SUMMARY
BACKGROUND: The balance between clot formation (coagulation) and degradation (fibrinolysis) is disrupted in
patients with bleeding disorders or severe hemorrhage, resulting in increased bleeding. The extent to which
fibrinolysis contributes to bleeding from the onset of acute bleeding in these contexts is unclear, and this gap in
knowledge limits the development of therapies that inhibit fibrinolysis. Current therapeutics are often not sufficient
for patients with bleeding disorders; for example, antifibrinolytic drugs are widely used to manage acute bleeding
in these patients, but are unsuitable for long-term prophylaxis due to short half-lives that decrease efficacy.
RATIONALE: We hypothesize that fibrinolysis contributes to the incidence and severity of bleeding starting from
the onset of bleeding, and that long-acting RNA agents targeting plasminogen, the zymogen precursor of the
main fibrinolytic enzyme plasmin, can decrease fibrinolysis and help manage bleeding long-term for people
across all classes of bleeding disorders, including diagnosed and undiagnosed disorders. This knowledge will
provide insights into the mechanisms that impact outcomes in traumatic hemorrhage. These RNA agents will be
delivered using lipid nanoparticles (LNPs), a clinically approved delivery platform we have unique expertise in.
SPECIFIC AIMS: We will develop an RNA therapeutic targeting plasminogen, using small interfering RNA
(siRNA)-mediated gene silencing (siPlg). This therapy is long-acting over several weeks per dose and highly
specific for the target protein, which decreases the burden of frequent administration. The specific aims of this
proposal are to (1) Develop approaches for knockdown of plasminogen and inhibition of fibrinolysis in mice, pigs,
and dogs, and characterize the safety of this approach; (2) Determine the contribution of fibrinolysis to blood
loss at the onset and in early stages of hemorrhage in a traumatic injury model in swine; and (3) Investigate the
role of fibrinolysis in bleeding disorders by testing the effect of plasminogen knockdown in mouse and canine
models of hemophilia A, and mouse models of hemophilia B and von Willebrand disease.
INNOVATION: This work has both mechanistic and therapeutic innovation. It will immediately determine the
contribution of fibrinolysis to bleeding in early stages of severe hemorrhage and in bleeding disorders. This will
likely be the first development of RNA-LNP therapies targeting circulating proteins in large animal models of
bleeding. These findings will address a fundamental question in trauma and hemostasis research. Separately, it
will aid the development of improved, long-acting therapies for controlling bleeding in bleeding disorders. These
therapies would be particularly beneficial for women with menorrhagia caused by bleeding disorders.
EXPECTED OUTCOMES: We expect to (1) Develop species-specific siPlg that have minimal adverse health
effects; and (2-3) Demonstrate that knocking down plasminogen inhibits fibrinolysis and decreases blood loss in
swine models of traumatic hemorrhage, and in murine and canine models of bleeding disorders.
