Advances in Diagnosis and Treatments for Immune Thrombocytopenia

Abstract

Immune thrombocytopenia (ITP) is an acquired hemorrhagic condition characterized by the accelerated clearance of platelets caused by antiplatelet autoantibodies. A platelet count in peripheral blood <100 × 10⁹/L is the most important criterion for the diagnosis of ITP. However, the platelet count is not the sole diagnostic criterion, and the diagnosis of ITP is dependent on additional findings. ITP can be classified into three types, namely, acute, subchronic, and persistent, based on disease duration. Conventional therapy includes corticosteroids, intravenous immunoglobulin, splenectomy, and watch-and-wait. Second-line treatments for ITP include immunosuppressive therapy [eg, anti-CD20 (rituximab)], with international guidelines, including rituximab as a second-line option. The most recently licensed drugs for ITP are the thrombopoietin receptor agonists (TRAs), such as romiplostim and eltrombopag. TRAs are associated with increased platelet counts and reductions in the number of bleeding events. TRAs are usually considered safe, effective treatments for patients with chronic ITP at risk of bleeding after failure of first-line therapies. Due to the high costs of TRAs, however, it is unclear if patients prefer these agents. In addition, some new agents are under development now. This manuscript summarizes the pathophysiology, diagnosis, and treatment of ITP. The goal of all treatment strategies for ITP is to achieve a platelet count that is associated with adequate hemostasis, rather than a normal platelet count. The decision to treat should be based on the bleeding severity, bleeding risk, activity level, likely side effects of treatment, and patient preferences.

Keywords

ITP anti-GPIIb/IIIa antibody bleeding risk corticosteroid splenectomy immune suppressive therapy thrombopoietin receptor agonist new agent

Introduction

Immune thrombocytopenia (ITP) is an acquired hemorrhagic condition characterized by the accelerated clearance of platelets caused by antiplatelet autoantibodies such as anti-glycoprotein (GP) IIb/IIIa.^1,2 Autoreactive B- and T-cells have important roles in antibody generation in ITP patients.³ GPIIb/IIIa is one of the major target antigens recognized by platelet-reactive CD4 ⁺ T-cells.^4–6 A platelet count in peripheral blood of <100 × 10⁹/L is the most important criterion for the diagnosis of ITP.^1,2 However, the platelet count is not the sole diagnostic criterion, and the diagnosis of ITP is dependent on additional findings. ITP can be classified into three types, namely, acute, subchronic, and persistent, based on disease duration.² ITP in adults tends to be subchronic/persistent, relapsing, and very often, refractory to treatment.⁷

Conventional therapy includes corticosteroids, intravenous immunoglobulin (IVIg), splenectomy, and watch-and-wait.^1,2 Initially, 70%-80% of patients respond to corticosteroids, and 10%-30% attain durable remission.² Splenectomy is avoided in young children because of infection risk and because the prevalence of spontaneous resolution of ITP is high.⁸ Second-line treatments for ITP include immunosuppressive therapy [eg, anti-CD20 (rituximab)],⁹ with international guidelines including rituximab as a second-line option.^10,11

In contrast to immunosuppressive therapy, physiological therapy was recently introduced.¹² These drugs include thrombopoietin receptor agonists (TRAs), such as romiplostim and eltrombopag.^13–16 TRAs are associated with increased platelet counts and a reduced number of bleeding events. TRAs are usually considered as a safe and an effective treatment for patients with chronic ITP at risk of bleeding after failure of first- or second-line therapies.

Herein, we discuss the accepted therapies available for ITP, as well as potential for the use of alternative drugs and novel treatments. We also describe the pathogenesis and clinical manifestations involved in the development and progression of ITP.

Assessment of the Literature

Reference articles were identified by an internet search of the PubMed database. The search key words were idiopathic thrombocytopenic purpura and immune thrombocytopenia.

Pathophysiology

The most commonly identified antigenic targets of ITP autoantibodies are GPIIb/IIIa and GPIb/IX; a considerable number of ITP patients have antibodies directed to multiple platelet antigens.¹⁷ Several lines of evidence link T-cells to the pathogenesis of ITP. The T-cell changes implicated include excessive activation and proliferation of platelet antigen-reactive cytotoxic T-cells, production of abnormal helper T-cells (Th), and abnormalities in the number and function of regulatory T-cells (Tregs).^18–21 Platelet-reactive CD4⁺ T-cells have been found in the blood samples of ITP patients, with the major target antigen being GPIIb/IIIa.⁴

CD4⁺ Th cells are divided into four main subsets (1, 2, 3, and 17) according to the type of cytokines they produce.^22,23 Th1 cells are involved mainly in macrophage activation and production of interferon gamma and interleukin (IL)-2. Th2 cells are involved mainly in secretion of IL-4 and IL-10, the humoral immune response, and inactivation of several macrophage functions.²² Th3 cells generate transforming growth factor-β1. Th17 cells generate IL-17 and modulate immune responses.^22,24 Several studies have found evidence supporting polarization of Th cells in the immune response in patients with chronic ITP.^21–30 Those reports have suggested that a peripheral regulatory mechanism controlling these GP-reactive T-cells is necessary to prevent autoimmunity (Fig. 1).^20,31

Figure 1.

Pathogenesis of ITP. Activated macrophages in the reticuloendothelial system transfer the antigenic information such as GPIIb/IIIa peptide to autoreactive CD4⁺ T-cells. autoreactive CD4⁺ T-cells and antibody-producing B-cells maintain antiplatelet autoantibody production in ITP patients. there exists a continuous pathogenic loop in ITP. treg in ITP has less functional strength than it used to.

