Platelets play a central role in hemostasis via injury site-selective multi-step mechanisms of: (1) Adhesion to vWF and collagen, (2) Fibrinogen-mediated aggregation to form the primary hemostatic plug, (3) Biointerfacial presentation of anionic phosphatidylserine (PS) on the activated platelet surface for procoagulant amplification of thrombin (hence fibrin), and (4) clot-localized secretion of platelet granule contents (e.g. inorganic polyphosphate, PolyP) to locally enhance fibrin stability. These mechanisms are significantly compromised in non-compressible traumatic hemorrhage, which remains a major cause of mortality. The `gold standard' for treating such hemorrhage is massive transfusion of whole blood or components (platelets, plasma, RBC). Especially, platelet transfusion has shown tremendous clinical benefit in saving lives in trauma. However, platelets are rarely available in resource-limited hospitals and unavailable pre-hospital, due to challenges of storage, portability, high risk of bacterial contamination and very short shelf-life (~5 days). We aim at addressing this challenge by designing biomaterials-based `artificial platelet' nanoconstructs. To this end, utilizing a previous R01 award (HL121212) we developed self-assembled lipid-peptide nanoconstructs that mimic and integrate the platelet mechanisms of (1) and (2) stated above. This design showed hemostatic ability in vitro and in thrombocytopenic mouse tail-bleeding models, and modest efficacy in severe trauma models. Building on this, we now propose to mimic the mechanisms of (3) and (4) on a liposomal template by designing unique enzyme- responsive lipopeptides, that will subsequently allow integration of all four mechanisms onto a single nanoconstruct for a superior artificial platelet design. Our central hypothesis is `Modular amplification of hemostasis via mimicry of platelet's biointerfacial and secretory mechanisms within an artificial platelet construct can significantly attenuate hemorrhage and enhance survival in trauma'. To test this, our Specific Aims are to: (1) Evaluate stimuli (plasmin)-triggered exposure of PS on lipidic nanoconstructs for platelet-inspired amplification of thrombin (hence fibrin) site-specifically in trauma; (2) Evaluate stimuli (thrombin)-triggered release of inorganic polyphosphate (PolyP) as a payload from lipidic nanoconstructs for injury site-targeted stabilization of fibrin clot; and (3) Integrate these independent synergistic components in artificial platelet nanoconstructs to evaluate hemostatic efficacy and survival in rodent trauma model. The traumatic insult to vascular endothelium results in enhanced secretion of tissue plasminogen activator (hence plasmin) at the clot site, resulting in rapid fibrin degradation (hyperfibrinolysis) and compromising clot stability. Exploiting this plasmin to expose PS on `artificial platelet' surface will allow enhanced thrombin (and hence fibrin) generation to offset hyperfibrinolysis. This thromb...