scholarly journals Use of Targeted High-Throughput Sequencing for Genetic Classification of Patients with Bleeding Diathesis and Suspected Platelet Disorder

TH Open ◽  
2018 ◽  
Vol 02 (04) ◽  
pp. e445-e454 ◽  
Author(s):  
Oliver Andres ◽  
Eva-Maria König ◽  
Karina Althaus ◽  
Tamam Bakchoul ◽  
Peter Bugert ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Genetic Diagnosis ◽  
Individual Risk ◽  
Recurrent Bleeding ◽  
Bleeding Diathesis ◽  
Genetic Classification ◽  
Diagnostic Algorithms ◽  
Platelet Disorder ◽  
Platelet Disorders

AbstractInherited platelet disorders (IPD) form a rare and heterogeneous disease entity that is present in about 8% of patients with non-acquired bleeding diathesis. Identification of the defective cellular pathway is an important criterion for stratifying the patient's individual risk profile and for choosing personalized therapeutic options. While costs of high-throughput sequencing technologies have rapidly declined over the last decade, molecular genetic diagnosis of bleeding and platelet disorders is getting more and more suitable within the diagnostic algorithms. In this study, we developed, verified, and evaluated a targeted, panel-based next-generation sequencing approach comprising 59 genes associated with IPD for a cohort of 38 patients with a history of recurrent bleeding episodes and functionally suspected, but so far genetically undefined IPD. DNA samples from five patients with genetically defined IPD with disease-causing variants in WAS, RBM8A, FERMT3, P2YR12, and MYH9 served as controls during the validation process. In 40% of 35 patients analyzed, we were able to finally detect 15 variants, eight of which were novel, in 11 genes, ACTN1, AP3B1, GFI1B, HPS1, HPS4, HPS6, MPL, MYH9, TBXA2R, TPM4, and TUBB1, and classified them according to current guidelines. Apart from seven variants of uncertain significance in 11% of patients, nine variants were classified as likely pathogenic or pathogenic providing a molecular diagnosis for 26% of patients. This report also emphasizes on potentials and pitfalls of this tool and prospectively proposes its rational implementation within the diagnostic algorithms of IPD.

scholarly journals Introducing high-throughput sequencing into mainstream genetic diagnosis practice in inherited platelet disorders

Haematologica ◽  
2017 ◽  
Vol 103 (1) ◽  
pp. 148-162 ◽  
Author(s):  
José M. Bastida ◽  
María L. Lozano ◽  
Rocío Benito ◽  
Kamila Janusz ◽  
Verónica Palma-Barqueros ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Genetic Diagnosis ◽  
Platelet Disorders

scholarly journals Accurate fetal variant calling in the presence of maternal cell contamination

2019 ◽  
Author(s):  
Elena Nabieva ◽  
Satyarth Mishra Sharma ◽  
Yermek Kapushev ◽  
Sofya K. Garushyants ◽  
Anna V. Fedotova ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Chorionic Villus ◽  
Genetic Diagnosis ◽  
Variant Calling ◽  
Data Availability ◽  
Training Data ◽  
Sequencing Data ◽  
Maternal Cell ◽  
Fetal Dna

AbstractHigh-throughput sequencing of fetal DNA is a promising and increasingly common method for the discovery of all (or all coding) genetic variants in the fetus, either as part of prenatal screening or diagnosis, or for genetic diagnosis of spontaneous abortions. In many cases, the fetal DNA (from chorionic villi, amniotic fluid, or abortive tissue) can be contaminated with maternal cells, resulting in the mixture of fetal and maternal DNA. This maternal cell contamination (MCC) undermines the assumption, made by traditional variant callers, that each allele in a heterozygous site is covered, on average, by 50% of the reads, and therefore can lead to erroneous genotype calls. We present a panel of methods for reducing the genotyping error in the presence of MCC. All methods start with the output of GATK HaplotypeCaller on the sequencing data for the (contaminated) fetal sample and both of its parents, and additionally rely on information about the MCC fraction (which itself is readily estimated from the high-throughput sequencing data). The first of these methods uses a Bayesian probabilistic model to correct the fetal genotype calls produced by MCC-unaware HaplotypeCaller. The other two methods “learn” the genotype-correction model from examples. We use simulated contaminated fetal data to train and test the models. Using the test sets, we show that all three methods lead to substantially improved accuracy when compared with the original MCC-unaware HaplotypeCaller calls. We then apply the best-performing method to three chorionic villus samples from spontaneously terminated pregnancies.Code and training data availabilityhttps://github.com/bazykinlab/ML-maternal-cell-contamination

scholarly journals A high-throughput sequencing test for diagnosing inherited bleeding, thrombotic, and platelet disorders

