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IntroductionThis study aims to define the phenotypic and molecular spectrum of the two clinical forms of β-galactosidase (β-GAL) deficiency, GM1-gangliosidosis and mucopolysaccharidosis IVB (Morquio disease type B, MPSIVB).MethodsClinical and genetic data of 52 probands, 47 patients with GM1-gangliosidosis and 5 patients with MPSIVB were analysed.ResultsThe clinical presentations in patients with GM1-gangliosidosis are consistent with a phenotypic continuum ranging from a severe antenatal form with hydrops fetalis to an adult form with an extrapyramidal syndrome. Molecular studies evidenced 47 variants located throughout the sequence of the GLB1 gene, in all exons except 7, 11 and 12. Eighteen novel variants (15 substitutions and 3 deletions) were identified. Several variants were linked specifically to early-onset GM1-gangliosidosis, late-onset GM1-gangliosidosis or MPSIVB phenotypes. This integrative molecular and clinical stratification suggests a variant-driven patient assignment to a given clinical and severity group.ConclusionThis study reports one of the largest series of b-GAL deficiency with an integrative patient stratification combining molecular and clinical features. This work contributes to expand the community knowledge regarding the molecular and clinical landscapes of b-GAL deficiency for a better patient management.

Background Thymidine phosphorylase (TP), encoded by the TYMP gene, is a cytosolic enzyme essential for the nucleotide salvage pathway. TP catalyzes the phosphorylation of the deoxyribonucleosides, thymidine and 2 '-deoxyuridine, to thymine and uracil. Biallelic TYMP variants are responsible for Mitochondrial NeuroGastroIntestinal Encephalomyopathy (MNGIE), an autosomal recessive disorder characterized in most patients by gastrointestinal and neurological symptoms, ultimately leading to death. Studies on the impact of TYMP variants in cellular systems with relevance to the organs affected in MNGIE are still scarce and the role of TP in adipose tissue remains unexplored. Methods Deep phenotyping was performed in three patients from two families carrying homozygous TYMP variants and presenting with lipoatrophic diabetes. The impact of the loss of TP expression was evaluated using a CRISPR-Cas9-mediated TP knockout (KO) strategy in human adipose stem cells (ASC), which can be differentiated into adipocytes in vitro. Protein expression profiles and cellular characteristics were investigated in this KO model. Results All patients had TYMP loss-of-function variants and first presented with generalized loss of adipose tissue and insulin-resistant diabetes. CRISPR-Cas9-mediated TP KO in ASC abolished adipocyte differentiation and decreased insulin response, consistent with the patients' phenotype. This KO also induced major oxidative stress, altered mitochondrial functions, and promoted cellular senescence. This translational study identifies a new role of TP by demonstrating its key regulatory functions in adipose tissue. Conclusions The implication of TP variants in atypical forms of monogenic diabetes shows that genetic diagnosis of lipodystrophic syndromes should include TYMP analysis. The fact that TP is crucial for adipocyte differentiation and function through the control of mitochondrial homeostasis highlights the importance of mitochondria in adipose tissue biology., Mairie de Paris grant [R18139DD]; Societe Francophone du Diabete [R19114DD]; Fondation pour la Recherche Medicale [ARF20170938613, EQU202003010517]; Fondation pour la Recherche Medicale grant [EQU201903007868]; Agence nationale de la recherche grant [ANR-21-CE17-0002-01], Mairie de Paris grant R18139DD (JG) Societe Francophone du Diabete grant R19114DD (JG) Fondation pour la Recherche Medicale grants ARF20170938613 & EQU202003010517 (JG) Agence nationale de la recherche grant ANR-21-CE17-0002-01 (JG) Fondation pour la Recherche Medicale grant EQU201903007868 (IJ, CV, BF)

Recently a new three-group clinical classification was reported by an International Consortium to stratify CMML patients with regard to prognosis. The groups were defined as follows: (1) Myelodysplastic (MD)-CMML: WBC ≤ 10 × 109/l, circulating immature myeloid cells (IMC) = 0, no splenomegaly; (2) MD/MP (overlap)–CMML: WBC 10–20 × 109/l or WBC ≤ 10 × 109/l but IMC > 0 and/or splenomegaly; (3) Myeloproliferative (MP)-CMML: WBC > 20 × 109/l. By analysing EBMT Registry patients who underwent allo-HCT for CMML between 1997 and 2016, we aimed to determine the impact of this classification on transplantation outcome and to make a comparison with the conventional WHO classification (CMML-0/CMML-1/CMML-2). Patient grouping was based on the data registered at time of transplantation, with IMC replaced by peripheral blasts. Among 151 patients included in the analysis, 38% were classified as MD-CMML, 42% as MD/MP-CMML and 20% as MP-CMML. With a median survival of 17 months in the whole series, MD-CMML patients were distinguished as a low-risk group with higher CR rate at transplant and a longer post-transplant 2-year progression-free survival in comparison to others (44.5% vs 33.5%, respectively), whereas the WHO classification was superior in identifying high-risk patients (CMML-2) with inferior survival outcomes.

Acute myeloid leukemia (AML) is a hematological malignancy with an undefined heritable risk. Here we perform a meta-analysis of three genome-wide association studies, with replication in a fourth study, incorporating a total of 4018 AML cases and 10488 controls. We identify a genome-wide significant risk locus for AML at 11q13.2 (rs4930561; P = 2.15 × 10−8; KMT5B). We also identify a genome-wide significant risk locus for the cytogenetically normal AML sub-group (N = 1287) at 6p21.32 (rs3916765; P = 1.51 × 10−10; HLA). Our results inform on AML etiology and identify putative functional genes operating in histone methylation (KMT5B) and immune function (HLA)., Genome wide association studies in cancer are used to understand the heritable genetic contribution to disease risk. Here, the authors perform a genome wide association study in European patients with acute myeloid leukemia and identify loci associated with risk of developing the disease.

Objectives To determine the prevalence of Melanocortin-4 Receptor (MC4R) mutations in a cohort of children and adolescents with overweight or obesity and to determine whether treatment responses differed between carriers and noncarriers. Methods Using target region capture sequencing, an MC4R mutation screen was performed in 1261 Danish children and adolescents enrolled at a tertiary multidisciplinary childhood obesity treatment center. Measurements of anthropometrics, blood pressure, fasting blood biochemistry including lipid and hormone levels, and dual-energy X-ray absorptiometry were performed at baseline and throughout treatment. Results Of 1209 children and adolescents that met all criteria to be included in the described analyses, 30 (2.5%) carried damaging or unresolved MC4R mutations. At baseline, mutation carriers exhibited higher concentrations of plasma thyroid-stimulating hormone (p = 0.003), and lower concentrations of plasma thyroxine (p = 0.010) compared to noncarriers. After a median of 1 year of treatment (range 0.5–4.0 years), body mass index (BMI) standard deviation score (SDS) was reduced in noncarriers but not in carriers, and this difference in treatment response was statistically significant (p = 0.005). Furthermore, HDL cholesterol was reduced in carriers, a response significantly different from that of noncarriers (p = 0.017). Conclusion Among Danish children and adolescents with overweight or obesity entering a tertiary lifestyle intervention, 2.5% carried damaging or unresolved MC4R mutations. In contrast to noncarriers, carriers of damaging or unresolved MC4R mutations failed to reduce their BMI SDS during obesity treatment, indicating a need for personalized treatment based on the MC4R genotype.

Thrombotic microangiopathy (TMA) is a rare and potentially life-threatening condition that can be caused by a heterogeneous group of diseases, often affecting the brain and kidneys. TMAs should be classified according to etiology to indicate targets for treatment. Complement dysregulation is an important cause of TMA that defines cases not related to coexisting conditions, that is, primary atypical hemolytic uremic syndrome (HUS). Ever since the approval of therapeutic complement inhibition, the approach of TMA has focused on the recognition of primary atypical HUS. Recent advances, however, demonstrated the pivotal role of complement dysregulation in specific subtypes of patients considered to have secondary atypical HUS. This is particularly the case in patients presenting with coexisting hypertensive emergency, pregnancy, and kidney transplantation, shifting the paradigm of disease. In contrast, complement dysregulation is uncommon in patients with other coexisting conditions, such as bacterial infection, drug use, cancer, and autoimmunity, among other disorders. In this review, we performed a critical appraisal on complement dysregulation and the use of therapeutic complement inhibition in TMAs associated with coexisting conditions and outline a pragmatic approach to diagnosis and treatment. For future studies, we advocate the term complement-mediated TMA as opposed to the traditional atypical HUS-type classification.

