| 1. |
- Andersson, Nadine, et al.
(författare)
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Genetic screening of children with suspected inherited bleeding disorders
- 2020
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Ingår i: Haemophilia. - : Wiley. - 1351-8216 .- 1365-2516. ; 26:2, s. 314-324
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Tidskriftsartikel (refereegranskat)abstract
- INTRODUCTION: Genetic screening using high-throughput DNA sequencing has become a tool in diagnosing patients with suspected inherited bleeding disorders (IBD). However, its usefulness and diagnostic efficacy in children is unclear.AIM: To evaluate the diagnostic efficacy of genetic screening for IBD in children and downstream further testing.METHODS: After informed consent, children (<18 years) with suspected IBD underwent genetic screening with 94 selected genes.RESULTS: A total of 68 heterozygous class 3-5 variants were detected in 30 children, 2.3 variants per patient. Directed specific functional testing was performed after genetic screening in a subset of patients. Adhering to the ACMG guidelines, the results of functional testing together with family history and previous publications classified three variants as likely disease causing (class 4) and two variants as disease causing (class 5), all in children with thrombocytopenia. The overall diagnostic rate was 16.7% (5/30). Children with thrombocytopenia had a significantly higher rate of significant genetic findings, 5/9 (55.6%) vs. 0/21 (0%; P = .0009).CONCLUSION: We conclude that performing genetic screening in children is an effective tool especially for children with inherited thrombocytopenia and has the possibility to diagnose platelet disorders adequately early in life. Children with bleeding diathesis, normal coagulation work-up and without thrombocytopenia are unlikely to be diagnosed by genetic screening. Ethical issues such as incidental findings, variants associated with cancer and the interpretation of the genetic results into clinical practice remain problematic.
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| 2. |
- Fager Ferrari, Marcus, et al.
(författare)
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A rare heterozygous variant in FGB (Fibrinogen Merivale) causing hypofibrinogenemia in a Swedish family
- 2020
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Ingår i: Blood Coagulation and Fibrinolysis. - 1473-5733. ; 31:7, s. 481-484
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Tidskriftsartikel (refereegranskat)abstract
- Fibrinogen is essential for normal hemostasis. Congenital fibrinogen disorders (afibrinogenemia, hypofibrinogenemia, dysfibrinogenemia and hypodysfibrinogenemia), caused by pathogenic variants in the genes FGA, FGB and FGG, have the potential of causing bleeding diathesis and/or thrombotic events of variable severity. We describe a case of familial hypofibrinogenemia in a Swedish family. The proband is a 27-year-old woman, with a history of significant bleeding diathesis. She was diagnosed with moderate hypofibrinogenemia (0.8 g/l), and genetic screening identified a rare heterozygous missense variant in FGB (c.854G>A, p.Arg285His) (Fibrinogen Merivale) previously described in a New Zealand European family with symptomatic hypofibrinogenemia. The father, sister and brother of the proband also harbored the FGB variant, segregating with hypofibrinogenemia (0.9-1.2 g/l). The proband showed a more severe bleeding phenotype compared with her other hypofibrinogenemic family members; this was attributed to a concomitant platelet dysfunction, also present in her normofibrinogenemic mother.
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| 3. |
- Fager Ferrari, Marcus, et al.
(författare)
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Germline heterozygous variants in genes associated with familial hemophagocytic lymphohistiocytosis as a cause of increased bleeding
- 2018
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Ingår i: Platelets. - : Informa UK Limited. - 0953-7104 .- 1369-1635. ; 29:1, s. 56-64
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Tidskriftsartikel (refereegranskat)abstract
- Familial hemophagocytic lymphohistiocytosis (FHL) is caused by biallelic variants in genes regulating granule secretion in cytotoxic lymphocytes. In FHL3–5, the affected genes UNC13D, STX11 and STXBP2 have further been shown to regulate the secretion of platelet granules, giving rise to compromised platelet function. Therefore, we aimed to investigate platelet degranulation in patients heterozygous for variants in UNC13D, STX11 and STXBP2. During the work-up of patients referred to the Coagulation Unit, Skåne University Hospital, Malmö, Sweden and the Department of Hematology, Rigshospitalet, Copenhagen, Denmark due to bleeding tendencies, 12 patients harboring heterozygous variants in UNC13D, STX11 or STXBP2 were identified using targeted whole exome sequencing. Transmission electron microscopy (TEM) was used to assess the secretion of platelet dense granules following thrombin stimulation. Platelet degranulation, activation and aggregation were further assessed by flow cytometry (FC) and light transmission aggregometry (LTA) with lumi-aggregometry. In total, eight out of twelve (67%) patients showed impaired degranulation by at least one of the assays (TEM, FC and LTA). In the 12 patients, eight different heterozygous variants were identified. One variant was strongly associated with impaired degranulation, while four of the variants were associated with impaired granule secretion to a slightly lesser extent. One additional variant was found in six out of the twelve patients, and was associated with varying degrees of degranulation impairment. Accordingly, six out of the eight (75%) identified variants were associated with impaired platelet degranulation. Our results suggest that heterozygous variants in UNC13D, STX11 and STXBP2 are sufficient to cause platelet secretion defects resulting in increased bleeding.
