Defective Functional Activity of Platelets in Children with Severe Chronic Immune Thrombocytopenia and Its Correction By Romiplostim Treatment

BACKGROUND: Immune thrombocytopenic purpura (ITP) is an autoimmune disease characterized by antibody-mediated platelet destruction, which together with suboptimal platelet production lead to thrombocytopenia. Children with low platelet counts due to ITP are at an increased risk of bleeding, which is not always directly correlated to the severity of thrombocytopenia. Relationship of platelet functional activity and bleeding risk in ITP is poorly investigated. There are conflicting reports on whether platelets in adult ITP patients differ from normal ones, and no data for children are available. Information about the thrombopoietin receptor agonists (eltrombopag and romiplostim, which are increasingly used to treat ITP) effect on platelet quality is also scarce. Data for eltrombopag are inconclusive, and no studies for romiplostim are known.

OBJECTIVES: To investigate platelet functional activity before and during the romiplostim treatment of ITP in children.

METHODS: The study group consisted of patients from 4 to 11 years old (n=8) with chronic ITP; the control group included healthy donors (n=12). All patients initially had low platelet counts (<15*10⁹/L) and did not respond to the first- and second- line therapy. Blood samples were collected before treatment with romiplastim and then monthly until the onset of clinical remission. Platelets in whole blood were either left intact or activated with collagen-related peptide (0.18 mkg/ml) and thrombin receptor activating peptide (12.5 mkM), labelled and analysed by flow cytometry. To characterize platelet functions, we used fluorescencently labeled antibodies against glycoprotein Ib (CD42b), total and active integrin α_IIbβ₃ (CD61 and PAC-1), and P-selectin (CD62P); dense granule release was assessed using platelet loading with mepacrine, and procoagulant activity was determined with a phosphatidylserine marker annexin V.

RESULTS: All investigated parameters of platelet function in children with severe chronic ITP before treatment were greatly impaired. CD42b and CD61 were up to several-fold lower the normal values, while phosphatidylserine exposure, dense granule release and integrin activation upon activation were decreased by at least an order of magnitude. Romiplostim treatment improved platelet parameters, although not always to the normal level. Five patients out of eight responded partially or completely to the romiplostim therapy. The clinical response (relief of hemorrhagic manifestations) correlated well with improvement of the functional state of platelets, in one case, even without significant platelet count increase.

CONCLUSIONS. Our data suggest essential revision of the pathophysiology of severe chronic ITP in children: their platelets are not only small in their number, but also appear to have severe defects of all major functions. The mechanism of action of romiplostim in these patients might also be in some need of revision, as it seems to greatly improve not only quantity, but also quality of platelets. Our data demonstrate the important role of monitoring platelet functional activity in addition to platelet count.