Acute myeloid leukemia (AML) is a hematological malignancy with an undefined heritable risk. Here we perform a meta-analysis of three genome-wide association studies, with replication in a fourth study, incorporating a total of 4018 AML cases and 10488 controls. We identify a genome-wide significant risk locus for AML at 11q13.2 (rs4930561; P = 2.15 × 10 -8 ; KMT5B). We also identify a genome-wide significant risk locus for the cytogenetically normal AML sub-group (N = 1287) at 6p21.32 (rs3916765; P = 1.51 × 10 -10 ; HLA). Our results inform on AML etiology and identify putative functional genes operating in histone methylation (KMT5B) and immune function (HLA).

Objective The molecular basis of prostate cancer is highly heterogeneous. Our study aimed to perform the mutation analysis of KRAS, BRAF, PIK3CA, and immunohistochemical (IHC) evaluation of EGFR, HER2, p16, and PTEN to demonstrate new areas for targeted therapies. Methods A total of 24 prostatectomy samples diagnosed with adenocarcinoma were analyzed by microarray hybridization. Also, these samples were IHC stained for EGFR, HER2, P16, and PTEN. The cases were divided into two groups based on low and high Gleason scores. All findings were compared with the clinicopathological parameters of the patients. Results While KRAS mutation was in 3/24 (12.5%) of our cases, BRAF and PIK3CA mutations were not detected. There was no significant difference between the groups in terms of KRAS mutation frequency. HER2 was immunohistochemically negative in all samples. There was no correlation between EGFR, P16 immunopositivity, and clinicopathological features. Conclusion KRAS mutation frequency is similar to those in Asian populations. BRAF and PIK3CA mutation frequencies have been reported in the literature in the range of 0-15% and 0-10.4%, respectively, consistent with our study findings. HER2 immunoexpression is a controversial issue in the literature. EGFR and p16 expressions may not correlate with the stage. WOS:000641856000186 2-s2.0-85104773618 PubMed: 33878842

Integrating somatic mutation analysis and gene expression profiling distinguishes pediatric AML subtypes with differential prognoses and clinical risks., Genomic characterization of pediatric patients with acute myeloid leukemia (AML) has led to the discovery of somatic mutations with prognostic implications. Although gene-expression profiling can differentiate subsets of pediatric AML, its clinical utility in risk stratification remains limited. Here, we evaluate gene expression, pathogenic somatic mutations, and outcome in a cohort of 435 pediatric patients with a spectrum of pediatric myeloid-related acute leukemias for biological subtype discovery. This analysis revealed 63 patients with varying immunophenotypes that span a T-lineage and myeloid continuum designated as acute myeloid/T-lymphoblastic leukemia (AMTL). Within AMTL, two patient subgroups distinguished by FLT3-ITD and PRC2 mutations have different outcomes, demonstrating the impact of mutational composition on survival. Across the cohort, variability in outcomes of patients within isomutational subsets is influenced by transcriptional identity and the presence of a stem cell–like gene-expression signature. Integration of gene expression and somatic mutations leads to improved risk stratification. Significance: Immunophenotype and somatic mutations play a significant role in treatment approach and risk stratification of acute leukemia. We conducted an integrated genomic analysis of pediatric myeloid malignancies and found that a combination of genetic and transcriptional readouts was superior to immunophenotype and genomic mutations in identifying biological subtypes and predicting outcomes. This article is highlighted in the In This Issue feature, p. 549

Olgun, Gulden/0000-0002-4467-1610; Sahin, Erdi/0000-0002-5792-2888; Tastan, Oznur/0000-0001-7058-5372; Akcimen, Fulya/0000-0003-0931-5247; Kartal, Ece/0000-0002-7720-455X WOS:000542467300001 PubMed ID: 32579787 The last decade has proven that amyotrophic lateral sclerosis (ALS) is clinically and genetically heterogeneous, and that the genetic component in sporadic cases might be stronger than expected. This study investigates 1,200 patients to revisit ALS in the ethnically heterogeneous yet inbred Turkish population. Familial ALS (fALS) accounts for 20% of our cases. The rates of consanguinity are 30% in fALS and 23% in sporadic ALS (sALS). Major ALS genes explained the disease cause in only 35% of fALS, as compared with similar to 70% in Europe and North America. Whole exome sequencing resulted in a discovery rate of 42% (53/127). Whole genome analyses in 623 sALS cases and 142 population controls, sequenced within Project MinE, revealed well-established fALS gene variants, solidifying the concept of incomplete penetrance in ALS. Genome-wide association studies (GWAS) with whole genome sequencing data did not indicate a new risk locus. Coupling GWAS with a coexpression network of disease-associated candidates, points to a significant enrichment for cell cycle- and division-related genes. Within this network, literature text-mining highlightsDECR1, ATL1, HDAC2, GEMIN4, andHNRNPA3as important genes. Finally, information on ALS-related gene variants in the Turkish cohort sequenced within Project MinE was compiled in the GeNDAL variant browser (www.gendal.org). TUBITAKTurkiye Bilimsel ve Teknolojik Arastirma Kurumu (TUBITAK) [109S075]; Bogazici University Research FundsBogazici University [15B01P1]; Suna and Inan Kirac Foundation [2005-2020] TUBITAK, Grant/Award Number: 109S075; Bogazici University Research Funds, Grant/Award Number: 15B01P1; Suna and Inan Kirac Foundation, Grant/Award Number: 2005-2020

Background: EGFR-tyrosine kinase inhibitors (EGFR-TKIs) including afatinib, dacomitinib, erlotinib, gefitinib, and osimertinib have proven efficacy in terms of progression-free survival (PFS) in patients with non-small-cell lung cancer (NSCLC) harboring EGFR mutations. However, an overall view for comparing efficacy and toxicity on a meta-level is lacking. This study compared efficacy and toxicity of first-line treatment with five different EGFR-TKIs by conducting a network meta-analysis (NMA).Methods: A systematic review was performed, aiming to find eligible literature. Data of PFS, overall survival (OS), objective response rate (ORR), and adverse events were extracted. An NMA based on Bayesian statistics was established to synthesize the efficacy and toxicity of all treatments.Results: Thirteen randomized controlled trials, including data from 3,539 patients with EGFR-mutated NSCLC, were analyzed. Rank probabilities showed that osimertinib had a potentially better efficacy in terms of PFS and OS compared to all other TKIs. For ORR, afatinib and osimertinib showed a trend of superiority compared to the other four TKIs. Furthermore, there was a high risk of diarrhea and rash for patients treated with afatinib or dacomitinib as well as a moderate risk for treatment with erlotinib, gefitinib, and osimertinib.Conclusion: Our study showed a favorable efficacy of osimertinib in terms of PFS and OS compared to all other EGFR-TKIs in patients with NSCLC harboring activating EGFR mutations. Furthermore, gefitinib, erlotinib, and osimertinib were associated with fewer toxicities compared to the other TKIs. Therefore, osimertinib is indicated as a preferable first-line TKI in patients with activating EGFR-mutated NSCLC.