CD4⁺ Tregs have critical roles in the maintenance of peripheral tolerance by suppression of the activation and proliferation of many cell types.^32,33 Tregs are divided into two subtypes, namely, naturally occurring Tregs and induced Tregs, based on their ontogeny and mode of action. Naturally occurring Tregs are generated in the thymus gland and constitutively express cytotoxic T-lymphocyte-associated antigen-4 and transcription factor forkhead-box p3 (Foxp3).²⁰ The previously mentioned Th3 cells are Foxp3 negative, but these cells are included in Tregs.³⁴ A reduction in Treg function has been reported in several autoimmune diseases.³³ Some studies in ITP patients have also shown reduced levels of Foxp3 and abnormal function of Tregs.^35–37

The major physiological role of B-cells is antibody production.^38,39 The number of circulating B-cells secreting anti-GPIIb/IIIa antibodies has been reported to be increased in patients with ITP.^40,41 The dysregulation of B-cell development has also been associated with ITP.²⁸ Serum concentrations of B-cell activating factor were shown to be significantly increased in patients with active ITP.⁴² B-cells are also efficient at presenting antigens, including low concentrations of antigens, to T-cells.^28,39 These findings suggest that a rational approach to ITP treatment could involve B-cell depletion, such as with rituximab.^38,39

The spleen plays an important role in pathways leading to thrombocytopenia. This organ is the primary site of antibody production, as well as being important in the clearance of antibody-coated platelets (Fig. 2).^43–46

Figure 2.

Mechanisms leading to the thrombocytopenia. Human macrophages express Fc receptor that binds IgG specifically. Liver and spleen are dominant organs for the clearance of IgG-coated platelets. There is also direct cytotoxicity by complement such as C5b-9.

Human macrophages express several Fc receptors that bind IgG specifically.⁴⁷ There are two classes of Fc receptors: high affinity (FcγRI) and low affinity (FcγRII, FcγRIII). Removal of opsonized platelets is dependent on low-affinity receptors.^48,49

Clinical Features

Findings such as attack of fever, bone or joint pain, infection with human immunodeficiency virus, morphologic abnormalities of the skeleton or soft tissue, nonpetechial rash, lymphadenopathy, abnormal hemoglobin level, white blood cell count, abnormal white cell morphology, a family history of low platelets, or easy bruising are not typical of ITP and should prompt additional diagnostics such as bone marrow evaluation to rule out other disorders.^2,10,11

Most children have bruising or purpura, although they are asymptomatic.⁵⁰ There may be tiny red spots or larger areas of purpura. Pediatric ITP often occurs ~2 weeks after a viral infection. Most children with ITP recover within six weeks. ITP is acute in 70%-80% of children and chronic in 20%-30%.

In adults,⁵¹ ITP arises gradually and does not usually follow a viral illness. Symptoms are highly variable: no symptoms, purpura, mild bleeding, or severe bleeding. Unlike pediatric ITP, most adults with ITP continue to have a low platelet count indefinitely (chronic ITP). However, the differences between ITP in adults and children are not as clear as suggested here.

Laboratory Measurements

A presumptive diagnosis of ITP can be made if patient history, physical examination, complete blood count, and examination of peripheral blood smears do not suggest other causes of thrombocytopenia (Table 1). There is no gold standard test that can reliably establish the diagnosis. It remains unclear whether bone marrow examination is necessary for all patients with ITP.¹⁰ Assays for antibodies for specific platelet GPs are not recommended routinely because levels of platelet-associated IgG are elevated in ITP and non-ITP.¹⁷ However, existence of anti-GPIIb/IIIa antibody may be useful for the diagnosis of ITP (Table 1).⁴⁰

Table 1.

Diagnosis of ITP.

Thrombocytopenia (<100 × 10⁹/L) without morphologic evidence for dysplasia in the peripheral blood film

The presence of any three or more, including at least one of ③, ④, and ⑤, of the following laboratory findings:

①

Absence of anemia

②

Normal leukocyte count

③

Increased anti-GPllb/llla antibody producing B cell frequency

④

Increased platelet-associated anti-GPllb/llla antibody level

⑤

Elevated percentage of reticulated platelets

⑥

Normal or slightly increased plasma TPO level (<300 pg/ml)

the following diseases or conditions are excluded

Drug- or radiation-induced TP, aplastic anemia, MDS, PNH, SLE, leukemia, malignant lymphoma, DIC, TTP, sepsis, sarcoidosis, virus infection, Bernard-Soulier syndrome, Wiskott-Aldrich syndrome, May-Hegllin abnormality, Kasabach-Merritt syndrome

Abbreviations: GP, glycoprotein; TPO, thrombopoietin; TP, thrombocytopenia; MDS, myelodisplastic syndrome; PNH, paroxysmal night hemoglobinemia; SLE, systemic lupus erythematosus; DIC, disseminated intravascular coagulation; TTP, thrombotic thrombocytopenia.

Differential Diagnoses

The diagnosis of primary ITP is one of exclusions. Thrombocytopenia can be caused by systemic disease, infection, drugs, or primary hematologic disorders.¹⁰ Bleeding after surgery, dentistry, and trauma must be considered when estimating the possible duration of chronic thrombocytopenia or an alternative bleeding disorder. Differential diagnoses can classify patients into two groups. One group includes patients with ITP, such as systemic lupus erythematosus, whereas the other includes patients with non-ITP, such as aplastic anemia and leukemia. Although aplastic anemia may present initially as isolated thrombocytopenia, it is followed within days, weeks, months, or even years by anemia and leukopenia. Moreover, leukemia never presents as isolated thrombocytopenia. International consensus documents and guidelines should be consulted regarding differences between primary ITP and the other conditions.^10,11

Treatment

Initial Management

Determination of the threshold for a minimum platelet count or specific age at which a typical patient with ITP should be treated is difficult.¹¹ The goal of all treatment strategies for ITP is to achieve a platelet count that is associated with adequate hemostasis, rather than a normal platelet count.¹¹ Therefore, if a bleeding symptom is not found, treatment may not be needed. Patients should be treated with platelet-enhancing agents if the platelet count is <30 × 10⁹/L and mucosal bleeding has started, although a threshold of 30 × 10⁹/L may not be suitable for children. Treatment may also be appropriate if: follow-up cannot be assured; there are concerns regarding the activity or risk of bleeding; and there is a need for procedures associated with bleeding risk. In conclusion, the decision to treat should be based on bleeding severity, bleeding risk, activity level, likely side effects of treatment, and patient preferences.^51–54

First-Line Therapy

Corticosteroids

Corticosteroids remain the most commonly used first-line therapy for ITP. Treatment is initiated with prednisolone or prednisone (1-2 mg/kg per day, p.o., single or divided doses). Approximately, two-thirds of patients achieve a complete or partial response with corticosteroids at these doses, with most responses occurring within the first week of treatment.⁵⁵ In general, corticosteroids are initially effective. A study comparing high- and low-dose corticosteroids in adult patients found that low-dose therapy was initially more effective, although long-term effects have not been determined.^11,56 However, because of their side effects, long-term corticosteroids should be avoided in children with acute ITP.¹¹

