental disorders in astrocytes and neurons.

Solute carrier family 6 member 1 (SLC6A1) is abundantly expressed in the developing brain even before the CNS is formed. Its encoded GABA transporter 1 (GAT-1) is responsible for the reuptake of GABA into presynaptic neurons and glia, thereby modulating neurotransmission. GAT-1 is expressed globally in the brain, in both astrocytes and neurons. The GABA uptake function of GAT-1 in neurons cannot be compensated for by other GABA transporters, while the function in glia can be partially replaced by GABA transporter 3. Recently, many variants in SLC6A1 have been associated with a spectrum of epilepsy syndromes and neurodevelopmental disorders, including myoclonic atonic epilepsy, childhood absence epilepsy, autism, and intellectual disability, but the pathomechanisms associated with these phenotypes remain unclear. The presence of GAT-1 in both neurons and astrocytes further obscures the role of abnormal GAT-1 in the heterogeneous disease phenotype manifestations. Here we examine the impact on transporter trafficking and function of 22 SLC6A1 variants identified in patients with a broad spectrum of phenotypes. We also evaluate changes in protein expression and subcellular localization of the variant GAT-1 in various cell types, including neurons and astrocytes derived from human patient induced pluripotent stem cells. We found that a partial or complete loss-of-function represents a common disease mechanism, although the extent of GABA uptake reduction is variable. The reduced GABA uptake appears to be due to reduced cell surface expression of the variant transporter caused by variant protein misfolding, endoplasmic reticulum retention, and subsequent degradation. Although the extent of reduction of the total protein, surface protein, and the GABA uptake level of the variant transporters is variable, the loss of GABA uptake function and endoplasmic reticulum retention is consistent across induced pluripotent stem cell-derived cell types, including astrocytes and neurons, for the surveyed variants. Interestingly, we did not find a clear correlation of GABA uptake function and the disease phenotypes, such as myoclonic atonic epilepsy versus developmental delay, in this study. Together, our study suggests that impaired transporter protein trafficking and surface expression are the major disease-associated mechanisms associated with pathogenic SLC6A1 variants. Our results resemble findings from pathogenic variants in other genes affecting the GABA pathway, such as GABAA receptors. This study provides critical insight into therapeutic developments for SLC6A1 variant-mediated disorders and implicates that boosting transporter function by either genetic or pharmacological approaches would be beneficial.

Cancer-specific mortality associated with germline genetic testing results among women with breast cancer or ovarian cancer treated with chemotherapy.

10517 Background: Breast and ovarian cancer patients increasingly undergo germline genetic testing. While studies suggest a greater chemotherapy benefit for carriers of BRCA1/2 pathogenic variants, little is known about whether pathogenic variants in other genes are associated with cancer mortality. Methods: Georgia and California Surveillance, Epidemiology and End Results (SEER) registry records of women diagnosed with breast cancer or ovarian cancer from 2013-2017 were linked to results of clinical germline genetic testing from four participating laboratories. Patients were included if they linked to a genetic result, had stages I-III breast cancer or I-IV epithelial ovarian cancer and received chemotherapy. Multivariable Cox proportional hazard models were used to examine the association of genetic results, demographic and clinical factors with cancer-specific mortality. Results: 21,348 breast and 4,320 ovarian cancer patients were analyzed with median follow-up of 41 months. Pathogenic variants were present in 12% of patients with estrogen and progesterone receptor-positive, HER2-negative breast cancer, 9% with HER2-positive breast cancer, 17% with triple-negative breast cancer and 18% with ovarian cancer. Pathogenic variants were most common in BRCA1/2, CHEK2, PALB2, ATM and BRIP1. Among triple-negative breast cancer patients, mortality was lower with pathogenic variants in BRCA1 (hazard ratio (HR) 0.27, 95% confidence interval (CI) 0.17-0.45) and genes other than BRCA1/2 (HR 0.33, CI 0.13-0.81) versus no pathogenic variant. Genetic results were not associated with mortality in other breast cancer subtypes. Among ovarian cancer patients, mortality was lower with pathogenic variants in BRCA2 (HR 0.36, CI 0.26-0.49) and in genes other than BRCA1/2 (HR 0.48, CI 0.33-0.70). Conclusions: Among breast and ovarian cancer patients treated with chemotherapy, those with germline pathogenic variants in several cancer-associated genes had equivalent or lower short-term mortality than those testing negative. These results may guide patient counseling and clinical trial design.

