The ultimate aim of this project is to produce a drug for the prevention and treatment of heparin induced thrombocytopenia and thrombosis (HITT), a serious complication of heparin therapy for which there is currently no specific treatment. Heparin is a naturally-occurring anticoagulant that prevents the formation of clots and extension of existing clots within the blood. Major medical applications of heparin include dialysis, cardiac catheterization, and cardiopulmonary bypass surgery. Heparin therapy is usually safe and effective;however some patients develop a serious complication known as heparin induced thrombocytopenia (HIT). HIT is caused by an immunological reaction that targets platelets leading to a low platelet count (thrombocytopenia). HIT increases the risk of blood clots forming within blood vessels and blocking the flow of blood (thrombosis), this condition is known as heparin induced thrombocytopenia and thrombosis (HITT). We estimate the rate of occurrence of HITT to be ~10-20 cases/yr/hospital. Affected individuals have a 20-50% risk of developing new thromboembolic events, a mortality rate ~20%, and an additional ~10% of patients require amputations or suffer other major morbidity. It is generally accepted that the clinical manifestations of HITT are caused by antibodies that recognize a complex composed of heparin and platelet factor 4 (PF4). The interaction of PF4 tetramers with heparin results in the formation of ultralarge complexes (ULCs) which represent the major antigen recognized by pathogenic HITT antibodies. Importantly, PF4 must form tetramers as a prerequisite of ULC formation. In preliminary experiments we have selected PF4 as a potential drug target, identified a potential drug binding site at the dimer-dimer interface of PF4, computationally screened over one million small molecules to find those with potential to bind this site, and experimentally confirmed that several of these compounds disrupt PF4 tetramerization in vitro. The goal for phase 1 of this project is to identify a diverse set of lead compounds with favorable pharmacological properties for development as PF4 antagonists. To evaluate the feasibility of developing a PF4 antagonist-based therapeutic for HITT, we will begin by predicting which molecules from our existing list of preliminary leads are the most likely to bind the PF4 target site using physics-based computational methods. We will then experimentally validate the predicted PF4 antagonists for the ability to disrupt ULC and/or PF4 tetramer formation using in vitro assays. Finally, the validated active lead compounds will be optimized as necessary to create drug candidates that are suitable for in vivo testing. Successful completion of this first phase will enable the application of in vivo assays to demonstrate efficacy, potency, and lack of toxicity in an animal model system, and experimental structural characterization of active PF4-small molecule complexes in Phase 2.
The ultimate aim of this project is to produce a drug for the prevention and treatment of heparin induced thrombocytopenia and thrombosis (HITT), a serious complication of heparin therapy for which there is currently no specific treatment. As a large number of hospitalized patients are exposed to heparin, HITT is a major cause of treatment-induced morbidity and mortality in this patient population. An effective treatment for HITT would limit or prevent the complications of this disease, and may prove effective for other immune- mediated thrombolytic diseases including antiphospholipid syndrome (APS).
| Sachais, Bruce S; Rux, Ann H; Cines, Douglas B et al. (2012) Rational design and characterization of platelet factor 4 antagonists for the study of heparin-induced thrombocytopenia. Blood 119:5955-62 |