WOS: 000463722100007, PubMed ID: 30489254, Objective: Familial Mediterranean fever (FMF) is the most common autoinflammatory disease. Most of the identified disease-causing mutations are located on exon 10. As the number of studies about the effect of the exonal location of the mutation and its phenotypic expression is limited, we aimed to investigate whether the exonic location of the Mediterranean fever (MEFV) mutation has an effect on the clinical manifestation in patients with FMF. Methods: Study population was derived from the main FMF registry that included 2246 patients from 15 different rheumatology clinics. We categorized the mutations according to their exon locations and retrieved the clinical and demographic information from the database. Results: Patients having the MEFV mutations on exon 2 or 10 (n: 1526) were divided into three subgroups according to the location of the MEFV mutations: Group 1 (exon 2 mutations), Group 2 (exon 10 mutations), and Group 3 (both exon 2 and exon 10 mutations). Group 2 patients were of a significantly younger age at onset, and erysipel-like erythema, arthritis, amyloidosis, and a family history of FMF were more common in this group. Conclusion: Patients with FMF and exon 10 mutations show more severe clinical symptoms and outcome. Exon 2 mutations tend to have a better outcome.

Background/Aims: Acute myeloid leukemia (AML) of French-American-British (FAB) subtypes M0 and M1 are both poorly differentiated AML, but their mutational spectrum and molecular characteristics remain unknown. This study aimed to explore the mutational spectrum and prognostic factors of AML-M0 and M1. Methods: Sixty-five AML patients derived from The Cancer Genome Atlas (TCGA) database were enrolled in this study. Whole-genome sequencing was performed to depict the mutational spectrum of each patient. Clinical characteristics at diagnosis, including peripheral blood (PB) white blood cell counts (WBC), blast percentages in PB and bone marrow (BM), FAB subtypes and the frequencies of known recurrent genetic mutations were described. Survival was estimated using the Kaplan-Meier methods and log-rank test. Univariate and multivariate Cox proportional hazard models were constructed for event-free survival (EFS) and overall survival (OS), using a limited backward elimination procedure. Results: Forty-six patients had more than five recurrent genetic mutations. FLT3 had the highest mutation frequency (n=20, 31%), followed by NPM1 (n=18, 28%), DNMT3A (n=16, 25%), IDH1 (n=14, 22%), IDH2 (n=12, 18%), RUNX1 (n=11, 17%) and TET2 (n=7, 11%). Univariate analysis showed that age ≥60 years and TP53 mutations had adverse effect on EFS (P=0.015, P=0.036, respectively) and OS (P=0.003, P=0.004, respectively), WBC count ≥50×109/L and FLT3-ITD negatively affected EFS (P=0.003, P=0.034, respectively), whereas NPM1 mutations had favorable effect on OS (P=0.035) and allogeneic hematopoietic stem cell transplantation (allo-HSCT) on EFS and OS (all P< 0.001). Multivariate analysis suggested that allo-HSCT and NPM1 mutations were independent favorable prognostic factors for EFS and OS (all P< 0.05), WBC count ≥50×109/L was an independent risk factor for EFS (P=0.002) and TP53 mutations for OS (P=0.043). Conclusions: Our study provided new insights into the mutational spectrum and molecular signatures of AML-M0 and M1. We proposed that FLT3-ITD, NPM1 and TP53 be identified as markers for risk stratification of AML-M0 and M1. Patients with AML-M0 and M1 would likely benefit from allo-HSCT.

Thrombotic microangiopathy (TMA) is a pattern of endothelial damage that can be found in association with diverse clinical conditions such as malignant hypertension. Although the pathophysiological mechanisms differ, accumulating evidence links complement dysregulation to various TMA syndromes and in particular the atypical hemolytic uremic syndrome. Here, we evaluated the role of complement in nine consecutive patients with biopsy-proven renal TMA attributed to severe hypertension. Profound hematologic symptoms of TMA were uncommon. In six Out of nine patients, we found mutations C3 in three, CFI in one, CD46 in one, and/or CFH in two patients either with or without the risk CFH-H3 haplotype in four patients. Elevated levels of the soluble C5b-9 and renal deposits of C3c and C5b-9 along the vasculature and/or glomerular capillary wall, confirmed complement activation in vivo. In contrast to patients without genetic defects, patients with complement defects invariably progressed to end-stage renal disease, and disease recurrence after kidney transplantation seems common. Thus, a subset of patients with hypertension-associated TMA falls within the spectrum of complement-mediated TMA, the prognosis of which is poor. Hence, testing for genetic complement abnormalities is warranted in patients with severe hypertension and TMA on renal biopsy to adopt suitable treatment options and prophylactic measures.

Background: Sodium intake, but not potassium or fluid intake, has been associated with higher renal cell cancer (RCC) risk. However, risk factors may differ by molecular sub-types of the tumour. In renal physiology, electrolyte and water homeostasis is facilitated by ion transport mechanisms (ITM). Aberrant regulation of ITM genes, for example by promoter CpG island methylation, may modify associations between sodium, potassium and fluid intake and RCC risk.Methods: We identified ARHGDIG, ATP1A1, SCNN1B and SLC8A3 as ITM genes exhibiting RCC-specific promoter methylation and down-regulation. Methylation-specific polymerase chain reaction (PCR) was used to analyse promoter CpG island methylation in tumour DNA of 453 RCC cases from the Netherlands Cohort Study (n = 120 852) after 20.3 years of follow-up. Diet was measured at baseline using food-frequency questionnaires. Cox regression analyses were restricted to clear-cell (cc) RCC (n = 306) and stratified by tumours with no, low (1 gene) and high (>= 2 genes) methylation.Results: Sodium intake (high vs low) increased ccRCC risk particularly in tumours with a high methylation index: hazard ratio (HR) [95% confidence interval (CI)]: 2.04 (1.16-3.58), whereas heterogeneity across the methylation index was not significant (P-heterogeneity = 0.26). Potassium intake was differentially associated with ccRCC risk (P-heterogeneity = 0.008); the risk for high (vs low) potassium intake was low for unmethylated tumours [HR (95% CI): 0.60 (0.36-1.01)], but high for tumours with a high methylation index [HR (95% CI): 1.60 (0.96-2.65)]. Risks similarly differed for fluid intake, though not significantly (P-heterogeneity = 0.54).Conclusions: Our findings suggest for the first time that dietary intakes are differentially associated with ccRCC risk according to molecular subtypes defined by ITM gene-specific promoter methylation.

Exome sequencing in diabetes presents a diagnostic challenge because depending on frequency, functional impact, and genomic and environmental contexts, HNF1A variants can cause maturity-onset diabetes of the young (MODY), increase type 2 diabetes risk, or be benign. A correct diagnosis matters as it informs on treatment, progression, and family risk. We describe a multi-dimensional functional dataset of 73 HNF1A missense variants identified in exomes of 12,940 individuals. Our aim was to develop an analytical framework for stratifying variants along the HNF1A phenotypic continuum to facilitate diagnostic interpretation. HNF1A variant function was determined by four different molecular assays. Structure of the multi-dimensional dataset was explored using principal component analysis, k-means, and hierarchical clustering. Weights for tissue-specific isoform expression and functional domain were integrated. Functionally annotated variant subgroups were used to re-evaluate genetic diagnoses in national MODY diagnostic registries. HNF1A variants demonstrated a range of behaviors across the assays. The structure of the multi-parametric data was shaped primarily by transactivation. Using unsupervised learning methods, we obtained high-resolution functional clusters of the variants that separated known causal MODY variants from benign and type 2 diabetes risk variants and led to reclassification of 4% and 9% of HNF1A variants identified in the UK and Norway MODY diagnostic registries, respectively. Our proof-of-principle analyses facilitated informative stratification of HNF1A variants along the continuum, allowing improved evaluation of clinical significance, management, and precision medicine in diabetes clinics. Transcriptional activity appears a superior readout supporting pursuit of transactivation-centric experimental designs for high-throughput functional screens. publishedVersion

OBJECTIVE This study aims to evaluate the predictive impacts of cigarette smoking on treatment outcomes of EGFR tyrosine kinase inhibitors (TKIs) in Non-Small Cell Lung Cancer (NSCLC) patients with activating EGFR mutations. METHODS We retrospectively evaluated the data of 46 patients with metastatic NSCLC (adenocarcinoma) and EGFR mutation (exon 19 deletion, exon 21 mutation, and exon 18 activating mutation) treated with EGFR-TKI between 2012 and 2017. RESULTS Median age was 61 (range 30-80), and 56.5% (26/46) was female. Median follow-up was 39 months. The rate of smoking was 41.3% (19/46). The EGFR mutations were present in the patients, exon 19 deletion in 29 patients (64%), exon 21 mutation in 13 patients (28%) and exon 18 activating mutations in four patients (8%). Progression-free survival (PFS) was 21 months in smokers, whereas it was 25 months in non-smokers (p=0.330). Median PFS was 21 months for patients using EGFR-TKI in the first-line (35 patients), and 13 months in the second-line setting (11 patients). CONCLUSION There were no statistically significant PFS differences between the smoker and non-smoker groups. Smokers should be tested for EGFR mutations, as some patients may benefit from EGFR TKI treatment for longer than reported in the literature.

