Evolution of Platelet Function and Bleeding in Children and Adults with Chronic Immune Trombocytopenia on Romiplostim Treatment

INTRODUCTION. Platelets in immune thrombocytopenia (ITP) are not only reduced in quantity, but were also reported to have impaired function. There is evidence that this might be the mechanism behind the clinically established fact that severity of bleeding in ITP does not correllate well with the degree of thrombocytopenia. However, there are no established methods or guidelines to predict bleeding risks based on the platelet function tests, and little information about the effects of thrombopoietin mimetics on platelet function.

MATERIALS. We studied evolution of platelet function and clinical bleeding symptoms in 32 adult and 44 pediatric patients with chronic ITP 32 over six months of romiplostim treatment using flow cytometry and thrombelastography (TEG). As controls, samples of 20 healthy donors were used. The median age of adult patients was 62 (19-80 years), that of pediatric patients was 10 (1-18 years). All had previous history of treatment (one to three lines of therapy). Platelet function was characterized by flow cytometry before and after activation with SFLLRN plus collagen-related peptide. Levels of CD42b, CD61, CD62P, PAC1, annexin V binding, and mepacrine uptake and release were evaluated. Bleeding score was determined using a standardized consensus-based ITP-specific bleeding assessment tool developed by the International Working Group on ITP. Thrombelastography (TEG) was employed to characterize integral hemostasis function. Investigations were performed in accordance with the Declaration of Helsinki under a protocol approved by the NSPC CHOI Ethical Committee, and written informed consent was obtained from all donors and patients. Comparisons between groups were performed using Mann-Whitney non-parametric test.

RESULTS. Bleeding before the start of therapy was observed in 10 adults and 25 children. There was no significant difference in platelet count between the bleeding and non-bleeding adults (median 25 versus 23, respectively). For children, it was significantly (median 19 versus 83, p<0.05) lower for those with bleeding. Both adult's and children's platelets were significantly pre-activated by all parameters (PAC1, CD62p, annexin V) and increased in size (forward scatter, CD42b, CD61, mepacrine uptake) compared with normal ones. The TEG lag time parameters R and K were significantly increased in the ITP patients, while α angle and MA were decreased. Platelet pre-activation, size, and TEG parameter changes were significantly greater (by 50-100%) in the adult patients with bleeding phenotype compared with those who had no hemorrhage. Evaluation of their predictive capabilities with regard to bleeding risks using ROC analysis produced AUC values between 0.78 and 0.91, allowing design of a risk calculator. The results for children were similar when adjusted for the platelet count difference in the bleeding and non-bleeding groups. Romiplostim treatment led to a rapid (<1 month) decrease of procoagulant annexin V-positive platelets in the circulation to the normal level, followed by a more gradual normalization of platelet size and decrease of PAC1-positive platelets over several months. In some adult and pediatric patients, disappearance of clinical bleeding syndromes on romiplostim was associated with normalization of platelet function parameters in the absence of platelet count increase.

CONCLUSIONS. Our data indicate that: a) platelets in chronic ITP are pre-activated, large and slowly reacting; b) these changes are associated with bleeding independently of platelet count, and can be used for risk stratification; c) romiplostim treatment decreases platelet size and pre-activation.