Key Points rhTPO is a potentially effective and safe treatment option for ITP during pregnancy.

In addition to antiplatelet autoantibodies, CD8+ cytotoxic T lymphocytes (CTLs) play an important role in the increased platelet destruction in immune thrombocytopenia (ITP). Recent studies have highlighted that platelet desialylation leads to platelet clearance via hepatocyte asialoglycoprotein receptors (ASGPRs). Whether CD8+ T cells induce platelet desialylation in ITP remains unclear. Here, we investigated the cytotoxicity of CD8+ T cells towards platelets and platelet desialylation in ITP. We found that the desialylation of fresh platelets was significantly higher in ITP patients with positive cytotoxicity of CD8+ T cells than those without cytotoxicity and controls. In vitro, CD8+ T cells from ITP patients with positive cytotoxicity induced significant platelet desialylation, neuraminidase-1 expression on the platelet surface, and platelet phagocytosis by hepatocytes. To study platelet survival and clearance in vivo, CD61 knockout mice were immunized and their CD8+ splenocytes were used. Platelets co-cultured with these CD8+ splenocytes demonstrated decreased survival in the circulation and increased phagocytosis in the liver. Both neuraminidase inhibitor and ASGPRs competitor significantly improved platelet survival and abrogated platelet clearance caused by CD8+ splenocytes. These findings suggest that CD8+ T cells induce platelet desialylation and platelet clearance in the liver in ITP, which may be a novel mechanism of ITP.

To evaluate whether the serum thrombopoietin levels in pregnancy-associated immune thrombocytopenia (ITP) differ from those in gestational thrombocytopenia, and reveal the possibility of thrombopoietin serving as a marker for differential diagnosis. Serum thrombopoietin concentration was determined in ITP in pregnancy (n = 35), gestational thrombocytopenia (n = 31), healthy pregnancy (n = 32), age-matched nonpregnant ITP (n = 32) and nonpregnant healthy controls (n = 35) by ELISA. The serum thrombopoietin level of ITP in pregnancy (1283 ± 646 pg/mL) was significantly higher than gestational thrombocytopenia (187 ± 64 pg/mL) (P < 0.01), although the platelet counts of these two disorders may overlap. Twenty-nine of 35 patients with ITP in pregnancy had thrombopoietin values >500 pg/mL, whereas none of the gestational thrombocytopenia patients' thrombopoietin levels exceeded 500 pg/mL. In addition, ITP in pregnancy presented a markedly higher thrombopoietin level than nonpregnant ITP (88 ± 41 pg/mL) (P < 0.01), indicating that the pathogenesis of pregnant and nonpregnant ITP was different. Our findings suggest that measurement of serum thrombopoietin concentration provides valuable diagnostic information for differentiating ITP in pregnancy from gestational thrombocytopenia. Thrombopoietin represents a reliable marker for ITP in pregnancy.