Objective: The aim was to systematically review the characteristics of ovarian cancer in women with Lynch syndrome (LS) and evaluate the role of surveillance in detection of ovarian cancer in LS. Methods: All studies between 1979 and 2015 of women with ovarian cancer and LS or at 50% risk of LS were evaluated. Two reviewers independently evaluated eligible studies and extracted data on age at diagnosis, histological type, FIGO stage, and way of detection according to pre-specified criteria. The studies were assessed for quality using the Newcastle-Ottawa quality assessment scales. Results: The quality score of the 49 identified studies was at least 6 out of 8 and provide clinical information on 747 LS women with ovarian cancer. The mean age at diagnosis was 45.3 (range 19-82) years. Most frequent mutations were MSH2 (47%) and MLH1 (38%). Histopathological data were available for 445 women. The most frequently reported histological type was mixed type (mucinous/endometrioid/clear cell carcinomas) (n = 136; 31%). Most tumours (281, 65%) were diagnosed at an early stage (FIGO I/II). Six studies evaluating the effect of surveillance of ovarian cancer, reported that seven of 22 (32%) ovarian cancers were found during surveillance, 6/7 (86%) were detected at an early stage. Conclusion: This systematic review describes that ovarian cancer in women with LS has a wide age-range of onset, is often diagnosed at an early stage with frequently endometrioid/clear cell histology. Data about the role of surveillance in detection of ovarian cancer in women with LS are scarce however detection at an early stage seems possible. (C) 2015 Elsevier Ltd. All rights reserved.

Objective: The aim was to systematically review the characteristics of ovarian cancer in women with Lynch syndrome (LS) and evaluate the role of surveillance in detection of ovarian cancer in LS.Methods: All studies between 1979 and 2015 of women with ovarian cancer and LS or at 50% risk of LS were evaluated. Two reviewers independently evaluated eligible studies and extracted data on age at diagnosis, histological type, FIGO stage, and way of detection according to pre-specified criteria. The studies were assessed for quality using the Newcastle-Ottawa quality assessment scales.Results: The quality score of the 49 identified studies was at least 6 out of 8 and provide clinical information on 747 LS women with ovarian cancer. The mean age at diagnosis was 45.3 (range 19-82) years. Most frequent mutations were MSH2 (47%) and MLH1 (38%). Histopathological data were available for 445 women. The most frequently reported histological type was mixed type (mucinous/endometrioid/clear cell carcinomas) (n = 136; 31%). Most tumours (281, 65%) were diagnosed at an early stage (FIGO I/II). Six studies evaluating the effect of surveillance of ovarian cancer, reported that seven of 22 (32%) ovarian cancers were found during surveillance, 6/7 (86%) were detected at an early stage.Conclusion: This systematic review describes that ovarian cancer in women with LS has a wide age-range of onset, is often diagnosed at an early stage with frequently endometrioid/clear cell histology. Data about the role of surveillance in detection of ovarian cancer in women with LS are scarce however detection at an early stage seems possible. (C) 2015 Elsevier Ltd. All rights reserved.

Pheochromocytomas (PCC) and abdominal paragangliomas (PGL) display a highly diverse genetic background and recent gene expression profiling studies have shown that PCC and PGL (together PPGL) alter either kinase signaling pathways or the pseudo-hypoxia response pathway dependent of the genetic composition. Recurrent mutations in the Harvey rat sarcoma viral oncogene homolog (HRAS) have recently been verified in sporadic PPGLs. In order to further establish the HRAS mutation frequency and to characterize the associated expression profiles of HRAS mutated tumors, 156 PPGLs for exon 2 and 3 hotspot mutations in the HRAS gene was screened, and compared with microarray-based gene expression profiles for 93 of the cases. The activating HRAS mutations G13R, Q61R, and Q61K were found in 10/142 PCC (7.0%) and a Q61L mutation was revealed in 1/14 PGL (7.1%). All HRAS mutated cases included in the mRNA expression profiling grouped in Cluster 2, and 21 transcripts were identified as altered when comparing the mutated tumors with 91 HRAS wild-type PPGL. Somatic HRAS mutations were not revealed in cases with known PPGL susceptibility gene mutations and all HRAS mutated cases were benign. The HRAS mutation prevalence of all PPGL published up to date is 5.2% (49/950), and 8.8% (48/548) among cases without a known PPGL susceptibility gene mutation. The findings support a role of HRAS mutations as a somatic driver event in benign PPGL without other known susceptibility gene mutations. HRAS mutated PPGL cluster together with NF1- and RET-mutated tumors associated with activation of kinase-signaling pathways. Funding Agencies|Swedish Cancer Foundation; StratCan; Swedish Research Council; Cancer Research Foundations of Radiumhemmet; Karolinska Institutet; Stockholm County Council

Author(s): Ustun, Celalettin; Morgan, Elizabeth; Moodie, Erica EM; Pullarkat, Sheeja; Yeung, Cecilia; Broesby-Olsen, Sigurd; Ohgami, Robert; Kim, Young; Sperr, Wolfgang; Vestergaard, Hanne; Chen, Dong; Kluin, Philip M; Dolan, Michelle; Mrozek, Krzysztof; Czuchlewski, David; Horny, Hans-Peter; George, Tracy I; Kristensen, Thomas Kielsgaard; Ku, Nam K; Yi, Cecilia Arana; Moller, Michael Boe; Marcucci, Guido; Baughn, Linda; Schiefer, Ana-Iris; Hilberink, JR; Pullarkat, Vinod; Shanley, Ryan; Kohlschmidt, Jessica; Coulombe, Janie; Salhotra, Amandeep; Soma, Lori; Cho, Christina; Linden, Michael A; Akin, Cem; Gotlib, Jason; Hoermann, Gregor; Hornick, Jason; Nakamura, Ryo; Deeg, Joachim; Bloomfield, Clara D; Weisdorf, Daniel; Litzow, Mark R; Valent, Peter; Huls, Gerwin; Perales, Miguel-Angel; Borthakur, Gautam | Abstract: BackgroundAlthough the prognosis of core-binding factor (CBF) acute myeloid leukemia (AML) is better than other subtypes of AML, 30% of patients still relapse and may require allogeneic hematopoietic cell transplantation (alloHCT). However, there is no validated widely accepted scoring system to predict patient subsets with higher risk of relapse.MethodsEleven centers in the US and Europe evaluated 247 patients with t(8;21)(q22;q22).ResultsComplete remission (CR) rate was high (92.7%), yet relapse occurred in 27.1% of patients. A total of 24.7% of patients received alloHCT. The median disease-free (DFS) and overall (OS) survival were 20.8 and 31.2 months, respectively. Age, KIT D816V mutated (11.3%) or nontested (36.4%) compared with KIT D816V wild type (52.5%), high white blood cell counts (WBC), and pseudodiploidy compared with hyper- or hypodiploidy were included in a scoring system (named I-CBFit). DFS rate at 2 years was 76% for patients with a low-risk I-CBFit score compared with 36% for those with a high-risk I-CBFit score (P l 0.0001). Low- vs high-risk OS at 2 years was 89% vs 51% (P l 0.0001).ConclusionsI-CBFit composed of readily available risk factors can be useful to tailor the therapy of patients, especially for whom alloHCT is not need in CR1 (ie, patients with a low-risk I-CBFit score).