IVIg

IVIg therapy is administered to patients requiring rapid or urgent elevation of platelet count (eg, intraoperative or life-threatening bleeding). IVIg increases the platelet count in 70%-80% of treated patients, often within days. Several IVIg regimens have been employed, but many hematologists prefer the convenience of a 1 g/kg/day infusion for 1-2 days.⁵⁷ IVIg also affects humoral and cellular immunity by influencing the expression and activity of Fc receptors.⁵⁸ Although its incidence is low, intracerebral hemorrhage remains a severe side effect of IVIg.¹¹

Anti-D Immunoglobulin

Anti-D immunoglobulin (anti-D) binds to Rh(D) antigen on erythrocytes, thereby leading to clearance of antibody-coated cells and inhibiting the clearance of opsonized platelets by the reticuloendothelial system.⁵⁹ Therefore, anti-D is effective only in RhD positive individuals. Anti-D has been reported effective in approximately 50%-70% of patients treated with this agent.⁶⁰ If anti-D is chosen as therapy, care must be taken because of the risk of severe hemolysis that has been reported with some products.¹¹ However, subcutaneous administration abolishes the risk of hemolysis in most patients. Avoidance of use of anti-D in patients with a positive direct antiglobulin test has been recommended.⁶¹ In addition, fatal renal failure and disseminated intravascular coagulation are reported as side effects of anti-D.^11,62

Management of Helicobacter pylori-associated ITP. Secondary ITP can occur in patients with H. pylori infection.¹⁰ The eradication therapy should be administered in patients who are found to have H. pylori infection.¹¹

Second-Line Therapy

Splenectomy

Splenectomy is considered by some scholars to be the gold standard treatment for ITP. In particular, it is recommended as a second-line treatment for adults unresponsive for corticosteroid therapy. Initially, 65%-70% of patients show a complete response, whereas 60%-70% show a long-term response.^45,63 Types of splenectomy are open splenectomy and laparoscopic splenectomy, but the latter is associated with fewer complications than the former.⁶⁴ Splenectomy-related complications include infection, bleeding, thrombosis, and relapse.⁶⁵ In particular, the risk of infection is the major cause of mortality after splenectomy.⁶⁶

Rituximab

Rituximab is a chimeric monoclonal antibody that targets CD20 B-cell surface antigen.^9,26,67,68 Several studies have reported significant responses before and after splenectomy with the use of rituximab.^67–71 Despite targeting CD20 on B-cells, the mechanism of action of rituximab may involve more complex immunologic modulation.⁷² In one report, successful therapy correlated with normalization of distribution of T-cell subsets,²⁶ and in another report, it correlated with reappearance of normal numbers and function of Tregs.⁷³ Adverse effects of rituximab include infusion reactions, serum sickness, and cardiac arrhythmia. Rituximab can also be used off-license as a second-line option in certain types of refractory ITP, but long-term safety is not known.⁷⁴ In addition, a recent meta-analysis could not find that rituximab was effective.⁷⁵

Third-Line Therapy

TRAs

Thrombocytopenia may result not only from platelet destruction but also from antibody-mediated damage to megakaryocytes.⁴⁵ Thus, ITP in some patients may be due to impaired production of platelets.⁴⁵ Therefore, recombinant human TPO (rhTPO) has been administered in some ITP patients,^76,77 and as a result, the improvement in thrombocytopenia has been remarkable. However, all clinical trials with rhTPO were stopped after development of antibodies against rhTPO was observed in healthy volunteers.⁷⁸ Since then, the development of TRAs has progressed. Two of these new TRAs, such as romiplostim and eltrombopag, have been licensed for use in patients with chronic ITP. Binding of TRAs to the thrombopoietin receptor results in activation of intracellular signaling pathways such as JAK-STAT and mitogen-activated protein kinase (MAPK) that lead to increased production of platelets.⁷⁹ Although TRAs may markedly improve thrombocytopenia, their safety is questionable, as shown by an increasing list of severe side effects.⁷⁵

Romiplostim

Romiplostim is a recombinant fusion protein peptibody composed of two IgG1 constant regions (Fc fragments) linked to a peptide domain containing four binding sites for the thrombopoietin receptor.⁸⁰ Some evidence on the use of romiplostim in adults with ITP suggests that it increases the platelet count and reduces bleeding.^81–84 Romiplostim-related adverse effects have been mild-moderate and not led to treatment cessation.^15,85 Rare adverse effects include mild-to-moderate postinjection headache, fatigue, and arthralgia.^81,85 Serious adverse events that continue to be under investigation include increased reticulum cells in the bone marrow,⁸⁶ increased proliferation of leukemic blasts,⁸⁷ and thrombosis.⁸⁸ However, the frequency of these adverse events appears to be low.

Eltrombopag

Eltrombopag is a small nonpeptide molecule that binds to the thrombopoietin receptor via its trans-membrane domain and activates JAK-STAT and MAPK intracellular pathways to increase platelet production.^79,89,90 Once-daily oral administration of eltrombopag was found to be effective and safe in patients with chronic ITP.^13,91,92 Patients with chronic ITP and platelet count <30,000/μL received eltrombopag (50 mg/day) or standard of care for six months in a double-blind phase III study.¹⁶ Prevalence of adverse events in adult ITP was 73% for patients receiving eltrombopag compared with 25% for patients receiving placebo.⁹² All adverse events were mild-moderate in severity, and increases in the prevalence of nasopharyngitis and level of alanine amino-transferase were the most frequently reported adverse events in the eltrombopag group.⁹² The most common adverse events of eltrombopag in childhood ITP were headache, infection of the upper respiratory tract, and nasopharyngitis.⁹³ Long-term safety and efficacy of eltrombopag has been investigated in the EXTEND study, which enrolled 301 patients, with 84 and 28 patients being treated for ≥3 and ≥4 years, respectively.⁹⁴

