scholarly journals αIIbβ3 variants defined by next-generation sequencing: Predicting variants likely to cause Glanzmann thrombasthenia

2015 ◽  
Vol 112 (15) ◽  
pp. E1898-E1907 ◽  
Author(s):  
Lorena Buitrago ◽  
Augusto Rendon ◽  
Yupu Liang ◽  
Ilenia Simeoni ◽  
Ana Negri ◽  
...  
Keyword(s):  
Amino Acids ◽  
Next Generation Sequencing ◽  
Hek293 Cells ◽  
Next Generation ◽  
Glanzmann Thrombasthenia ◽  
Missense Variants ◽  
Sorting Intolerant From Tolerant ◽  
Fibrinogen Binding ◽  
Novel Variants ◽  
Generation Sequencing

Next-generation sequencing is transforming our understanding of human genetic variation but assessing the functional impact of novel variants presents challenges. We analyzed missense variants in the integrin αIIbβ3 receptor subunit genes ITGA2B and ITGB3 identified by whole-exome or -genome sequencing in the ThromboGenomics project, comprising ∼32,000 alleles from 16,108 individuals. We analyzed the results in comparison with 111 missense variants in these genes previously reported as being associated with Glanzmann thrombasthenia (GT), 20 associated with alloimmune thrombocytopenia, and 5 associated with aniso/macrothrombocytopenia. We identified 114 novel missense variants in ITGA2B (affecting ∼11% of the amino acids) and 68 novel missense variants in ITGB3 (affecting ∼9% of the amino acids). Of the variants, 96% had minor allele frequencies (MAF) < 0.1%, indicating their rarity. Based on sequence conservation, MAF, and location on a complete model of αIIbβ3, we selected three novel variants that affect amino acids previously associated with GT for expression in HEK293 cells. αIIb P176H and β3 C547G severely reduced αIIbβ3 expression, whereas αIIb P943A partially reduced αIIbβ3 expression and had no effect on fibrinogen binding. We used receiver operating characteristic curves of combined annotation-dependent depletion, Polyphen 2-HDIV, and sorting intolerant from tolerant to estimate the percentage of novel variants likely to be deleterious. At optimal cut-off values, which had 69–98% sensitivity in detecting GT mutations, between 27% and 71% of the novel αIIb or β3 missense variants were predicted to be deleterious. Our data have implications for understanding the evolutionary pressure on αIIbβ3 and highlight the challenges in predicting the clinical significance of novel missense variants.

scholarly journals αIIbβ3 Variants Defined By Next Generation Sequencing: Implications for Predicting Variants Likely to Cause Glanzmann Thrombasthenia and Alloimmune Disorders

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4151-4151
Author(s):  
Claudia Lorena Buitrago ◽  
Augusto Rendon ◽  
Ernest Turro ◽  
Yupu Liang ◽  
Ilenia Simeoni ◽  
...  
Keyword(s):  
Amino Acids ◽  
Missense Variant ◽  
The Novel ◽  
Glanzmann Thrombasthenia ◽  
Sequencing Project ◽  
Missense Variants ◽  
Exome Sequencing Project ◽  
Whole Exome ◽  
Novel Variants ◽  
Generation Sequencing

Abstract # Authors contributed equally to this work. ~ Currently at Genomics England Ltd, London, United Kingdom Next generation sequencing is transforming our understanding of human genetic variation and is becoming a routine part of human genetic analysis. The identification of millions of new variants, which are mainly rare and assessing their implications for human health presents new challenges to researchers and clinicians. We have analyzed missense variants in the ITGB2A and ITGB3 genes obtained from whole exome and whole genome sequencing (WES & WGS) data from 5 databases: The Human Genome Mutation Database, the 1000 Genomes project, the UK10K Whole Exome Sequencing project, the UK10K Whole Genome Sequencing project, and The National Heart, Lung and Blood Institute Exome Sequencing Project. Together, these encompass variants of the platelet αIIbβ3 integrin receptor from ~32,000 alleles derived from 16,108 individuals. We identified 111 missense variants that have previously been associated with Glanzmann thrombasthenia (GT), 20 variants associated with alloimmune thrombocytopenia, and 5 variants associated with aniso/macrothrombocytopenia. None of the GT variants were found in the last four databases, indicating that they have minor allele frequencies (MAF) less than ~0.01%, attesting to both their rarity and the likelihood that they entered the population within the last ~2,500 years. We also identified 114 novel missense variants in ITGB2A affecting ~11% of the amino acids and 68 novel missense variants in ITGB3 affecting ~9% of the amino acids. 96% of the novel variants had MAF <0.1%, indicating their rarity. Based on sequence conservation, MAF, and/or location of the substituted residue on a complete model of αIIbβ3 that suggested a possible effect on protein folding, we selected three novel variants (αIIb P943A and P176H, and β3 C547G) that affect amino acids previously associated with GT for expression in HEK 293 cells. Both αIIb P176H and β3 C547G severely affected αIIbβ3 expression, whereas αIIb P943A had only a partial effect on expression and no effect on DTT-induced fibrinogen binding. We were not surprised that the latter variant did not have a severe effect on expression or function because it has an MAF (0.46%) that is much higher than the MAFs of the other GT-causing variants. To estimate the percentage of the 114 novel identified variants that are likely to be deleterious we used 3 different algorithms, CADD, Polyphen 2-HDVI, and SIFT. The algorithms showed moderate concordance in their rankings of the likelihood that a variant is deleterious. To compare their predictive powers, we performed receiver operating characteristic (ROC) analysis based on their ability to discriminate confirmed GT missense variants (positive controls) from alloantigens (negative controls); the area under the curve (AUC) values were 0.91, 0.88, and 0.90, respectively. At cutoff values that achieved greater than 95% sensitivity for each algorithm: 1) the specificity values were 75%, 65%, and 60%, and 2) the percentages of novel αIIb+β3 missense variants predicted to be deleterious were 43%, 56%, and 58%. Polyphen 2-HDVI and SIFT identified αIIb P176H and β3 C547G as highly likely to be deleterious and αIIb P943A as much less likely to be deleterious, whereas CADD did not differentiate them in the same way. We conclude that ~1.1% of individuals in the populations studied carry at least one missense variant in αIIb or β3 and that 0.6% carry a variant that might be deleterious and therefore may result in a hemorrhagic GT-like phenotype. The rarity of almost all of the novel missense variants identified indicates that they entered the population recently. Despite having detailed knowledge of the structure and function of αIIbβ3, it is difficult to predict with certainty the impact of any single missense variant. This will pose serious challenges as more individuals undergo WES and WGS; we anticipate that linkage to health record data, as will happen for the UK 100,000 Genomes project, will aid clinical interpretation. Finally, “hypomorphic” gene variants that produce only a partial decrease in expression, such as αIIb P943A, may contribute to the wide variation in αIIbβ3 surface expression observed in the healthy population. Disclosures No relevant conflicts of interest to declare.

