Update on thrombotic thrombocytopenic purpura

The pathophysiology of various types of thrombotic microangiopathies is coming progressively into focus and therapeutic advances are likely to follow at a rapid pace. The discussion below focuses on thrombotic thrombocytopenic purpura.

Thrombotic thrombocytopenic purpura (TTP), the most severe microvascular occlusive thrombotic microangiopathy, is characterized by systemic platelet aggregation, organ ischemia, profound thrombocytopenia and fragmentation of erythrocytes. The red blood cell fragmentation occurs as blood flows through turbulent areas of the microcirculation partially occluded by platelet aggregates. Schistocytes, or “split” red cells, appear on the peripheral blood smear (1% to 4% or more of total RBCs) as an indication of microangiopathic hemolytic anemia. Serum levels of L-lactate dehydrogenase are extremely elevated as a consequence of hemolysis and the leakage of LDH from ischemic or necrotic tissue cells.

The systemic platelet clumping in TTP is often associated with platelets <20,000/mcL. Occlusive ischemia of the brain or the gastrointestinal tract is common and renal dysfunction may occur. In current clinical practice, the triad of thrombocytopenia, schistocytosis and an impressively elevated serum LDH are sufficient to suggest the diagnosis.

Joel Moake

Monomers of VWF (250,000 Da) are linked by disulfide bonds VWF multimers with molecular masses that may reach the millions of Da. VWF multimers are constructed predominantly within endothelial cells and stored within Weibel-Palade bodies as immense, coiled ultra-large VWF multimers. When stimulated, endothelial cells secrete the ultra-large VWF multimers in long strings that remain anchored to the cell membrane. The long VWF multimeric strings are extremely “sticky” to the glycoprotein Iba components of platelet GPIb-IX-V surface receptors. The initial adherence of platelets via GPIba receptors to the long VWF strings and the subsequent coherence of additional platelets to each other (aggregation) via activated GPIIb/IIIa receptors produces potentially occlusive platelet thrombi.

A specific VWF–cleaving protease that circulates in normal plasma rapidly cleaves the highly adhesive long VWF strings as they are secreted from endothelial cells by cleaving a tyrosine 1605-1606 methionine peptide bond in one or more susceptible VWF monomeric subunits. The protease is number 13 in a family of 19 distinct ADAMTS–type metalloprotease enzymes.

ADAMTS13 is a disintegrin and metalloprotease with eight thrombospondin 1–like domains composed of an aminoterminal metalloprotease domain followed by: a disintegrin domain; a thrombospondin 1–like domain; a cysteine-rich domain and an adjacent spacer portion; seven additional thrombospondin 1–like domains; and two other different types of domains that resemble each other at the carboxyl-terminal end of the molecule.

ADAMTS13 is a Zn²⁺–requiring and Ca²⁺–requiring 190,000 Da glycosylated protein that is encoded on chromosome 9q34. It is produced predominantly in endothelial cells for slow, constitutive release into the circulation. Endothelial cells can be stimulated to secrete long VWF strings by inflammatory cytokines (tumor necrosis factor-alpha [TNFA], interleukin-8 and IL-6, Shiga toxins, estrogen or other agonists). ADAMTS13 is inhibited by EDTA and therefore functional assays of the enzyme are usually performed using plasma anticoagulated with citrate (a weaker divalent cation binder than EDTA).

ADAMTS13–deficient types of TTP

The severe deficiency of ADAMTS13 activity in familial or acquired autoantibody-mediated TTP patient plasma results in the failure to cleave long VWF multimers as they emerge in long strings from the surface of stimulated endothelial cells. Familial TTP is rare and usually (but not always) appears initially in infancy or childhood. It also may recur as “chronic relapsing TTP” episodes at about three-week intervals. Patients with familial, relapsing TTP have less than about 5% to 10% of normal plasma ADAMTS13 (both during and between episodes).

The absent or severely reduced plasma ADAMTS13 activity in familial TTP is a consequence of homozygous (or double heterozygous) mutations in both of the ADAMTS13 alleles that result in deficient production or release of ADAMTS13 molecules. Mutations in familial TTP have been detected all along the gene, in regions encoding different domains. In severe familial ADAMTS13 deficiency, TTP episodes usually commence in infancy or childhood.