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| 4. |
- Leinøe, Eva, et al.
(författare)
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Application of whole-exome sequencing to direct the specific functional testing and diagnosis of rare inherited bleeding disorders in patients from the Öresund Region, Scandinavia
- 2017
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Ingår i: British Journal of Haematology. - : Wiley. - 0007-1048. ; 179:2, s. 308-322
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Tidskriftsartikel (refereegranskat)abstract
- Rare inherited bleeding disorders (IBD) are a common cause of bleeding tendency. To ensure a correct diagnosis, specialized laboratory analyses are necessary. This study reports the results of an upfront diagnostic strategy using targeted whole exome sequencing. In total, 156 patients with a significant bleeding assessment tool score participated in the study, of which a third had thrombocytopenia. Eighty-seven genes specifically associated with genetic predisposition to bleeding were analysed by whole exome sequencing. Variants were classified according to the five-tier scheme. We identified 353 germline variants. Eight patients (5%) harboured a known pathogenic variant. Of the 345 previously unknown variants, computational analyses predicted 99 to be significant. Further filtration according to the Mendelian inheritance pattern, resulted in 59 variants being predicted to be clinically significant. Moreover, 34% (20/59) were assigned as novel class 4 or 5 variants upon targeted functional testing. A class 4 or 5 variant was identified in 30% of patients with thrombocytopenia (14/47) versus 11% of patients with a normal platelet count (12/109) (P < 0·01). An IBD diagnosis has a major clinical impact. The genetic investigations detailed here extricated our patients from a diagnostic conundrum, thus demonstrating that continuous optimization of the diagnostic work-up of IBD is of great benefit.
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| 5. |
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| 6. |
- Fager Ferrari, Marcus, et al.
(författare)
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Collagen remodelling and plasma ascorbic acid levels in patients suspected of inherited bleeding disorders harbouring germline variants in collagen‐related genes
- 2021
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Ingår i: Haemophilia. - : Wiley. - 1351-8216 .- 1365-2516. ; 27:1, s. 69-77
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Tidskriftsartikel (refereegranskat)abstract
- INTRODUCTION: Variants in collagen-related genes COL1A1, COL3A1, COL5A1 andCOL5A2 are associated with Ehlers-Danlos syndrome (EDS), a heterogeneous groupof connective tissue disorders strongly associated with increased bleeding. Of patientswith incompletely explained bleeding diathesis, a relatively high proportion were shown to harbour at least one heterozygous variant of unknown significance (VUS) in one of these genes, the vast majority without meeting the clinical criteria for EDS.AIM: To investigate the functional consequences of the identified variants by assessingthe formation and degradation of types I, III and V collagen, in addition to plasmalevels of ascorbic acid (AA).METHODS: A total of 31 patients harbouring at least one heterozygous VUS in COL1A1, COL3A1, COL5A1 or COL5A2 and 20 healthy controls were assessed using monoclonal antibodies targeting neo-epitopes specific for collagen formation and degradation. Plasma AA levels were measured in patients using high-performance liquid chromatography.RESULTS: Serum levels of C5M (degradation of type V collagen) were decreased inpatients compared with healthy controls (p = .033). No significant differences werefound in biomarkers for remodelling of types I and III collagen. A significant negativecorrelation between bleeding (ISTH-BAT score) and plasma AA levels was shown(r = −.42; r2 = .17; p = .020). Suboptimal or marginally deficient AA status was foundin 8/31 patients (26%).CONCLUSION: Functional investigations of collagen remodelling were not able to identify any clear associations between the identified variants and increased bleeding.The negative correlation between plasma AA levels and ISTH-BAT score motivatesfurther investigations.
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| 7. |
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| 8. |
- Fager Ferrari, Marcus, et al.