New Agents

Anti-CD40 Ligand Monoclonal Antibody

The GPIIb/IIIa-reactive T-cell is a target for therapeutic strategies involving selective suppression of the pathogenic autoimmune response in ITP patients. One strategy is disruption of a costimulatory signal by blocking the interaction between CD40 on antigen-presenting cells and CD40 ligand (CD40L) on activated CD4⁺ T-cells; this interaction is essential for T-cell priming and the T-cell-dependent humoral immune response.⁹⁵ Therefore, anti-CD40L antibody could be effective against ITP.⁹⁶

IDEC-131 is a humanized monoclonal antibody against human CD40L. It binds to the trimer of CD40L on T-cells with high specificity and activity, thereby preventing CD40 signaling.⁹⁷ Kuwana et al⁹⁸ reported that CD40/CD40L blockade therapy by IDEC-131 could be effective against refractory ITP through selective suppression of autoreactive T- and B-cells to platelet antigens. Another trial with anti-CD40L monoclonal antibody showed an overall response of 24% in 46 refractory patients with ITP.⁹⁹ Anti-CD40L antibody could become a therapeutic strategy against ITP.^98–100

Anti-CD20 Monoclonal Antibody

Veltuzumab is a humanized anti-CD20 monoclonal antibody with complementarity-determining regions identical to rituximab. It has unique characteristics in terms of significantly improved complement-dependent cytotoxicity.¹⁰¹ Veltuzumab is active at a fraction of the conventional clinical dose of rituximab, and survival studies in lymphoma models have shown significantly higher potency of this antibody over rituximab.¹⁰¹ Additional studies are needed to assess the efficacy and safety of anti-CD20 antibody as first-line therapy in adults with ITP.¹⁰² Low-dose veltuzumab (s.c.) appears to be convenient, well-tolerated, and with promising effects against relapsed ITP.¹⁰³

Spleen Tyrosine Kinase Inhibitor

Spleen tyrosine kinase (Syk) is a cytoplasmic protein-tyrosine kinase that associates with the Fcγ receptor in various inflammatory cells.¹⁰⁴ Syk can couple immune-cell receptors to intracellular signaling pathways that regulate cellular responses to extracellular antigens and anti-gen-Ig complexes of particular importance to the initiation of inflammatory responses.¹⁰⁵ ITP patients have accelerated clearance of circulating IgG-coated platelets via Fcγ receptor-bearing macrophages in the spleen and liver. A pilot study found that the Syk inhibitor R788 restored platelet counts to ≥50% in adults with refractory ITP.¹⁰⁶ In addition, adverse events were gastrointestinal (diarrhea, nausea, emesis).¹⁰⁶ However, the progressive condition with this drug is unclear now.

Other New Agents

Trials of several drugs for the treatment of ITP are now underway. The list of new drugs/molecules and studies related to these drugs is included in Table 2.²⁴,⁷⁴,⁷⁷,⁷⁸,⁸⁹,⁹⁰,^100–103,^107–116

Table 2.

New drugs of ITP.

	REFERENCES
1. Nonspecific immunosupressant
① Cyclosporin A (CyA)	107
② Etanercept (soluble TNF receptor)	108
③ MMF (inhibitory effect on T-cell)	109
2. Molecule targetting therapy
① Anti-CD20 (rituximab, veltuzumab)	24, 99, 101-103, 110
② Anti-CD40L (IDEC-131; CD154)	95-100
③ Anti-CD52 (alemtuzumab; Campath-1 H)	111
④ Anti-CD80/86 (abatacept; CTLA4-lg)	112
⑤ Anti-IL2 receptor (daclizumab)	113
3. Platelet increasing therapy
① rhTPO (PEG-rHuMGDF)	74, 114
② TRAs (eltrombopag, romiplostim)	77, 78, 89, 90
③ New TRA (avatrombopag)	115
④ Other (NIP-004)	116

Abbreviation: MMF, mycophenolate mofetil.

Conclusion

ITP is an acquired hemorrhagic condition characterized by the accelerated clearance of platelets caused by antiplatelet autoantibodies such as anti-GP IIb/IIIa. The platelet count in peripheral blood is <100 × 10⁹/L. ITP can be classified into three types, namely, acute, subchronic, and persistent, based on disease duration. ITP in adults tends to be sub-chronic/persistent, relapsing, and very often, refractory to treatment. Conventional therapy includes corticosteroids, IVIg, splenectomy, and watch-and-wait. Initially, 70%-80% of patients respond to corticosteroids, and 10%-30% attain durable remission. Splenectomy is avoided in young children because of infection risk and the high prevalence of spontaneous resolution of ITP. Second-line treatments for ITP include immunosuppressive therapy (eg, rituximab). Third-line therapies include TRAs, such as romiplostim and eltrombopag. TRAs are associated with increased platelet counts and reductions in the number of bleeding events. TRAs are considered as a safe and an effective treatment for patients with chronic ITP at risk of bleeding after failure of first- or second-line therapies. Determination of a threshold minimum platelet count or specific age at which a typical patient with ITP should be treated is difficult. The goal of all treatment strategies for ITP is to achieve a platelet count that is associated with adequate hemostasis, rather than a normal platelet count. Therefore, if bleeding symptoms are not found, treatment may not be needed. Patients should be treated with platelet-enhancing agents if the platelet count is <30 × 10⁹/L and mucosal bleeding has started, although a threshold of 30 × 10⁹/L may not be suitable for children. Treatment may also be appropriate if follow-up cannot be assured, if there are concerns for bleeding due to high levels of activity, or if there is a need for procedures associated with bleeding risk.

New therapies and recommendations have emerged in the last decade. However, deciding who should be treated with which treatment option and for how long is not known. The decision to treat should be based on bleeding severity, bleeding risk, activity level, likely side effects of treatment, and patient preference. A flowchart detailing the management and therapeutic options for ITP is illustrated in Figure 3. It can provide guidance, but should not replace clinical judgment.^2,10,11,75

Figure 3.

Treatment algorithm for ITP.

Author Contributions

Conceived the concepts: SN. Wrote the first draft of the manuscript: SN. Developed the structure and arguments for the paper: SN. Made critical revisions: SN. The author reviewed and approved of the final manuscript.

References

Cines

D.B.

, Blanchette

V.S.

Immune thrombocytopenic purpura.

N Engl J Med. 2002; 346: 995–1008.

Rodeghiero

, Stasi

, Gernsheimer

. Standardization of terminology, definitions and outcome criteria in immune thrombocytopenic purpura of adults and children: report from an international working group. Blood. 2009; 113: 2386–2393.