scholarly journals Next-generation sequencing reveals three novel variants in Polish patients with Usher syndrome

Klinika Oczna ◽  
2018 ◽  
Vol 2018 (4) ◽  
pp. 189-194
Author(s):  
Anna Wawrocka ◽  
Joanna Walczak-Sztulpa ◽  
Anna Skorczyk-Werner ◽  
Łukasz Kuszel ◽  
Magdalena Socha ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Usher Syndrome ◽  
Next Generation ◽  
Novel Variants ◽  
Generation Sequencing

Molecular genetic diagnosis by next-generation sequencing in a cohort of Mexican patients with haemophilia and report of novel variants

2020 ◽  
Vol 83 ◽  
pp. 102423 ◽  
Author(s):  
Laura Villarreal-Martínez ◽  
Marisol Ibarra-Ramirez ◽  
Geovana Calvo-Anguiano ◽  
José de Jesús Lugo-Trampe ◽  
Hilda Luna-Záizar ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Molecular Genetic ◽  
Genetic Diagnosis ◽  
Next Generation ◽  
Novel Variants ◽  
Molecular Genetic Diagnosis ◽  
Generation Sequencing ◽  
Mexican Patients

scholarly journals A universal primer-independent next-generation sequencing approach for investigations of norovirus outbreaks and novel variants

2017 ◽  
Vol 7 (1) ◽  
Author(s):  
Jannik Fonager ◽  
Marc Stegger ◽  
Lasse Dam Rasmussen ◽  
Mille Weismann Poulsen ◽  
Jesper Rønn ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Next Generation ◽  
Universal Primer ◽  
Novel Variants ◽  
Generation Sequencing

The utility of next-generation sequencing technologies in diagnosis of Mendelian mitochondrial diseases and reflections on clinical spectrum

2021 ◽  
Vol 34 (4) ◽  
pp. 417-430
Author(s):  
Melis Kose ◽  
Esra Isik ◽  
Ayça Aykut ◽  
Asude Durmaz ◽  
Engin Kose ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Exome Sequencing ◽  
Diagnostic Process ◽  
Unknown Etiology ◽  
Next Generation ◽  
Related Gene ◽  
Targeted Exome Sequencing ◽  
Novel Variants ◽  
Gene Defects ◽  
Generation Sequencing

Abstract Objectives Diagnostic process of mitochondrial disorders (MD) is challenging because of the clinical variability and genetic heterogeneity of these conditions. Next-Generation Sequencing (NGS) technology offers a high-throughput platform for nuclear MD. Methods We included 59 of 72 patients that undergone WES and targeted exome sequencing panel suspected to have potential PMDs. Patients who were included in the analysis considering the possible PMD were reviewed retrospectively and scored according to the Mitochondrial Disease Criteria Scale. Results Sixty-one percent of the patients were diagnosed with whole-exome sequencing (WES) (36/59) and 15% with targeted exome sequencing (TES) (9/59). Patients with MD-related gene defects were included in the mito group, patients without MD-related gene defects were included in the nonmito group, and patients in whom no etiological cause could be identified were included in the unknown etiology group. In 11 out of 36 patients diagnosed with WES, a TES panel was applied prior to WES. In 47 probands in 39 genes (SURF1, SDHAF1, MTO1, FBXL4, SLC25A12, GLRX5, C19oRF12, NDUFAF6, DARS2, BOLA3, SLC19A3, SCO1, HIBCH, PDHA1, PDHAX, PC, ETFA, TRMU, TUFM, NDUFS6, WWOX, UBCD TREX1, ATL1, VAC14, GFAP, PLA2G6, TPRKB, ATP8A2, PEX13, IGHMBP2, LAMB2, LPIN1, GFPT1, CLN5, DOLK) (20 mito group, 19 nonmito group) 59 variants (31 mito group, 18 nonmito group) were detected. Seven novel variants in the mito group (SLC25A12, GLRX5, DARS2, SCO1, PC, ETFA, NDUFS6), nine novel variants in the nonmito group (IVD, GCDH, COG4, VAC14, GFAP, PLA2G6, ATP8A2, PEX13, LPIN1) were detected. Conclusions We explored the feasibility of identifying pathogenic alleles using WES and TES in MD. Our results show that WES is the primary method of choice in the diagnosis of MD until at least all genes responsible for PMD are found and are highly effective in facilitating the diagnosis process.

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