Glycoprotein-specific antibodies modulates platelet phenotypes in an epitope-dependent manner

Introduction: Immune thrombocytopenia (ITP) is an acquired autoimmune disorder characterized by a decreased platelet count due to accelerated platelet destruction and impaired megakaryopoiesis. Antibodies targeting platelet glycoproteins (GPs) are implicated in the pathogenesis of ITP, although the exact mechanisms remain uncertain. Accumulating evidence suggests that resting human platelets develop different phenotypes upon stimulation with agonists such as aggregatory, apoptotic, procoagulatory or necroptotic phenotype. In this study, we investigated the effect of GP-specific monoclonal antibodies (mAbs) that mimic ITP-antibodies on platelet phenotypes.

Method: Washed platelets were incubated with various GP-specific mAbs targeting CD41 (GPIIb), CD61 (GPIIIa), CD42a (GPIX), and CD42b (GPIbα). Different platelet phenotypes, e.g. activation, aggregation, procoagulant activity, and apoptosis were assessed by flow cytometry using markers such as CD62P, CD63, PAC-1, Annexin V, and TMRE. Additionally, ITP-Sera were incubated with washed platelets from healthy donors and platelet phenotypes were assessed by flow cytometry using the same markers as mentioned above.

Results: We observed heterogeneous effects of GP-specific mAbs on platelets. Two mAb-clones, one against CD41 and the other against CD61 induced platelet activation, procoagulant activity, and apoptosis after 1h of incubation. The mAb targeting CD41 further induced aggregation. Other mAbs targeting the same GPs but binding to a different epitope did only activate the platelets. Furthermore, some antibodies, targeting GPIIb/IIIa inhibited procoagulant phenotype formation and apoptosis in platelets. Two anti CD42b-mAb-clones triggered a procoagulant phenotype, and mainly apoptosis in platelets after 6h of incubation. Preliminary data from the ITP-cohort (n=103) showed that 16% of the ITP-Sera also induced platelet activation, apoptosis and a procoagulant phenotype in washed platelets.

Conclusion: These results highlight the diverse effects of GP-specific mAbs on platelet function, suggesting potential heterogeneity in ITP pathogenesis. Further investigations utilizing the ITP patient samples are planned to elucidate the clinical relevance of these findings.

Publication History

Article published online:
13 February 2025