A high proportion of patients with acute myeloid leukemia who achieve minimal residual disease negative status ultimately relapse because a fraction of pathological clones remains undetected by standard methods. We designed and validated a high-throughput sequencing method for minimal residual disease assessment of cell clonotypes with mutations of NPM1, IDH1/2 and/or FLT3-single nucleotide variants. For clinical validation, 106 follow-up samples from 63 patients in complete remission were studied by sequencing, evaluating the level of mutations detected at diagnosis. The predictive value of minimal residual disease status by sequencing, multiparameter flow cytometry, or quantitative polymerase chain reaction analysis was determined by survival analysis. The sequencing method achieved a sensitivity of 10 -4 for single nucleotide variants and 10 -5 for insertions/deletions and could be used in acute myeloid leukemia patients who carry any mutation (86% in our diagnostic data set). Sequencing-determined minimal residual disease positive status was associated with lower disease-free survival (hazard ratio 3.4, P=0.005) and lower overall survival (hazard ratio 4.2, P, This study was supported by the Subdirección General de Investigación Sanitaria (Instituto de Salud Carlos III, Spain) grants PI13/02387 and PI16/01530, and the CRIS against Cancer foundation grant 2014/0120. ML holds a postdoctoral fellowship of the Spanish Ministry of Economy and Competitiveness (FPDI-2013- 16409). PRP holds a postdoctoral fellowship of the Spanish Instituto de Salud Carlos III: Contrato Predoctoral de Formación en Investigación en Salud i-PFIS (IFI 14/00008).

tThe U12-dependent (minor) spliceosome excises a rare group of introns that are characterized by ahighly conserved 5splice site and branch point sequence. Several new congenital or somatic diseaseshave recently been associated with mutations in components of the minor spliceosome. A commontheme in these diseases is the detection of elevated levels of transcripts containing U12-type introns,of which a subset is associated with other splicing defects. Here we review the present understandingof minor spliceosome diseases, particularly those associated with the specific components of the minorspliceosome. We also present a model for interpreting the molecular-level consequences of the differentdiseases.

Over fifty missense mutations in the gene coding for valosin-containing protein (VCP) are associated with a unique autosomal dominant adult-onset progressive disease associated with combinations of proximo-distal inclusion body myopathy (IBM), Paget's disease of bone (PDB), frontotemporal dementia (FTD), and amyotrophic lateral sclerosis (ALS). We report the clinical, histological, and molecular findings in four new patients/families carrying novel VCP mutations: c.474 G > A (p.M158I); c.478 G > C (p.A160P); c.383G > C (p.G128A); and c.382G > T (p.G128C). Clinical features included myopathy, PDB, ALS and Parkinson's disease though frontotemporal dementia was not an associated feature in these families. One of the patients was noted to have severe manifestations of PDB and was suspected of having neoplasia. There were wide inter- and intra-familial variations making genotype-phenotype correlations difficult between the novel mutations and frequency or age of onset of IBM, PDB, FTD, ALS and Parkinson's disease. Increasing awareness of the full spectrum of clinical presentations will improve diagnosis of VCP-related diseases and thus proactively manage or prevent associated clinical features such as PDB. (C) 2018 Published by Elsevier B.V.

Purpose Recommendations for treating patients who carry a BRCA1/2 gene are mainly based on cumulative lifetime risks (CLTRs) of breast cancer determined from retrospective cohorts. These risks vary widely (27% to 88%), and it is important to understand why. We analyzed the effects of methods of risk estimation and bias correction and of population factors on CLTRs in this retrospective clinical cohort of BRCA1/2 carriers. Patients and Methods The following methods to estimate the breast cancer risk of BRCA1/2 carriers were identified from the literature: Kaplan-Meier, frailty, and modified segregation analyses with bias correction consisting of including or excluding index patients combined with including or excluding first-degree relatives (FDRs) or different conditional likelihoods. These were applied to clinical data of BRCA1/2 families derived from our family cancer clinic for whom a simulation was also performed to evaluate the methods. CLTRs and 95% CIs were estimated and compared with the reference CLTRs. Results CLTRs ranged from 35% to 83% for BRCA1 and 41% to 86% for BRCA2 carriers at age 70 years width of 95% CIs: 10% to 35% and 13% to 46%, respectively). Relative bias varied from −38% to +16%. Bias correction with inclusion of index patients and untested FDRs gave the smallest bias: +2% (SD, 2%) in BRCA1 and +0.9% (SD, 3.6%) in BRCA2. Conclusion Much of the variation in breast cancer CLTRs in retrospective clinical BRCA1/2 cohorts is due to the bias-correction method, whereas a smaller part is due to population differences. Kaplan-Meier analyses with bias correction that includes index patients and a proportion of untested FDRs provide suitable CLTRs for carriers counseled in the clinic.

PurposeRecommendations for treating patients who carry a BRCA1/2 gene are mainly based on cumulative lifetime risks (CLTRs) of breast cancer determined from retrospective cohorts. These risks vary widely (27% to 88%), and it is important to understand why. We analyzed the effects of methods of risk estimation and bias correction and of population factors on CLTRs in this retrospective clinical cohort of BRCA1/2 carriers.Patients and MethodsThe following methods to estimate the breast cancer risk of BRCA1/2 carriers were identified from the literature: Kaplan-Meier, frailty, and modified segregation analyses with bias correction consisting of including or excluding index patients combined with including or excluding first-degree relatives (FDRs) or different conditional likelihoods. These were applied to clinical data of BRCA1/2 families derived from our family cancer clinic for whom a simulation was also performed to evaluate the methods. CLTRs and 95% CIs were estimated and compared with the reference CLTRs.ResultsCLTRs ranged from 35% to 83% for BRCA1 and 41% to 86% for BRCA2 carriers at age 70 years width of 95% CIs: 10% to 35% and 13% to 46%, respectively). Relative bias varied from -38% to +16%. Bias correction with inclusion of index patients and untested FDRs gave the smallest bias: +2% (SD, 2%) in BRCA1 and +0.9% (SD, 3.6%) in BRCA2.ConclusionMuch of the variation in breast cancer CLTRs in retrospective clinical BRCA1/2 cohorts is due to the bias-correction method, whereas a smaller part is due to population differences. Kaplan-Meier analyses with bias correction that includes index patients and a proportion of untested FDRs provide suitable CLTRs for carriers counseled in the clinic. (C) 2015 by American Society of Clinical Oncology

Familial hypertrophic cardiomyopathy (HCM) is usually caused by autosomal dominant pathogenic mutations in genes encoding sarcomeric or sarcomere-associated cardiac muscle proteins. The disease mainly affects adults, although young children with severe HCM have also been reported. We describe four unrelated neonates with lethal cardiomyopathy, and performed molecular studies to identify the genetic defect. We also present a literature overview of reported patients with compound heterozygous or homozygous pathogenic MYBPC3 mutations and describe their clinical characteristics. All four children presented with feeding difficulties, failure to thrive, and dyspnea. They died from cardiac failure before age 13 weeks. Features of left ventricular noncompaction were diagnosed in three patients. In the fourth, hypertrabeculation was not a clear feature, but could not be excluded. All of them had septal defects. Two patients were compound heterozygotes for the pathogenic c.2373dup p.(Trp792fs) and c.2827C > T p.(Arg943*) mutations, and two were homozygous for the c.2373dup and c.2827C > T mutations. All patients with biallelic truncating pathogenic mutations in MYBPC3 reported so far (n=21) were diagnosed with severe cardiomyopathy and/or died within the first few months of life. In 62% (13/21), septal defects or a patent ductus arteriosus accompanied cardiomyopathy. In contrast to heterozygous pathogenic mutations, homozygous or compound heterozygous truncating pathogenic MYBPC3 mutations cause severe neonatal cardiomyopathy with features of left ventricular noncompaction and septal defects in approximately 60% of patients.