BACKGROUND: Thrombopoietin (TPO), the major physiological regulator of platelet formation, binds to and activates TPO receptor on megakaryocytes, thereby promoting platelet production. Recombinant human thrombopoietin (rhTPO) is a novel therapeutic option for patients with immune thrombocytopenia (ITP) in China. However, the safety and efficacy of rhTPO for ITP patients in pregnancy is unclear. We report two ITP patients in pregnancy successfully treated with rhTPO. METHODS: Patient one, who was diagnosed as ITP according to the ASH guidelines two years ago, was a 28-year-old G2P1 pregnant woman, with a platelet count of 10×109/L and gingival bleeding. Before rhTPO therapy her TPO level was 231pg/mL. Patient two, who was diagnosed as ITP one year ago, was a 23-year-old primigravid woman with gross hematuria at 22-week gestation. Her platelet count was 19×109/L and the level of TPO in her plasma was 107pg/mL before rhTPO admission. Both patients failed to respond to prednisone (1mg/kg, 14d) and intravenous immunoglobulin (IVIG, 0.4g/kg, 5d). Due to persistent hemorrhage, they received rhTPO at a daily dose of 300U/kg daily (3SBIO Pharmaceutical Co., LTD, Shenyang, China). When their platelet counts rose above 50×109/L, maintenance therapy (rhTPO 300U/kg, every other day) started. This treatment protocol was approved by the Medical Ethical Committee of Qilu Hospital, Shandong University. RESULTS: The platelet count of patient one rose to 165×109/L after 10 days of rhTPO therapy. Then rhTPO maintenance therapy was applied. On the 39th week, she vaginally delivered a female infant with normal platelet count. The TPO level of the newborn was 127pg/mL. In the second case, after seven days of rhTPO therapy, her platelet count rose to 58×109/L, then we started rhTPO maintenance therapy. On the 40th week, she delivered a female infant without any signs of neonatal ITP. The level of TPO in the infant's plasma was 174pg/mL. Maintenance therapy continued after delivery with a dose of 300U/kg per week. The platelet counts of patients during treatment were illustrated in Figure 1. At a 1-month follow-up visit, no adverse events were observed in both infants. CONCLUSION: RhTPO may be an effective and fast-onset treatment for ITP in pregnancy. Future studies are necessary to investigate the safety and efficacy of rhTPO in this clinical setting. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Immune thrombocytopenia (ITP) is an acquired autoimmune disease characterized by transient or persistent decrease of platelet count. ITP is the most common cause of thrombocytopenia in early pregnancy. Patients with severe thrombocytopenia (platelet count < 20 ×109/L) are at risk of spontaneous bleeding, postpartum hemorrhage and placental abruption. The aim of this study is to determine the efficacy and the safety of recombinant human thrombopoietin (rhTPO) in the management of ITP in pregnancy. This study is registered at www.clinicaltrials.gov as NCT02391272. Patients at eight centers in China were enrolled. Enrollment criteria were pregnant women aged between 18 and 50, who failed to respond to first-line treatments of ITP and/or were refractory to platelet transfusion. Patients' platelet counts were below 30×109/L with bleeding manifestations. Gestational age of the patients was over 12 weeks. Informed consent was obtained from each patient in accordance with the Declaration of Helsinki. Thirty-one patients were enrolled into the study. The median age of the pregnant ITP patients was 26 years (range 19 - 39 years) and 90.6% (29/31) were primigravidae. The median gestational age was 24 weeks (12 - 38 weeks). The median baseline platelet count was 10×109/L (range 1 - 29×109/L). 74.2% (23/31) of these patients were diagnosed as ITP before pregnancy and 25.8 % (8/31) during pregnancy. All eligible participants received rhTPO at an initial dose of 300U/kg once daily subcutaneously for 14 days, 74.2% (23/31) of these patients responded to the initial 14-day rhTPO therapy, including 10 CR and 13 R. Eight patients were NR though their platelet counts rose mildly. The median platelet count of responder was 100×109/L (range from 30 to 250×109/L) at day 14. Then the responders received sequential maintenance therapy through the end of week 12 after delivery. To reduce the risk of thrombocytosis during maintenance, dose was tapered to 300U/kg every other day when platelet counts exceeded 50 ×109/L and treatment stopped when platelet counts above 100×109/L. Only one responder had a transient loss of response due to influenza. After dose adjustment of rhTPO from 300U/kg every other day to 300U/kg every day, the platelet count exceeded 50×109/L in the next visit. The platelet counts gradually dropped after withdrawal of rhTPO. The relapse free survival (platelet count at least 30×109/L) at week 4 and week 12 after withdrawal of rhTPO was 69.6%(16/23) and 21.7%(5/23), respectively (Figure 1). Safety and adverse events were evaluated in all 31 participants. rhTPO was well tolerated. Only mild previously reported adverse events were observed, including one case of dizziness, one of fatigue and one of pain at injection site. There were no new reported adverse events during the observation period and no adverse event-related study withdrawals. In all the 31 newborns, the median age of gestation was 39 weeks (range 36-40, 3 cases had age of gestation< 37 weeks); median birth weight was 3.1 kg (range 2.3-4.2kg, 2 had birth weight<2.5 kg) and the median platelet count at birth was 132×109/L (range 53-263 ×109/L, 9 with platelet<100×109/L) (Table 1). The incidence of premature labor, birth weight and platelet count were similar to previous study on pregnancy outcome in patients with ITP. None of the neonates had bleeding at birth. All 31 infants were evaluated for safety. No adverse effect on the development of newborns was observed during 6 months follow-up. One case of abdominal distension had been reported, which was supposed not associated with rhTPO. This study demonstrates that rhTPO is a potentially effective, safe and fast-onset treatment for ITP in pregnancy refractory to fist-line therapy and platelet transfusion. Our study paved the way for the clinical application of rhTPO and other thrombopoietic agents in the management of ITP in pregnancy. Disclosures No relevant conflicts of interest to declare.