Semple

J.W.

, Milev

, Cosgrave

. Differences in serum cytokines levels in acute and chronic autoimmune thrombocytopenic purpura: relationship to platelet phenotype and antiplatelet T-cell reactivity. Blood. 1996; 87: 4245–4254.

Kuwana

, Kaburaki

, Ikeda

Autoreactive T cells to platelet GPIIb-IIIa in immune thrombocytopenic purpura: role in production of antiplatelet autoanti-body.

J Clin Invest. 1998; 102: 1393–1402.

Nomura

, Matsuzaki

, Ozaki

. Clinical significance of HLA-DRB1^*0410 in Japanese patients with idiopathic thrombocytopenic purpura. Blood. 1998; 91: 3616–3622.

Kuwana

, Kaburaki

, Kitasato

. Immunodominant epitopes on gly-coprotein IIb-IIIa recognized by autoreactive T cells in patients with immune thrombocytopenic purpura. Blood. 2001; 98: 130–139.

Michel

, Rauzy

O.B.

, Thoraval

F.R.

. Characteristics and outcome of immune thrombocytopenia in elderly: results from a single center case-controlled study. Am J Hematol. 2011; 86: 980–984.

Labarque

, Van Geet

Clinical practice: immune thrombocytopenia in paediatrics.

Eur J Pediatr. 2014; 173: 163–172.

Stasi

, Pagano

, Stipa

, Amadori

Rituximab chimeric anti-CD20 monoclonal antibody treatment for adults with chronic idiopathic thrombocytopenic purpura.

Blood. 2001; 98: 952–957.

10.

Provan

, Stasi

, Newland

A.C.

. International consensus report on the investigation and management of primary immune thrombocytopenia. Blood. 2010; 115: 168–175.

11.

Neunert

, Lim

, Crowther

. The American Society of Hematology 2011 evidence-based practice guideline for immune thrombocytopenia. Blood. 2011; 117: 4190–4207.

12.

Ozkan

M.C.

, Sahin

, Saydam

Immune thrombocytopenic purpura: new biological therapy of an old disease.

Curr Med Chem. 2015; 22: 1956–1962.

13.

Bussel

J.B.

, Cheng

, Saleh

M.N.

. Eltrombopag for the treatment of chronic idiopathic thrombocytopenic purpura. N Engl J Med. 2007; 357: 2237–2247.

14.

Bussel

J.B.

, Kuter

D.J.

, Pullarkat

, Lyons

R.M.

, Guo

, Nichol

J.L.

Safety and efficacy of long-term treatment with romiplostim in thrombocytopenic patients with chronic ITP.

Blood. 2009; 113: 2161–2171.

15.

Kuter

D.J.

, Rummel

, Roccia

. Romiplostim or standard of care in patients with immune thrombocytopenia. N Engl J Med. 2010; 363: 1889–1899.

16.

Cheng

, Saleh

M.N.

, Marcher

. Eltrombopag for management of chronic immune thrombocytopenia (RAISE): a 6-month, randomized, phase 3 study. Lancet. 2011; 377: 393–402.

17.

McMillan

, Wang

, Tani

Prospective evaluation of the immunobead assay for the diagnosis of adult chronic immune thrombocytopenic purpura (ITP).

J Thromb Haemost. 2003; 1: 485–491.

18.

Olsson

, Andersson

P.O.

, Jernás

. T-cell-mediated cytotoxicity toward platelets in chronic idiopathic thrombocytopenic purpura. Nat Med. 2003; 9: 1123–1124.

19.

Wang

T.T.

, Zhao

, Ren

. Type 1 and type 2 T-cell profiles in idiopathic thrombocytopenic purpura. Haematologica. 2005; 90: 914–923.

20.

Zhang

X.L.

, Peng

, Sun

J.Z.

. De novo induction of platelet-specific CD4⁺CD25⁺ regulatory T cells from CD4⁺CD25^– cells in patients with idio-pathic thrombocytopenic purpura. Blood. 2009; 113: 2568–2577.

21.

, Zhang

, Peng

, Hou

T cell immune abnormalities in immune throm-bocytopenia.

J Hematol Oncol. 2014; 7: 72–77.

22.

Mosmann

T.R.

, Coffman

R.L.

Th1 and Th2 cells: different patterns of lympho-kine secretion lead to different functional properties.

Annu Rev Immunol. 1989; 7: 145–173.

23.

Miossec

, Korn

, Kuchroo

V.K.

Mechanisms of disease: interleukin-17 and type 17 helper T cells.

N Engl J Med. 2009; 361: 888–898.

24.

Carrier

, Yuan

, Kuchroo

V.K.

, Weiner

H.L.

Th3 cells in peripheral tolerance. II. TGF-beta-transgenic Th3 cells rescue IL-2-deficient mice from autoimmunity.

J Immunol. 2007; 178: 172–178.

25.

Panitsas

F.P.

, Theodoropoulou

, Kouraklis

. Adult chronic idiopathic thrombocytopenic purpura (ITP) is the manifestation of a type-1 polarized immune response. Blood. 2004; 103: 2645–2647.

26.

Stasi

, Del Poeta

, Stipa

. Response to B-cell depleting therapy with rituximab reverts the abnormalities of T-cell subsets in patients with idiopathic thrombocytopenic purpura. Blood. 2007; 110: 2924–2930.

27.

Wang

J.D.

, Chang

T.K.

, Lin

H.K.

, Huang

F.L.

, Wang

C.J.

, Lee

H.J.

Reduced expression of transforming growth factor-p1 and correlated elevation of interleukin-17 and interferon-g in pediatric patients with chronic primary immune thrombocytopenia (ITP).

Pediatr Blood Cancer. 2011; 57: 636–640.

28.

McKenzie

C.G.J.

, Guo

, Freedman

, Semple

J.W.

Cellular immune dysfunction in immune thrombocytopenia (ITP).

Br J Haematol. 2013; 163: 10–23.

29.

Shan

N.N.

, Hu

, Hou

. Decreased Tim-3 and its correlation with Th1 cells in patients with immune thrombocytopenia. Thromb Res. 2014; 133: 52–56.

30.