ATP8B1 deficiency is a severe autosomal recessive liver disease resulting from mutations in the ATP8B1 gene characterized by a continuous phenotypical spectrum from intermittent (benign recurrent intrahepatic cholestasis; BRIC) to progressive familial intrahepatic cholestasis (PFIC). Current therapeutic options are insufficient, and elucidating the molecular consequences of mutations could lead to personalized mutation-specific therapies. We investigated the effect on pre-messenger RNA splicing of 14 ATP8B1 mutations at exon-intron boundaries using an in vitro minigene system. Eleven mutations, mostly associated with a PFIC phenotype, resulted in aberrant splicing and a complete absence of correctly spliced product. In contrast, three mutations led to partially correct splicing and were associated with a BRIC phenotype. These findings indicate an inverse correlation between the level of correctly spliced product and disease severity. Expression of modified U1 small nuclear RNAs (snRNA) complementary to the splice donor sites strongly improved or completely rescued splicing for several ATP8B1 mutations located at donor, as well as acceptor, splice sites. In one case, we also evaluated exon-specific U1 snRNAs that, by targeting nonconserved intronic sequences, might reduce possible off-target events. Although very effective in correcting exon skipping, they also induced retention of the short downstream intron. Conclusion: We systematically characterized the molecular consequences of 14 ATP8B1 mutations at exon-intron boundaries associated with ATP8B1 deficiency and found that the majority resulted in total exon skipping. The amount of correctly spliced product inversely correlated with disease severity. Compensatory modified U1 snRNAs, complementary to mutated donor splice sites, were able to improve exon definition very efficiently and could be a novel therapeutic strategy in ATP8B1 deficiency as well as other genetic diseases. (Hepatology 2015;61:1382-1391)

Background— Interpretation of missense variants can be especially difficult when the variant is also found in control populations. This is what we encountered for the LMNA c.992G>A (p.(Arg331Gln)) variant. Therefore, to evaluate the effect of this variant, we combined an evaluation of clinical data with functional experiments and morphological studies. Methods and Results— Clinical data of 23 probands and 35 family members carrying this variant were retrospectively collected. A time-to-event analysis was performed to compare the course of the disease with carriers of other LMNA mutations. Myocardial biopsies were studied with electron microscopy and by measuring force development of the sarcomeres. Morphology of the nuclear envelope was assessed with immunofluorescence on cultured fibroblasts. The phenotype in probands and family members was characterized by atrioventricular conduction disturbances (61% and 44%, respectively), supraventricular arrhythmias (69% and 52%, respectively), and dilated cardiomyopathy (74% and 14%, respectively). LMNA p.(Arg331Gln) carriers had a significantly better outcome regarding the composite end point (malignant ventricular arrhythmias, end-stage heart failure, or death) compared with carriers of other pathogenic LMNA mutations. A shared haplotype of 1 Mb around LMNA suggested a common founder. The combined logarithm of the odds score was 3.46. Force development in membrane-permeabilized cardiomyocytes was reduced because of decreased myofibril density. Structural nuclear LMNA -associated envelope abnormalities, that is, blebs, were confirmed by electron microscopy and immunofluorescence microscopy. Conclusions— Clinical, morphological, functional, haplotype, and segregation data all indicate that LMNA p.(Arg331Gln) is a pathogenic founder mutation with a phenotype reminiscent of other LMNA mutations but with a more benign course.

It is unknown how often cardiac arrhythmias occur in hypokalemic periodic paralysis (HypoPP) and if they are caused by hypokalemia alone or other factors. This systematic review shows that cardiac arrhythmias were reported in 27 HypoPP patients. Cases were confirmed genetically (13 with an R528H mutation in CACNA1S, 1 an R669H mutation in SCN4A) or had a convincing clinical diagnosis of HypoPP (13 genetically undetermined) if reported prior to the availability of genetic testing. Arrhythmias occurred during severe hypokalemia (11 patients), between attacks at normokalemia (4 patients), were treatment-dependent (2 patients), or unspecified (10 patients). Nine patients died from arrhythmia. Convincing evidence for a pro-arrhythmogenic factor other than hypokalemia is still lacking. The role of cardiac expression of defective skeletal muscle channels in the heart of HypoPP patients remains unclear. Clinicians should be aware of and prevent treatment-induced cardiac arrhythmia in HypoPP.

Aims Mutations in the lamin A/C gene (LMNA) cause a variety of clinical phenotypes, including dilated cardiomyopathy. LMNA is one of the most prevalent mutated genes in dilated cardiomyopathy, and is associated with a high risk of arrhythmias, sudden cardiac death, and heart failure. There are few data on the impact of age and gender on cardiac disease penetrance and mortality. Methods and results In a multicentre cohort of 269 LMNA mutation carriers, we evaluated gender-specific penetrance of cardiac involvement and major cardiac events. All-cause mortality of mutation carriers [standardized mortality ratio (SMR)] was determined. Cardiac disease penetrance was age dependent and almost complete at the age of 70 years. The presence of an LVEF ≤45% was significantly higher in men (P < 0.001). However, there was no difference between genders in the prevalence of atrioventricular block, atrial tachyarrhythmias, and non-sustained ventricular tachycardia. Malignant ventricular arrhythmias (26% vs. 8%) and end-stage heart failure (28% vs. 14%) were more common in men than in women (P < 0.001 and P = 0.006, respectively). All-cause mortality of mutation carriers was significantly increased [SMR 4.0, 95% confidence interval (CI) 2.8–5.2] between the ages of 15 and 75 years. Mortality in men was higher than in women (hazard ratio 2.2, 95% CI 1.2–4.3). Conclusions This large cohort of LMNA mutation carriers demonstrates a high cardiac disease penetrance and a high mortality in mutation carriers. Male mutation carriers have a worse prognosis due to a higher prevalence of malignant ventricular arrhythmias and end-stage heart failure.

Background Retinoblastoma (Rb) is a childhood cancer of the retina, commonly initiated by biallelic inactivation of the RB1 gene. Knowledge of the presence of a heritable RB1 mutation can help in risk management and reproductive decision making. We report here on RB1 mutation scanning in a unique nationwide cohort of Rb patients from the Netherlands.Methods From the 1173 Rb patients registered in the Dutch National Retinoblastoma Register until January 2013, 529 patients from 433 unrelated families could be included. RB1 mutation scanning was performed with different detection methods, depending on the time period.Results Our mutation detection methods revealed RB1 mutations in 92% of bilateral and/or familial Rb patients and in 10% of non-familial unilateral cases. Overall an RB1 germline mutation was detected in 187 (43%) of 433 Rb families, including 33 novel mutations. The distribution of the type of mutation was 37% nonsense, 20% frameshift, 21% splice, 9% large indel, 5% missense, 7% chromosomal deletions and 1% promoter. Ten per cent of patients were mosaic for the RB1 mutation. Six three-generation families with incomplete penetrance RB1 mutations were found. We found evidence that two variants, previously described as pathogenic RB1 mutations, are likely to be neutral variants.Conclusions The frequency of the type of mutations in the RB1 gene in our unbiased national cohort is the same as the mutation spectrum described worldwide. Furthermore, our RB1 mutation detection regimen achieves a high scanning sensitivity.