In other familial TTP patients with slightly higher plasma ADAMTS13 levels, overt TTP episodes may not develop until later in life (eg, during a first pregnancy). Accentuated secretion of long VWF multimeric strings by endothelial cells stimulated by estrogen (as during menses or pregnancy) or inflammatory cytokines may be required to provoke TTP episodes in these latter patients

Adults and some older children with acquired anti-ADAMTS13 autoantibody–mediated TTP have transient inhibition of ADAMTS13 to less than about 5% to 10% of normal only during acute episodes (or later recurrence). Plasma ADAMTS13 levels then increase toward normal during recovery. Recurrences occur at irregular intervals in about one out of three patients. Immunoglobulin G autoantibodies that inhibit plasma ADAMTS13 activity during acquired episodes are detected in most of these patients, indicating transiently defective immune regulation.

A small fraction of patients treated for arterial thrombosis with the platelet P2Y₁₂adenosine diphosphate receptor–inhibiting thienopyridine drugs ticlopidine (Ticlid) or clopidogrel (Plavix) develop TTP within a few weeks after the initiation of therapy. Autoantibodies that inhibit plasma ADAMTS13 have also been demonstrated in a few patients with ticlopidine or clopidogrel-associated TTP, indicating possible immune dysregulation induced by these similar thienopyridine compounds.

An alternative possibility is that the binding of thienopyridines to P2Y₁₂ molecules on endothelial cell (and other cell) types may in a fraction of exposed individuals initiate anomalous intracellular signaling patterns or provoke antibody production against thienopyridine (as hapten)-bound P2Y₁₂ protein complexes on cell surfaces. Malfunction or injury to endothelial cells and other cells with surface P2Y₁₂ receptors (lymphocytes and CD34 stem cells) may result.

TTP occurs occasionally late in pregnancy or immediately after delivery or in patients with HIV/AIDS.

Plasma ADAMTS13 activity in healthy adults ranges from about 50% to 180%. Activity is often reduced below normal in liver disease, disseminated malignancies, chronic metabolic and inflammatory conditions, pregnancy and newborns. With the exception of the occasional peripartum women who develop overt TTP, the ADAMTS13 activity in these conditions is not reduced to the extremely low values (ie, <5% to 10% of normal) found in patients with familial or acquired autoantibody- mediated TTP.

Therapy

Normal fresh-frozen plasma, the cryoprecipitate-poor fraction of plasma (cryosupernatant) and solvent/detergent-treated plasma all contain active ADAMTS13. The infusion about every two to three weeks of normal plasma into familial TTP patients lacking effective enzyme production/release prevents or reduces the frequency of TTP episodes.

Adults and some older children with acquired ADAMTS13 autoantibody-mediated TTP require daily plasma exchange until remission. Plasma exchange combines the infusion of fresh-frozen plasma or cryosupernatant (containing uninhibited ADAMTS13) with plasmapheresis (which may remove autoantibodies against ADAMTS13 and cytokines that stimulate endothelial cells to secrete long VWF strings).

Plasma exchange allows about 80% of acquired ADAMTS13 autoantibody–mediated TTP patients to survive an episode, usually with minimal organ damage. Lower titers of autoantibodies are associated with better responses to plasma exchange. Production of ADAMTS13 autoantibodies may be suppressed by high-dose glucocorticoids, four to eight weekly doses of rituximab (Rituxan, Genentech; monoclonal antibody against CD20 on B lymphocytes) or removal of autoantibody-producing cells by splenectomy.

ADAMTS13 has been partially purified and produced in recombinant active form for possible eventual therapeutic use. Because plasma ADAMTS13 levels of only about 5% are often sufficient to prevent or truncate TTP episodes, gene therapy may eventually extend remissions in familial TTP patients. An anti-VWF aptamer (oligonucleotide that binds to the VWF A1 domain and blocks its binding to platelet GPIba) has recently been synthesized.

In preliminary trials, this FDA- approved “orphan drug” (ARC-1779) was effective in the treatment of a few patients with relapsing and refractory types of TTP.

Joel Moake, MD, is a Senior Research Scientist at Rice University and is a member of the HemOnc Today Editorial Board.

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