Establishment of Various Specific Parameters for the Routine Diagnostics in Patients with Glanzmann Thrombasthenia

Objectives: The aim of the study was to establish specific parameters for the routine diagnostics of the Glanzmann Thrombasthenia (GT). GT is a hereditary function disorder caused by mutations in the genes encoding αIIbβ3. Patients with GT have a strong impairment of platelet function due to a quantitative or qualitative defect of the fibrinogen receptor. This receptor consists of two subunits, the glycoprotein IIb (GPIIb or αIIb) and the glycoprotein IIIa (GPIIIa or β3), and links activated platelets through fibrinogen bridges. Various clinical and functional parameters were determined in a collective consisting of eight patients.

Methods: The study investigated eight patients suffering from GT. First, various clinical laboratory parameters were determined. For the functional analysis we used the PFA-200 system, as well as the aggregometry (born method). For the typing of GT, a flow cytometric analysis was performed. In addition, the genes ITGA2B (αIIb) and ITGB3 (β3) were sequenced using the Sanger method.

Results: In our patient population a normal platelet count, pathologic PFA-closure times and an absence of aggregation in response to different agonists could be observed (except ristocetin). Subsequently, various surface proteins were quantified by flow cytometry. The absence of αIIb and β3 could be detected in six patients (patient 3-8). Two patients showed a receptor expression of about 5-15% in comparison to the donor population. Although both subunits were detected on the platelet surface, no activated receptor could be observed.

Sequencing showed a p.Gln626His mutation in the ITGA2B gene of patients 5 and 6 (sibling pair), resulting in a splicing anomaly. In patient 3 an exchange of valine to glutamine at position 422 could be detected. Patient 4 showed a deletion of 8 bp in the ITGA2B gene, which results in a frameshift. Both mutations have not been described so far. In patients 1 and 2, a new mutation could also be identified in exon 10 of the ITGB3 gene. This mutation results in an amino acid substitution, leading to destruction of a disulfide bridge.

Patients 7 and 8 each had only one heterozygous mutation located in the ITGA2B gene, however these patients showed the pathognomic characteristics of GT. These findings are discussed on the poster.

Conclusions: We recommend that patients with abnormal history of bleeding and standard advanced bleeding diagnostics (von Willebrand factor, single factor deficiency,...) visit a specialized laboratory with flow cytometry or sequencing capabilities.

In our examined patient collective we could find besides the typical characteristics of the GT (e.g. normal platelet count, pathological PFA-closure times and pathognomonic aggregation) an impairment in the surface expression of αIIbβ3 (quantitative and/or qualitative). With the Sanger sequencing we were able to determine homozygous pathogenic mutations in six patients, two patients showed one heterozygous mutation each.