BackgroundLamin A/C (LMNA) mutation carriers suffer from a variety of clinical phenotypes, including dilated cardiomyopathy (DCM). Although it has been suggested that carriers are at risk for thromboembolic complications, it is unknown whether this risk is higher than can be expected from the underlying cardiac abnormalities. The purpose of this study was to determine whether a LMNA mutation is associated with an increased risk of thromboembolic complications.MethodsWe compared a cohort of 76 LMNA mutation carriers with a cohort of 224 idiopathic DCM patients without a LMNA mutation, with respect to the prevalence of arterial and venous thromboembolic complications. Furthermore, we carried out a case–control study to explore whether a prothrombotic phenotype was present in LMNA mutation carriers without DCM or atrial tachyarrhythmias (n=14) and compared this with mutation negative relatives (n=13).ResultsThe prevalence of thromboembolic complications was higher in the cohort of LMNA mutation carriers than in DCM patients (22 vs 11%; p

Background Recently, we showed that the c.40_42delAGA (p.Arg14del) mutation in the phospholamban (PLN) gene can be identified in 10–15 % of Dutch patients with dilated cardiomyopathy or arrhythmogenic cardiomyopathy. The arrhythmogenic burden of the p.Arg14del mutation was illustrated by the high rate of appropriate ICD discharges and a positive family history for sudden cardiac death. Methods Our goal was to evaluate the geographical distribution and the origin of this specific mutation in the Netherlands and to get an estimation of the prevalence in a Dutch population cohort. Therefore, we investigated the postal codes of the places of residence of PLN p.Arg14del mutation carriers and places of birth of their ancestors. In addition, a large population-based cohort (PREVEND) was screened for the presence of this mutation. Results By April 2012, we had identified 101 probands carrying the PLN p.Arg14del mutation. A total of 358 family members were also found to carry this mutation, resulting in a total of 459 mutation carriers. The majority of mutation carriers live in the northern part of the Netherlands and analysing their grandparents’ places of birth indicated that the mutation likely originated in the eastern part of the province of Friesland. In the PREVEND cohort we identified six heterozygous PLN p.Arg14del mutation carriers out of 8,267 subjects (0.07 %). Conclusion The p.Arg14del mutation in the PLN gene is the most frequently identified mutation in Dutch cardiomyopathy patients. The mutation that arose 575–825 years ago is likely to have originated from the eastern part of the province of Friesland and is highly prevalent in the general population in the northern part of the Netherlands.

One of the most important advances in the treatment of non-small cell lung cancer (NSCLC) has been the identification of molecular alterations vulnerable to targeted inhibition, such as mutations in the epidermal growth factor receptor (EGFR) gene. EGFR tyrosine kinase inhibitors (EGFR-TKIs) are targeted agents used to treat EGFR mutation-positive advanced NSCLC showing significant improvements in terms of response rate (RR) and progression-free survival (PFS) compared to conventional chemotherapy. However, all patients eventually develop resistance to first-line EGFR-TKIs. The most common mechanism of acquired resistance is the secondary acquisition of a single missense mutation within exon 20 in the EGFR gene, known as the T790M mutation (49-60%). New agents targeting the T790M mutation have undergone clinical development, and among these, osimertinib has shown significant activity in relapsing EGFR mutation positive patients harbouring the T790M mutation. Although precision medicine is a reality for NSCLC, obtaining relevant tissue for repeated molecular analysis from these patients remains a challenge. In this article, a group of experts from the Spanish Society of Medical Oncology (SEOM) and the Spanish Lung Cancer Group (GECP) evaluated the role of rebiopsy and the potential application of plasma-testing methodologies in advanced EGFR mutation patients progressing after EGFR-TKI.

The hypoxia-inducible factor 1α (HIF-1α) and its microRNA target, miR-210, are candidate tumor-drivers of metabolic reprogramming in cancer. Neuroendocrine neoplasms such as paragangliomas (PGLs) are particularly appealing for understanding the cancer metabolic adjustments because of their associations with deregulations of metabolic enzymes, such as succinate dehydrogenase (SDH), and the von Hippel Lindau (VHL) gene involved in HIF-1α stabilization. However, the role of miR-210 in the pathogenesis of SDH-related tumors remains an unmet challenge. Herein is described an in vivo genetic analysis of the role of VHL, HIF1A and SDH on miR-210 by using knockout murine models, siRNA gene silencing, and analyses of human tumors. HIF-1α knockout abolished hypoxia-induced miR-210 expression in vivo but did not alter its constitutive expression in paraganglia. Normoxic miR-210 levels substantially increased by complete, but not partial, VHL silencing in paraganglia of knockout VHL-mice and by over-expression of p76del-mutated pVHL. Similarly, VHL-mutated PGLs, not those with decreased VHL-gene/mRNA dosage, over-expressed miR-210 and accumulate HIF-1α in most tumor cells. Ablation of SDH activity in SDHD-null cell lines or reduction of the SDHD or SDHB protein levels elicited by siRNA-induced gene silencing did not induce miR-210 whereas the presence of SDH mutations in PGLs and tumor-derived cell lines was associated with mild increase of miR-210 and the presence of a heterogeneous, HIF-1α-positive and HIF-1α-negative, tumor cell population. Thus, activation of HIF-1α is likely an early event in VHL-defective PGLs directly linked to VHL mutations, but it is a late event favored but not directly triggered by SDHx mutations. This combined analysis provides insights into the mechanisms of HIF-1α/miR-210 regulation in normal and tumor tissues potentially useful for understanding the pathogenesis of cancer and other diseases sharing similar underpinnings., This work was supported by Instituto de Salud Carlos III-Fondo de Investigación Sanitaria [FIS PI11/929]; Red Temática de Investigación Cooperativa en Cáncer [RD12/0036/0015] Instituto de Salud Carlos III (ISCIII), Spanish Ministry of Economy and Competitiveness & European Regional Development Fund (ERDF); the European Regional Development's funds (FEDER) (CIBERONC) and Obra Social CajAstur-Instituto Universitario de Oncología del Principado de Asturias.

Germline mutations of the gene encoding succinate dehydrogenase subunit B (SDHB) predispose to head-and-neck-paraganglioma (HNPGL), sympathetic PGL, pheochromocytoma and renal cell carcinoma for which regular surveillance is required. SDHB-associated tumors harbor germline and somatic mutations, consistent with Knudson's two-hit hypothesis. To assess the penetrance and optimal surveillance for different manifestations of SDHB mutation carriers. This study included all SDHB mutation carriers who were followed at the Department of Endocrinology at the University Medical Center of Groningen. Kaplan-Meier curves were used to assess the penetrance. Poisson process was used to assess the optimal age to start surveillance and intervals. Ninety-one SDHB-mutation carriers (38 men and 53 women) were included. Twenty-seven mutation carriers (30 %) had manifestations, with an overall penetrance 35 % at the age of 60 years. We calculated that optimal surveillance for HNPGL could start from an age of 27 years with an interval of 3.2 years. This study underscores the relatively low penetrance of disease in SDHB mutation carriers. Use of the Poisson approach provides a more accurate estimation of the age to initiate surveillance and length of intervals for HNPGL. These results may give rise to reconsider the current guidelines regarding the screening of these mutation carriers.

Background Multiple Endocrine Neoplasia type 1 (MEN1) is diagnosed when two out of the three primary MEN1-associated endocrine tumors occur in a patient. Up to 10–30 % of those patients have no mutation in the MEN1 gene. It is unclear if the phenotype and course of the disease of mutation-negative patients is comparable with mutation-positive patients and if these patients have true MEN1. The present study aims to describe and compare the clinical course of MEN1 mutation-negative patients with two out of the three main MEN1 manifestations and mutation-positive patients during long-term follow-up. Methods This is a cohort study performed using the Dutch MEN1 database, including > 90 % of the Dutch MEN1 population. Results A total of 293 (90.7 %) mutation-positive and 30 (9.3 %) mutation-negative MEN1 patients were included. Median age of developing the first main MEN1 manifestation was higher in mutation-negative patients (46 vs. 33 years) (P = 0.007). Mutation-negative patients did not develop a third main MEN1 manifestation in the course of follow-up compared to 48.3 % of mutation-positive patients (P