SOX10 Mutation Screening for 117 Patients with Kallmann Syndrome.

IntroductionKallmann syndrome (KS) is a genetically heterogeneous condition characterized by hypogonadotropic hypogonadism (HH) and olfactory dysfunction. Although SOX10, a causative gene for Waardenburg syndrome (WS) and peripheral demyelinating neuropathy, central demyelination, WS, and Hirschsprung disease (PCWH) has previously been implicated in KS, the clinical significance of SOX10 variants as the cause of KS remains uncertain.Patients and MethodsA total of 117 patients with KS underwent mutation screening of SOX10 and 14 other causative genes for KS/HH. Rare SOX10 variants were subjected to in silico and in vitro analyses. We also examined clinical data of the patients and their parents with SOX10 variants.ResultsSequence analysis identified 2 heterozygous variants of SOX10 (c.1225G > T, p.Gly409* and c.475C > T, p.Arg159Trp) in patients 1–3, as well as in the parents of patients 1 and 3. The variants were assessed as pathogenic/likely pathogenic, according to the American College of Medical Genomics guidelines. Both variants lacked in vitro transactivating activity for the MITF promoter and exerted no dominant-negative effects. Patients 1–3 carried no pathogenic variants in other genes examined. The patients presented with typical KS, while such features were absent in the parents of patients 1 and 3. None of the 5 variant-positive individuals exhibited hypopigmentation, while 1 and 2 individuals exhibited complete and partial hearing loss, respectively.ConclusionThese results provide evidence that SOX10 haploinsufficiency accounts for a small percentage of KS cases. SOX10 haploinsufficiency is likely to be associated with a broad phenotypic spectrum, which includes KS without other clinical features of WS/PCWH.

Analysis of the androgen receptor (AR) gene in a cohort of Indonesian undermasculinized 46, XY DSD patients

BackgroundPathogenic variants in the androgen receptor (AR) gene located on chromosome Xq11-12, are known to cause varying degrees of undermasculinization in 46, XY individuals. The aim of this study was to investigate the frequency of pathogenic variants in the AR gene in a cohort of 46, XY undermasculinized individuals from Indonesia who were suspected of having androgen insensitivity syndrome (AIS). All patients with 46, XY DSD referred to our center between 1994 and 2019 were collected from our clinical database. All 46, XY DSD patients without a prior molecular diagnosis with an external masculinization score (EMS) ≤ 9 were included in this study. All exons and intron–exon boundaries of AR gene were analyzed using Sanger sequencing to identify pathogenic variants of the AR gene.ResultsA cohort of 75 undermasculinized patients were selected for the study. Direct Sanger sequencing of all eight exons of the AR gene led to a genetic diagnosis in 11 patients (14.67%). All of the variants identified (p.Arg841His; p.Ile604Asn; p.Val731Met; p.Pro672Ser; p.Gln739Arg; p.Ser302Glufs*3) have been previously reported in patients with AIS.ConclusionsThis is the first study in Indonesia that highlights the significance of molecular analysis in providing a definitive diagnosis of AIS for patients with 46, XY DSD undermasculinization. This is an uncommon finding in the Indonesian population presenting with 46, XY DSD undermasculinization. A genetic diagnosis allows optimal clinical management and genetic counseling for patients and their families. As 46, XY DSD can be caused by pathogenic variants in other genes involved in gonadal development and differentiation, further genetic analysis, such as whole exome sequencing, should be carried out on those patients that did not carry an AR variant.