The limitation of genetic testing in diagnosing patients suspected for congenital platelet defects

Thrombosis and Hemostasi

Flow cytometric mepacrine fluorescence can be used for the exclusion of platelet dense granule deficiency

Background: δ-storage pool disease (δ-SPD) is a bleeding disorder characterized by a reduced number of platelet-dense granules. The diagnosis of δ-SPD depends on the measurement of platelet ADP content, but this test is time consuming and requires a relatively large blood volume. Flow cytometric analysis of platelet mepacrine uptake is a potential alternative, but this approach lacks validation, which precludes its use in a diagnostic setting. Objectives: To evaluate the performance of platelet mepacrine uptake as a diagnostic test for δ-SPD. Patients/Methods: Mepacrine fluorescence was determined with flow cytometry before and after platelet activation in 156 patients with a suspected platelet function disorder and compared with platelet ADP content as a reference test. Performance was analyzed with a receiver operating characteristic (ROC) curve. Results: Eleven of 156 patients had δ-SPD based on platelet ADP content. Mepacrine fluorescence was inferior to platelet ADP content in identifying patients with δ-SPD, but both mepacrine uptake (area under the ROC curve [AUC] 0.87) and mepacrine release after platelet activation (AUC 0.80) had good discriminative ability. In our tertiary reference center, mepacrine uptake showed high negative predicitive value (97%) with low positive predictive value (35%). Combined with a negative likelihood ratio of 0.1, these data indicate that mepacrine uptake can be used to exclude δ-SPD in patients with a bleeding tendency. Conclusion: Mepacrine fluorescence can be used as a screening tool to exclude δ-SPD in a large number of patients with a suspected platelet function disorder

Flow cytometric mepacrine fluorescence can be used for the exclusion of platelet dense granule deficiency

Background: δ-storage pool disease (δ-SPD) is a bleeding disorder characterized by a reduced number of platelet-d

The limitation of genetic testing in diagnosing patients suspected for congenital platelet defects

Thrombosis and Hemostasi

Mapping the Diagnostic Maze of Platelet Function Disorders

Platelet function disorders are characterized by (spontaneous) bruising, prolonged bleeding time and menorrhagia. Severe platelet function disorders can give rise to life threatening bleeding episodes, but unrecognized mild platelet function disorders can be equally dangerous. Therefore, the diagnosis of platelet function disorders is important. In almost half of the patients with platelet function disorders, the exact cause remains unclear, so there is an unmet need for improved diagnostic tests to identify a platelet function disorder in patients with a bleeding tendency. The aim of this thesis is to improve the diagnostic approach for platelet function disorders and to identify the pathophysiological mechanism in patients with bleeding symptoms of unknown cause. A potential test for the diagnosis of platelet function disorders, recommended by the International Society for Thrombosis and Haemostasis, is to measure agonist induced platelet reactivity on the flow cytometer. However, this test is barely used as a diagnostic tool because it lacks validation and standardization. In chapter 2 the challenges among validating and implementing new diagnostic tests for platelet function disorders are discussed. The main obstacle is the lack of a gold standard to compare a new assay, but it is feasible in a clearly defined patient population, in which many other causes for a bleeding disorder are excluded. Chapter 3 describes the validation of this flow cytometric platelet function test and showed that it has added value in diagnosing a platelet function disorder compared to light transmission aggregometry (LTA), the most commonly used platelet function test. However, it is not yet able to replace this assay and should be performed in addition to this test. Another application of this flow cytometric test is the use of mepacrine fluorescence for diagnosis dense granule deficiency. It has been shown in chapter 4 that mepacrine fluorescence is well suited as a screening tool for dense granule deficiency. Although genetic testing was suggested to be promising in diagnosing platelet function disorders, the diagnostic value of genetic analysis in patients with suspected platelet function disorders was limited, as described in chapter 5. Therefore, we are still in a better understanding of the role of genetics in platelet function disorders. In Chapter 6 we have identified and described a new mechanism of severe macrothrombocytopenia in two young sisters with a GNE mutation. This genetic variant results in hyposialylated platelets, which are rapidly removed from the circulation in the liver. Finally, chapter 7 shows how bleeding symptoms can occur as a consequence of monoclonal gammopathy and that those bleeding symptoms can be successfully managed by threating the monoclonal antibody. The work presented in this thesis contributes to the better understanding of platelet function disorders and its diagnosis. However, the diagnostic maze of platelet function disorders is not solved yet and deserves further research to help those patients to find their diagnosis

Clinical view versus guideline adherence in ferritin monitoring and initiating iron chelation therapy in patients with myelodysplastic syndromes

OBJECTIVES: In patients with myelodysplastic syndromes (MDS) with >20 transfusions and ferritin levels >1000 μg/L, international guidelines recommend iron chelation therapy (ICT). The study's objective was to determine guideline adherence and the intensity of ferritin monitoring in clinical practice. METHODS: We performed an observational population-based study using the HemoBase Registry, which contains data of all MDS patients diagnosed since 2005 in Friesland, the Netherlands. Clinical information on transfusions, ferritin measurements, ICT, and clinical performance as defined by age ≤80 years, Charlson Comorbidity Index 20 transfusions. In 57 of these 121 patients (47.1%), ferritin measurements were performed at least once. Clinical performance was significantly associated with monitoring ferritin around the 20th transfusion (RR: 2.49, p=0.016). Clinical performance was also associated with initiating ICT (RR: 5.99, p20 transfusions, and clinical performance was significantly associated with measuring ferritin. Our study suggests that in heavily transfused MDS patients, ferritin monitoring is primarily based on patients' clinical performance rather than guideline recommendations