(författare)
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Evaluation of the Sialidase Inhibitor Oseltamivir in GNE-associated Thrombocytopenia
- 2021
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Ingår i: Research and practice in thrombosis and haemostasis. - 2475-0379. ; 5:S2, s. 644-645
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Konferensbidrag (refereegranskat)abstract
- Background: GNE encodes UDP-N-acetyl-glucosamine-2-epimerase/N-acetylmannosamine kinase, the rate limiting enzyme of sialic acid biosynthesis. Biallelic variants in GNE have recently been associated with severe isolated macrothrombocytopenia, attributed to an increased clearance of desialylated platelets. Interestingly, treatment with the sialidase inhibitor oseltamivir has been reported to increase platelet counts in conditions such as immune thrombocytopenia (ITP) and influenza. We present a case of a 17-year-old boy (the proband) with severe congenital macrothrombocytopenia (platelet counts < 10 x 109/L). Whole genome sequencing revealed two previously undescribed compound heterozygous variants in GNE (c.416_426del, p.Ile139Argfs*4 and c.1352G>A, p.Arg451Gln). The proband was otherwise healthy, with no signs of GNE myopathy. Aims: To investigate the consequences of the identified variants in GNE and evaluate the effect of oseltamivir in GNE-associated thrombocytopenia. Methods: Sialylation of platelets, granulocytes, lymphocytes and monocytes was determined by flow cytometry in the proband and healthy controls (n = 5), using Sambucus nigra lectin (SNA) and Maackia amurensis lectin II (MAL II). Platelet sialylation was reassessed in the proband following treatment with oseltamivir (75 mg twice daily, off-label use). Informed consent was obtained from all participants. The study was approved by the regional ethical committee. Results: Sialylation of platelets and leukocytes was markedly decreased in the proband compared with the healthy controls, consistent with a deleterious effect of the compound heterozygous variants in GNE (Figure 1). Platelet sialylation was persistently decreased after 18 days of treatment with oseltamivir, and no clinically significant elevation of the platelet counts could be observed (Figure 1, Figure 2).
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| 9. |
- Hanes, Jozef, et al.
(författare)
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Evaluation of a novel immunoassay to detect p-tau Thr217 in the CSF to distinguish Alzheimer disease from other dementias.
- 2020
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Ingår i: Neurology. - 1526-632X. ; 95:22
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Tidskriftsartikel (refereegranskat)abstract
- To investigate whether tau phosphorylated at Thr217 (p-tau T217) assay in CSF can distinguish patients with Alzheimer disease (AD) from patients with other dementias and healthy controls.We developed and validated a novel Simoa immunoassay to detect p-tau T217 in CSF. There was a total of 190 participants from 3 cohorts with AD (n = 77) and other neurodegenerative diseases (n = 69) as well as healthy participants (n = 44).The p-tau T217 assay (cutoff 242 pg/mL) identified patients with AD with accuracy of 90%, with 78% positive predictive value (PPV), 97% negative predictive value (NPV), 93% sensitivity, and 88% specificity, compared favorably with p-tau T181 ELISA (52 pg/mL), showing 78% accuracy, 58% PPV, 98% NPV, 71% specificity, and 97% sensitivity. The assay distinguished patients with AD from age-matched healthy controls (cutoff 163 pg/mL, 98% sensitivity, 93% specificity), similarly to p-tau T181 ELISA (cutoff 60 pg/mL, 96% sensitivity, 86% specificity). In patients with AD, we found a strong correlation between p-tau T217 and p-tau T181, total tau and β-amyloid 40, but not β-amyloid 42.This study demonstrates that p-tau T217 displayed better diagnostic accuracy than p-tau T181. The data suggest that the new p-tau T217 assay has potential as an AD diagnostic test in clinical evaluation.This study provides Class III evidence that a CSF immunoassay for p-tau T217 distinguishes patients with AD from patients with other dementias and healthy controls.
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| 10. |
- Leinoe, Eva, et al.
(författare)
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Highly impaired platelet ultrastructure in two families with novel IKZF5 variants
- 2021
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Ingår i: Platelets. - : Informa UK Limited. - 0953-7104 .- 1369-1635. ; 32:4, s. 492-497
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Tidskriftsartikel (refereegranskat)abstract
- Heterozygous variants in the IKZF5 gene, encoding transcription factor Pegasus, were recently discovered to be causal of inherited thrombocytopenia (IT). We screened 90 patients suspected of inherited thrombocytopenia for variants in 101 genes associated with inherited bleeding disorders and report the clinical presentation of two Danish families with novel variants in IKZF5. Platelet ultrastructure and cytoskeleton were evaluated by immunofluorescent microscopy (IF) and found to be highly abnormal, demonstrating severe disturbances of distribution and expression of non-muscular myosin, filamin, β-tubulin and α tubulin. Number of alpha granules were reduced, and platelets elongated when evaluated by TEM. In both families a child carrying a rare IKZF5 variant was affected by developmental delay. The proband of family A presented with recurrent infections and was examined for an immunodeficiency. The concentration of naive B-cells was found moderately reduced by leucocyte subpopulation examination, indicating an impaired cellular immunity. T-cells were marginally low with reduced share and concentration of CD45RApos, CD31pos, CD4pos recent thymic immigrants as signs of reduced thymic output. The novel IKZF5 variants co-segregated with thrombocytopenia in both families and both probands had significant bleeding tendency. Through comprehensive characterizations of the platelet morphology and function linked to the specific phenotypes we add novel insight to IKZF5-associated thrombocytopenia, which may help to identify and classify more cases with IKZF5 associated IT.
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