Electrocardiogram (ECG) measurements play a key role in the diagnosis and prediction of cardiac arrhythmias and sudden cardiac death. ECG parameters, such as the PR, QRS, and QT intervals, are known to be heritable and genome-wide association studies of these phenotypes have been successful in identifying common variants; however, a large proportion of the genetic variability of these traits remains to be elucidated. The aim of this study was to discover loci potentially harboring rare variants utilizing variance component linkage analysis in 1547 individuals from a large family-based study, the Erasmus Rucphen Family Study (ERF). Linked regions were further explored using exome sequencing. Five suggestive linkage peaks were identified: two for QT interval (1q24, LOD = 2.63; 2q34, LOD = 2.05), one for QRS interval (1p35, LOD = 2.52) and two for PR interval (9p22, LOD = 2.20; 14q11, LOD = 2.29). Fine-mapping using exome sequence data identified a C > G missense variant (c.713C > G, p.Ser238Cys) in the FCRL2 gene associated with QT (rs74608430; P = 2.8 × 10-4, minor allele frequency = 0.019). Heritability analysis demonstrated that the SNP explained 2.42% of the trait's genetic variability in ERF (P = 0.02). Pathway analysis suggested that the gene is involved in cytosolic Ca2+ levels (P = 3.3 × 10-3) and AMPK stimulated fatty acid oxidation in muscle (P = 4.1 × 10-3). Look-ups in bioinformatics resources showed that expression of FCRL2 is associated with ARHGAP24 and SETBP1 expression. This finding was not replicated in the Rotterdam study. Combining the bioinformatics information with the association and linkage analyses, FCRL2 emerges as a strong candidate gene for QT interval. © 2016 Silva, Zorkoltseva, Amin, Demirkan, van Leeuwen, Kors, van den Berg, Stricker, Uitterlinden, Kirichenko, Witteman, Willemsen, Oostra, Axenovich, van Duijn and Isaacs.

Biotinidase deficiency is a rare inherited metabolic disorder that can cause severe neurological symptoms. To prevent severe clinical presentations, it was included in the Dutch neonatal screening programme in 2007. Since then the number of cases detected has been high. This study set out to describe the incidence of the disease, the clinical and demographic characteristics of the neonates identified and the type of mutations found. In the south-western Netherlands, 304 982 neonates were screened between 2007 and 2012; and 92 were identified for further testing. Confirmatory testing revealed 6 (7%) with a profound biotinidase deficiency (o10% enzyme activity), 44 (48%) with a partial deficiency (10-30%) and 42 (46%) with normal activity (> 30%). All six patients whose profound deficiency was confirmed had enzyme activities below 15% on neonatal screening. Mutation analysis was performed in 61 neonates: 5 'profound', 35 'partial' and 21 'normal'. All five 'profound' cases had two severe mutations. Comparison with the northern Netherlands showed that the frequency and types of mutation were representative for the Netherlands as a whole. The most common mutation detected was c.[1330G > C] (p.(Asp444His); 34%), which is considered to be mild, followed by three severe mutations c.[1368A > C], c.[1595C > T] and c.[1330G > C; 511G > A]. Seven new mutations were identified. We conclude that neonatal screening for profound biotinidase produces a high number of false positives. Biotinidase deficiency was profound in less than 10% of cases identified. As biotinidase activity lay below 15% on neonatal screening in all such cases, the screening threshold might be reduced to 15%.

Four main DNA mismatch repair (MMR) genes have been identified, MLH1, MSH2, MSH6, and PMS2, which when mutated cause susceptibility to Lynch syndrome (LS). LS is one of the most prevalent hereditary cancer syndromes in man and accounts for 1-3 % of unselected colorectal carcinomas and some 15 % of those with microsatellite instability and/or absent MMR protein. The International Society for Gastrointestinal Hereditary Tumours (InSiGHT) maintains a database for LS-associated mutations since 1996. The database was recently reorganized to efficiently gather published and unpublished data and to classify the variants according to a five-tiered scheme linked to clinical recommendations. This review provides an update of germline mutations causing susceptibility to LS based on information available in the InSiGHT database and the latest literature. MMR gene mutation profiles, correlations between genotype and phenotype, and possible mechanisms leading to the characteristic spectrum of tumors in LS are discussed in light of the different functions of MMR proteins, many of which directly serve cancer avoidance.

In this paper the Working Group on Myocardial and Pericardial Disease proposes a revised definition of dilated cardiomyopathy (DCM) in an attempt to bridge the gap between our recent understanding of the disease spectrum and its clinical presentation in relatives, which is key for early diagnosis and the institution of potential preventative measures. We also provide practical hints to identify subsets of the DCM syndrome where aetiology directed management has great clinical relevance.

A subset of patients with monogenic disorders lacks disease causing mutations in the protein coding region of the corresponding gene. Here we describe a recurrent germline mutation found in two unrelated patients with complete androgen insensitivity syndrome (CAIS) generating an upstream open reading frame (uORF) in the 5' untranslated region (5'-UTR) of the androgen receptor (AR) gene. We show in patient derived primary genital skin fibroblasts as well as in cell-based reporter assays that this mutation severely impacts AR function by reducing AR protein levels without affecting AR mRNA levels. Importantly, the newly generated uORF translates into a polypeptide and the expression level of this polypeptide inversely correlates with protein translation from the primary ORF of the AR thereby providing a model for AR-5'UTR mediated translational repression. Our findings not only add a hitherto unrecognized genetic cause to complete androgen insensitivity but also underline the importance of 5'UTR mutations affecting uORFs for the pathogenesis of monogenic disorders in general.

Mutations in epithelial growth factor receptor (EGFR), as well as in the EGFR downstream target KRAS are frequently observed in non-small cell lung cancer (NSCLC). Chronic obstructive pulmonary disease (COPD), an independent risk factor for developing NSCLC, is associated with an increased activation of EGFR. In this study we determined presence of EGFR and KRAS hotspot mutations in 325 consecutive NSCLC patients subjected to EGFR and KRAS mutation analysis in the diagnostic setting and for whom the pulmonary function has been determined at time of NSCLC diagnosis. Information about age at diagnosis, sex, smoking status, forced vital capacity (FVC) and forced expiratory volume in 1 sec (FEV1) was collected. Chronic obstructive pulmonary disease(COPD) was defined according to 2013 GOLD criteria. Chi-Square, student t-test and multivariate logistic regression were used to analyze the data. A total of 325 NSCLC patients were included, 193 with COPD and 132 without COPD. COPD was not associated with presence of KRAS hotspot mutations, while EGFR mutations were significantly higher in non-COPD NSCLC patients. Both female gender (HR 2.61; 95% CI: 1.56-4.39; pT and G>C transversions were significantly more frequent in females (86.2%) than in males (61.5%) (p = 0.008). The exon 19del mutation was more frequent in non-smokers (90%) compared to current or past smokers (36.8%). In conclusion, KRAS mutations are more common in females and smokers, but are not associated with COPD-status in NSCLC patients. EGFR mutations are more common in non-smoking NSCLC patients.

Brush border Myosin Ia (MYO1A) has been shown to be frequently mutated in colorectal tumors with microsatellite instability (MSI) and to have tumor suppressor activity in intestinal tumors. Here, we investigated the frequency of frameshift mutations in the A8 microsatellite in exon 28 of MYO1A in MSI gastric and endometrial tumors and found a high mutation rate in gastric (22/47; 46.8%) but not endometrial (3/48; 6.2%) tumors. Using a regression model, we show that MYO1A mutations are likely to confer a growth advantage to gastric, but not endometrial tumors. The mutant MYO1A(7A) protein was shown to lose its membrane localization in gastric cancer cells and a cycloheximide-chase assay demonstrated that the mutant MYO1A(7A) protein has reduced stability compared to the wild type MYO1A. Frequent MYO1A promoter hypermethylation was also found in gastric tumors. Promoter methylation negatively correlates with MYO1A mRNA expression in a series of 58 non-MSI gastric primary tumors (Pearson's r = -0.46; p = 0.0003) but not in a cohort of 54 non-MSI endometrial tumors and treatment of gastric cancer cells showing high MYO1A promoter methylation with the demethylating agent 5-aza-2'-deoxycytidine, resulted in a significant increase of MYO1A mRNA levels. We found that normal gastric epithelial cells, but not normal endometrial cells, express high levels of MYO1A. Therefore, when considered together, our findings suggest that MYO1A has tumor suppressor activity in the normal gastric epithelium but not in the normal endometrium and inactivation of MYO1A either genetically or epigenetically may confer gastric epithelial cells a growth advantage.