Lyu

, Li

, Hao

. Elevated semaphoring 5A correlated with Th1 polarization in patients with chronic immune thrombocytopenia. Thromb Res. 2015; 136: 859–864.

31.

Bao

, Bussel

J.B.

, Heck

. Improved regulatory T-cell activity in patients with chronic immune thrombocytopenia treated with thrombopoietic agents. Blood. 2010; 116: 4639–4645.

32.

Sakaguchi

Naturally arising CD4+ regulatory T cells for immunologic self-tolerance and negative control of immune responses.

Annu Rev Immunol. 2004; 22: 531–562.

33.

Von Boehmer

Mechanisms of suppression by suppressor T cells.

Nat Immunol. 2005; 6: 338–344.

34.

Wing

, Fehérvári

, Sakaguchi

Emerging possibilities in the development and function of regulatory T cells.

Int Immunol. 2006; 18: 991–1000.

35.

, Heck

, Patel

. Defective circulating CD25 regulatory T cells in patients with chronic immune thrombocytopenic purpura. Blood. 2008; 112: 1325–1328.

36.

Ling

, Cao

, Yu

, Ruan

Circulating dendritic cells subsets and CD4⁺ Foxp3⁺ regulatory T cells in adult patients with chronic ITP before and after treatment with high-dose dexamethasone.

Eur J Haematol. 2007; 79: 310–316.

37.

Liu

, Zhao

, Poon

M.C.

. Abnormality of CD4(+)CD25(+) regulatory T cells in idiopathic thrombocytopenic purpura. Eur J Haematol. 2007; 78: 139–143.

38.

Stasi

Rituximab in autoimmune hematologic diseases: not just a matter of B cells.

Semin Hematol. 2010; 47: 170–179.

39.

Godeau

B-cell depletion in immune thrombocytopenia.

Semin Hematol. 2013; 50(suppl 1): 575–582.

40.

Kuwana

, Kurata

, Fujimura

. Preliminary laboratory based diagnostic criteria for immune thrombocytopenic purpura: evaluation by multi-center prospective study. J Thromb Haemost. 2006; 4: 1936–1943.

41.

Chen

J.F.

, Yang

L.H.

, Chang

L.X.

, Feng

J.J.

, Liu

J.Q.

The clinical significance of circulating B cells secreting anti-glycoprotein IIb/IIIa antibody and platelet gly-coprotein IIb/IIIa in patients with primary immune thrombocytopenia.

Hematology. 2012; 5: 283–290.

42.

Emmerich

, Bal

, Barakat

. High-level serum B cell activating factor and promoter polymorphisms in patients with idiopathic thrombocytopenic purpura. Br J Haematol. 2007; 2: 309–314.

43.

Louwes

, Zeinali Lathori

O.A.

, Vellenga

, de Wolf

J.T.

Platelet kinetic studies in patients with idiopathic thrombocytopenic purpura.

Am J Med. 1999; 106: 430–434.

44.

Kuwana

, Okazaki

, Kaburaki

, Kawakami

, Ikeda

Spleen is a primary site for activation of platelet-reactive T and B cells in patients with immune thrombocytopenic purpura.

J Immunol. 2002; 168: 3675–3682.

45.

Stasi

Pathophysiology and therapeutic options in primary immune thrombocytopenia.

Blood Transfus. 2011; 9: 262–273.

46.

Audia

, Rossato

, Santegoets

. Splenic TFH expansion participates in B-cell differentiation and antiplatelet-antibody production during immune thrombocytopenia. Blood. 2014; 124: 2858–2866.

47.

Nimmerjahn

, Ravetch

J.V.

Fcγ receptors: old friends and new family members.

Immunity. 2006; 24: 19–28.

48.

Crow

A.R.

, Lazarus

A.H.

Role of Fcγ receptors in the pathogenesis and treatment of idiopathic thrombocytopenic purpura.

J Pediatr Hematol Oncol. 2003; 25: S14–S18.

49.

Asahi

, Nishimoto

, Okazaki

. Helicobacter pylori eradication shifts monocyte Fcγ receptor balance toward inhibitory FcγRIIB in immune thrombo-cytopenic purpura patients. J Clin Invest. 2008; 118: 2939–2949.

50.

Kühne

, Imbach

, Bolton-Maggs

P.H.

. Newly diagnosed idiopathic thrombocytopenic purpura in childhood: an observational study. Lancet. 2001; 358: 2122–2125.

51.

Stasi

, Stipa

, Masi

. Long-term observation of 208 adults with chronic idiopathic thrombocytopenic purpura. Am J Med. 1995; 98: 436–442.

52.

Frederiksen

, Schmidt

The incidence of idiopathic thrombocytopenic pur-pura in adults increases with age.

Blood. 1999; 94: 909–913.

53.

Neylon

A.J.

, Saunders

P.W.

, Howard

M.R.

, Proctor

S.J.

, Taylor

P.R.

, Northern Region Haematology Group. Clinically significant newly presenting autoimmune thrombocytopenic purpura in adults: a prospective study of a population-based cohort of 245 patients. Br J Haematol. 2003; 122: 966–974.

54.

Schoonen

W.M.

, Kucera

, Coalson

. Epidemiology of immune thrombocytopenic purpura in the general practice research database. Br J Haematol. 2009; 145: 235–244.

55.

Stasi

, Provan

Management of immune thrombocytopenic purpura in adults.

Mayo Clin Proc. 2004; 79: 504–522.

56.

Godeau

, Chevret

, Varet

. Intravenous immunoglobulin or high-dose methylprednisolone, with or without oral prednisone, for adults with untreated severe autoimmune thrombocytopenic purpura: a randomized, multicentre trial. Lancet. 2002; 359: 23–29.

57.

Stasi

Immune thrombocytopenia: pathophysiologic and clinical update.

Semin Thromb Hemost. 2012; 38: 454–462.

58.

Leontyev

, Katsman

, Branch

D.R.

Mouse background and IVIG dosage are critical in establishing the role of inhibitory Fcγ receptor for the amelioration of experimental ITP.

Blood. 2012; 119: 5261–5264.

59.

Cooper

Intravenous immunoglobulin and anti-RhD therapy in the management of immune thrombocytopenia.

Hematol Oncol Clin North Am. 2009; 23: 1317–1327.

60.

Crow

A.R.

