Novel Family of Insect Salivary Inhibitors Blocks Contact Pathway Activation by Binding to Polyphosphate, Heparin, and Dextran Sulfate

Abstract

Objective Polyphosphate and heparin are anionic polymers released by activated mast cells and platelets that are known to stimulate the contact pathway of coagulation. These polymers promote both the autoactivation of factor XII and the assembly of complexes containing factor XI, prekallikrein, and high-molecular-weight kininogen. We are searching for salivary proteins from blood-feeding insects that counteract the effect of procoagulant and proinflammatory factors in the host, including elements of the contact pathway. Approach and Results Here, we evaluate the ability of the sand fly salivary proteins, PdSP15a and PdSP15b, to inhibit the contact pathway by disrupting binding of its components to anionic polymers. We attempt to demonstrate binding of the proteins to polyphosphate, heparin, and dextran sulfate. We also evaluate the effect of this binding on contact pathway reactions. We also set out to determine the x-ray crystal structure of PdSP15b and examine the determinants of relevant molecular interactions. Both proteins bind polyphosphate, heparin, and dextran sulfate with high affinity. Through this mechanism they inhibit the autoactivation of factor XII and factor XI, the reciprocal activation of factor XII and prekallikrein, the activation of factor XI by thrombin and factor XIIa, the cleavage of high-molecular-weight kininogen in plasma, and plasma extravasation induced by polyphosphate. The crystal structure of PdSP15b contains an amphipathic helix studded with basic side chains that forms the likely interaction surface. Conclusions The results of these studies indicate that the binding of anionic polymers by salivary proteins is used by blood feeders as an antihemostatic/anti-inflammatory mechanism.

The polyphosphate–factor XII pathway drives coagulation in prostate cancer-associated thrombosis

“…by guest www.bloodjournal.org From or small molecules 54 may represent a promising strategy for interfering with cancer-driven coagulation and potentially, inflammatory reactions. 55 PolyP/FXII inhibition and deficiency in factors XI, XII, or HK interfered with prostasome-induced lethal PE (Figure 1). Similar to infusion of pure polyP 56 and platelet stimulation, 57 injection of prostasomes induced microvascular PE.…”

Factor XII: a novel target for safe prevention of thrombosis and inflammation

“…F12 −/− and wild‐type mice treated with FXIIa inhibitors or phosphatases were protected against pulmonary embolism induced by injection of purified polyP and activated platelets . Consistently, infusion of the proteins PdSP15a and PdSP15b from saliva of sand flies, which bind to polyP with high affinity, reduced FXIIa‐mediated clotting in vitro and attenuated vascular leakage in a model of skin inflammation in vivo . Universal heparin reversal agents (UHRAs) are synthetic amides that bind to polyanions and reverse the anticoagulant activity of heparin in a similar manner to protamine.…”

Factor XII Contact Activation

“…Recombinant PdSP15a/b, a sand-fly odorant-binding protein, binds to polyP and inhibits polyP/FXII-mediated BK formation and vascular leakage in mouse edema models. 37 Furthermore, synthetic polycationic polymers, polyamidoamine dendrimers (PAMAM), inhibit polyP-driven clotting in human plasma and interfere with platelet-driven arterial and venous thrombosis in murine models. Despite the potent antithrombotic activities of PAMAM, treated mice have low hemostatic disturbances.…”