Blood ◽  
2016 ◽  
Vol 127 (23) ◽  
pp. 2791-2803 ◽  
Author(s):  
Ilenia Simeoni ◽  
Jonathan C. Stephens ◽  
Fengyuan Hu ◽  
Sri V. V. Deevi ◽  
Karyn Megy ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Genetic Test ◽  
Targeted Sequencing ◽  
Sequencing Platform ◽  
Key Points ◽  
Platelet Disorders

Key Points Developed a targeted sequencing platform covering 63 genes linked to heritable bleeding, thrombotic, and platelet disorders. The ThromboGenomics platform provides a sensitive genetic test to obtain molecular diagnoses in patients with a suspected etiology.

scholarly journals Diagnostic high-throughput sequencing of 2,390 patients with bleeding, thrombotic and platelet disorders

2018 ◽  
Author(s):  
Kate Downes ◽  
Karyn Megy ◽  
Daniel Duarte ◽  
Minka Vries ◽  
Johanna Gebhart ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Clinical Phenotype ◽  
Careful Consideration ◽  
Clinical Diagnostics ◽  
Genomic Variation ◽  
Panel Test ◽  
Platelet Disorders ◽  
Normal Hemostasis ◽  
And Function

A targeted high-throughput sequencing (HTS) panel test for clinical diagnostics requires careful consideration of the inclusion of appropriate diagnostic-grade genes, the ability to detect multiple types of genomic variation with high levels of analytic sensitivity and reproducibility, and variant interpretation by a multi-disciplinary team (MDT) in the context of the clinical phenotype. We have sequenced 2,390 index patients using the ThromboGenomics HTS panel test of diagnostic-grade genes known to harbour variants associated with rare bleeding, thrombotic or platelet disorders (BPD). The diagnostic rate was determined by the clinical phenotype, with an overall rate of 50.4% for all thrombotic, coagulation, platelet count and function disorder patients and a rate of 6.2% for patients with unexplained bleeding disorders characterized by normal hemostasis test results. The MDT classified 756 unique variants, including copy number and intronic variants, as Pathogenic, Likely Pathogenic or Variants of Uncertain Significance. Almost half (49.7%) of these variants are novel and 41 unique variants were identified in 7 genes recently found to be implicated in BPD. Inspection of canonical hemostasis pathways identified 29 patients with evidence of oligogenic inheritance. A molecular diagnosis has been reported for 897 index patients providing evidence that introducing a HTS genetic test for BPD patients is meeting an important unmet clinical need.

scholarly journals Benefits of clinical criteria and high-throughput sequencing for diagnosing children with syndromic craniosynostosis

2020 ◽  
Author(s):  
Elin Tønne ◽  
Bernt Johan Due-Tønnessen ◽  
Inger-Lise Mero ◽  
Ulrikke Straume Wiig ◽  
Mari Ann Kulseth ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Diagnostic Yield ◽  
Single Gene ◽  
Genetic Diagnosis ◽  
Gene Analysis ◽  
Comparative Genomic ◽  
Genetic Syndromes ◽  
Clinical Criteria ◽  
Syndromic Craniosynostosis

AbstractAn accurate diagnosis of syndromic craniosynostosis (CS) is important for personalized treatment, surveillance, and genetic counselling. We describe detailed clinical criteria for syndromic CS and the distribution of genetic diagnoses within the cohort. The prospective registry of the Norwegian National Unit for Craniofacial Surgery was used to retrieve individuals with syndromic CS born between 1 January 2002 and 30 June 2019. All individuals were assessed by a clinical geneticist and classified using defined clinical criteria. A stepwise approach consisting of single-gene analysis, comparative genomic hybridization (aCGH), and exome-based high-throughput sequencing, first filtering for 72 genes associated with syndromic CS, followed by an extended trio-based panel of 1570 genes were offered to all syndromic CS cases. A total of 381 individuals were registered with CS, of whom 104 (27%) were clinically classified as syndromic CS. Using the single-gene analysis, aCGH, and custom-designed panel, a genetic diagnosis was confirmed in 73% of the individuals (n = 94). The diagnostic yield increased to 84% after adding the results from the extended trio-based panel. Common causes of syndromic CS were found in 53 individuals (56%), whereas 26 (28%) had other genetic syndromes, including 17 individuals with syndromes not commonly associated with CS. Only 15 individuals (16%) had negative genetic analyses. Using the defined combination of clinical criteria, we detected among the highest numbers of syndromic CS cases reported, confirmed by a high genetic diagnostic yield of 84%. The observed genetic heterogeneity encourages a broad genetic approach in diagnosing syndromic CS.

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