, Lazarus

A.H.

The mechanisms of action of intravenous immunoglobulin and polyclonal anti-D immunoglobulin in the amelioration of immune thrombo-cytopenic purpura: what do we really know?

Transfus Med Rev. 2008; 22: 103–116.

61.

Tarantino

M.D.

, Bussel

J.B.

, Cines

D.B.

. A closer look at intravascular hemolysis (IVH) following intravenous anti-D for immune thrombocytopenic purpura (ITP). Blood. 2007; 109: 5527.

62.

Gaines

A.R.

Disseminated intravascular coagulation associated with acute hemo-globinemia or hemoglobinuria following Rh(O)(D) immune globulin intravenous administration for immune thrombocytopenic purpura.

Blood. 2005; 106: 1532–1537.

63.

Viamelli

, Galli

, de Vivo

. Efficacy and safety of splenectomy in immune thrombocytopenic purpura: long-term results of 402 cases. Haematologica. 2005; 90: 72–77.

64.

Balagué

, Vela

, Targarona

E.M.

. Predictive factors for successful laparoscopic splenectomy in immune thrombocytopenic purpura: study of clinical and laboratory data. Surg Endosc. 2006; 20: 1208–1213.

65.

Ghanima

, Godeau

, Cines

D.B.

, Bussel

J.B.

How I treat immune thrombo-cytopenia: the choice between splenectomy or a medical therapy as a second-line treatment.

Blood. 2012; 120: 960–969.

66.

Portielje

J.E.

, Westendorp

R.G.

, Kluin-Nelemans

H.C.

, Brand

Morbidity and mortality in adults with idiopathic thrombocytopenic purpura.

Blood. 2001; 97: 2549–2554.

67.

Godeau

, Stasi

Is B-cell depletion still a good strategy for treating immune thrombocytopenia?

Presse Mal. 2014; 43: e79–e85.

68.

Auger

, Duny

, Rossi

J.F.

, Quittet

Rituximab before splenectomy in adults with primary idiopathic thrombocytopenic purpura: a meta-analysis.

Br J Haematol. 2012; 158: 386–398.

69.

Arnold

D.M.

, Dentali

, Crowther

M.A.

. Systematic review: efficacy and safety of rituximab for adults with idiopathic thrombocytopenic purpura. Ann Intern Med. 2007; 146: 25–33.

70.

Godeau

, Porcher

, Fain

. Rituximab efficacy and safety in adult splenectomy candidates with chronic immune thrombocytopenic purpura: results of a prospective multicenter phase 2 study. Blood. 2008; 112: 999–1004.

71.

Miyakawa

, Katsutani

, Yano

. Efficacy and safety of rituximab in Japanese patients with relapsed chronic immune thrombocytopenia refractory to conventional therapy. Int J Hematol. 2015; 102: 654–661.

72.

Kistangari

, McCrae

K.R.

Immune thrombocytopenia.

Hematol Oncol Clin North Am. 2013; 27: 495–520.

73.

Stasi

, Cooper

, Del Poeta

. Analysis of regulatory T-cell changes in patients with idiopathic thrombocytopenic purpura receiving B cell-depleting therapy with rituximab. Blood. 2008; 112: 1147–1150.

74.

Patel

V.L.

, Mahevas

, Lee

S.Y.

. Outcomes 5 years after response to rituximab therapy in children and adults with immune thrombocytopenia. Blood. 2012; 119: 5989–5995.

75.

Provan

, Newland

A.C.

Current management of primary immune thrombocytopenia.

Adv Ther. 2015; 32: 875–887.

76.

Nomura

, Dan

, Hotta

, Fujimura

, Ikeda

Effects of pegylated recombinant human megakaryocyte growth and development factor in patients with idiopathic thrombocytopenic purpura.

Blood. 2002; 100: 728–730.

77.

Kuter

D.J.

New thrombopoietic growth factors.

Blood. 2007; 109: 4607–4616.

78.

, Yang

, Xia

. Thrombocytopenia caused by the development of antibodies to thrombopoietin. Blood. 2001; 98: 3241–3248.

79.

Imbach

, Crowther

Thrombopoietin-receptor agonists for primary immune thrombocytopenia.

N Engl J Med. 2011; 365: 734–741.

80.

Wang

, Nichol

J.L.

, Sullivan

J.T.

Pharmacodynamics and pharmacokinetics of AMG 531, a novel thrombopoietin receptor ligand.

Clin Pharmacol Ther. 2004; 76: 628–638.

81.

Bussel

J.B.

, Kuter

D.J.

, George

J.N.

. AMG 531, a thrombopoiesis-stimulating protein, for chronic ITP. N Engl J Med. 2006; 355: 1672–1681.

82.

George

J.N.

, Mathias

S.D.

, Go

R.S.

. Improved quality of life for romiplostim-treated patients with chronic immune thrombocytopenic purpura: results from two randomized, placebo-controlled trials. Br J Haematol. 2009; 144: 409–415.

83.

Khellaf

, Michel

, Quittet

. Romiplostim safety and efficacy for immune thrombocytopenia in clinical practice: 2-year results of 72 adults in a romiplostim compassionate-use program. Blood. 2011; 118: 4338–4345.

84.

Kuter

D.J.

, Bussel

J.B.

, Newland

. Long-term treatment with romiplostim in patients with chronic immune thrombocytopenia: safety and efficacy. Br J Haematol. 2013; 161: 411–423.

85.

Chalmers

, Tarantino

M.D.

Romiplostim as a treatment for immune thrombocytopenia: a review.

J Blood Med. 2015; 6: 37–44.

86.

Rashidi

, Roullet

M.R.

Romiplostim-induced myelofibrosis.

Blood. 2013; 122: 2001.

87.

Dadfamia

, Lee

Progression of romiplostim myelofibrosis to myeloproliferative neoplasm.

Blood. 2014; 123: 1987.

88.

Ruggeri

, Tosetto

, Palandri

. Thrombotic risk in patients with primary immune thrombocytopenia is only mildly increased and explained by personal and treatment-related risk factors. J Thromb Haemost. 2014; 12: 1266–1273.

89.

Erickson-Miller

C.L.

, DeLorme

, Tian

S.S.

. Discovery and characterization of a selective, nonpeptidyl thrombopoietin receptor agonist. Exp Hematol. 2005; 33: 85–93.

90.

Erickson-Miller

C.L.

, Delorme

, Tian

S.S.

. Preclinical activity of eltrom-bopag (SB-497115), an oral, nonpeptide thrombopoietin receptor agonist. Stem Cells. 2009; 27: 424–430.

91.

Bussel

J.B.

, Provan

, Shamsi

. Effect of eltrombopag on platelet counts and bleeding during treatment of chronic idiopathic thrombocytopenic pur-pura: a randomised, double-blind, placebo-controlled trial. Lancet. 2009; 373: 641–648.

92.

Tomiyama

, Miyakawa

, Okamoto

. A lower starting dose of eltrom-bopag is efficacious in Japanese patients with previously treated chronic immune thrombocytopenia. J Thromb Haemost. 2012; 10: 799–806.

93.

Garzon

A.M.

, Mitchell

W.B.

Use of thrombopoietin receptor agonists in childhood immune thrombocytopenia.

Front Pediatr. 2015; 3: 70.

94.

Saleh

M.N.

, Bussel

J.B.

, Cheng

. Safety and efficacy of eltrombopag for treatment of chronic immune thrombocytopenia: results of the long-term, open-label EXTEND study. Blood. 2013; 121: 537–545.

95.

Van Kooten

, Banchereau

CD40-CD40 ligand.

J Leukoc Biol. 2000; 67: 2–17.

96.

McMillan

The pathogenesis of chronic immune thrombocytopenic purpura.

Semin Hematol. 2007; 44(suppl 5): S3–S11.

97.

Brams

, Black

, Padlan

E.A.

. A humanized anti-human CD154 monoclonal antibody blocks CD154-CD40 mediated human B cell activation. Int Immunopharmacol. 2001; 1: 277–294.

98.

Kuwana

, Nomura

, Fujimura

. Effect of a single injection of humanized anti-CD154 monoclonal antibody on the platelet-specific autoimmune response in patients with immune thrombocytopenic purpura. Blood. 2004; 103: 1229–1236.

99.

Patel

V.L.

, Schwartz

, Bussel

J.B.

The effect of anti-CD40 ligand in immune thrombocytopenic purpura.

Br J Haematol. 2008; 141: 545–548.

100.

Song

, Kim

, Kwon

, Koo

, Jo

Expression of CD154 (CD40L) on stimulated T lymphocytes in patients with idiopathic thrombocytopenic purpura.

Hematology. 2016; 21: 187–192.

101.

Goldenberg

D.M.

, Rossi

E.A.

, Stein

. Properties and structure-function relationships of veltuzumab (hA20), a humanized anti-CD20 monoclonal antibody. Blood. 2009; 113: 1062–1070.

102.

Gómez-Almaguer

Monoclonal antibodies in the treatment of immune thrombocytopenic purpura (ITP).

Hematology. 2012; 17(suppl 1): S25–S27.

103.

Liebman

H.A.

, Saleh

M.N.

, Bussel

J.B.

. Low-dose anti-CD20 veltuzumab given intravenously or subcutaneously is active in relapsed immune thrombocytopenia: a phase I study. Br J Haematol. 2013; 162: 693–701.

104.

Mócsai

, Ruland

, Tybulewicz

V.L.

The SYK tyrosine kinase: a crucial player in diverse biological functions.

Nat Rev Immunol. 2010; 10: 387–402.

105.

Geahlen

R.L.

Getting Syk: spleen tyrosine kinase as a therapeutic target.

Trends Pharmacol Sci. 2014; 35: 414–422.

106.

Podolanczuk

, Lazarus

A.H.

, Crow

A.R.

, Grossbard

, Bussel

J.B.

Of mice and men: an open-label pilot study for treatment of immune thrombocytopenic pur-pura by an inhibitor of Syk.

Blood. 2009; 113: 3154–3160.

107.

Hlusi

, Szotkowski

, Indrak

Refractory immune thrombocytopenia. Successful treatment with repeated cyclosporine A: two case reports.

Clin Case Rep. 2015; 3: 337–341.

108.

Litton

Refractory idiopathic thrombocytopenic purpura treated with the soluble tumor necrosis factor receptor etanercept.

Am J Hematol. 2008; 83: 344.

109.

Taylor

, Neave

, Solanki

. Mycophenolate mofetil therapy for severe immune thrombocytopenia. Br J Haematol. 2015; 171: 625–630.

110.

Dai

W.J.

, Zhang

R.R.

, Yang

X.C.

, Yuan

Y.F.

Efficacy of standard dose rituximab for refractory idiopathic thrombocytopenic purpura in children.

Eur Rev Med Pharmacol Sci. 2015; 19: 2379–2383.

111.

Cuker

, Coles

A.J.

, Sullivan

. A distinctive form of immune thrombo-cytopenia in a phase 2 study of alemtuzumab for the treatment of relapsing-remitting multiple sclerosis. Blood. 2011; 118: 6299–6305.

112.

Zhang

X.L.

, Peng

, Sun

J.Z.

. Modulation of immune response with cytotoxic T-lymphocyte-associated antigen 4 immunoglobulin-induced anergic T cells in chronic idiopathic thrombocytopenic purpura. J Thromb Haemost. 2008; 6: 158–165.

113.

Fogarty

P.F.

, Seggewiss

, McCloskey

D.J.

, Boss

C.A.

, Dunbar

C.E.

, Rick

M.E.

Anti-interleukin-2 receptor antibody (daclizumab) treatment of corticosteroid-refractory autoimmune thrombocytopenic purpura.

Haematologica. 2006; 91: 277–278.

114.

Zhou

, Xu

, Qin

. A multicenter randomized open-label study of rituximab plus rhTPO vs rituximab in corticosteroid-resistant or relapsed ITP. Blood. 2015; 125: 1541–1547.

115.

Bussel

J.B.

, Kuter

D.J.

, Aledort

L.M.

. A randomized trial of avatrombopag, an investigational thrombopoietin-receptor agonist, in persistent and chronic immune thrombocytopenia. Blood. 2014; 123: 3887–3894.

116.

Nurden

A.T.

, Viallard

J.F.

, Nurden

New-generation drugs that stimulate platelet production in chronic immune thrombocytopenic purpura.

Lancet. 2009; 273: 1562–1569.