Complement in the Pathogenesis of Acute Chest Syndrome

Project Summary Sickle Cell Disease (SCD) is characterized by vasoocclusion of blood vessels and chronic hemolytic anemia, resulting in recurrent and episodic acute organ damage. Acute chest syndrome (ACS) is a leading cause of hospitalization, requires intensive care, and is the most common cause of death in SCD. Despite this, ACS

continues to be managed only with supportive interventions, such as oxygen therapy, antibiotics, and red blood

cell transfusions that likely fail to directly treat the underlying pathophysiology of this condition. Thus, there is an unmet need for better therapeutic interventions in ACS. Our long-term research objective is to define key molecular factors that drive ACS pathogenesis and use this information to identify targets to prevent or treat this

condition. We initially demonstrated that patients with ACS exhibit increased levels of complement activation, including alternative complement pathway (ACP) specific products. Furthermore, in select patients, complement inhibition appears to reverse hemolysis and result in rapid clinical improvement. Our overarching hypothesis is

that hemolysis-induced activation of the ACP amplifies hemolysis, injures endothelial cells, and activates neutrophils, thereby resulting in ACS in SCD. The key contribution of this proposal will be characterizing the role of ACP in hemolysis and ACS in pre-clinical models. To accomplish this, we will define the mechanisms by which

complement contributes to the development of ACS (aim 1) by evaluating the influence and interaction of complement with erythrocytes and endothelium using a complementary approach of novel pre-clinical animal models, microfluidics, and in vitro assays to measure complement dysregulation. To accomplish this, we

generated a novel humanized sickle mouse model that is deficient in complement component C3 (SSC3KO), which is protected from hemin-mediated ACS. In addition to this novel genetic model and microfluidics, we will utilize high-dimensional flow cytometry to characterize the complement and neutrophil-dependent mechanisms

contributing to ACS (aim 2). Finally, we will test the efficacy of pharmacological inhibitors to inhibit complement pathways to prevent or treat ACS (aim 3), by targeting the alternative (factor B) and terminal (C5) pathways to prevent hemolysis, neutrophil activation, endothelial injury, and subsequent ACS. The contribution of this overall

proposal is expected to be significant because it will allow us to develop tools that can help predict the likelihood of ACS and also potentially identify critical therapeutic targets designed to prevent or abort ACS. We anticipate that the successful completion of these studies will define an essential contribution of complement activation to

the development of ACS and, more importantly, could lead to a paradigm shift in the mechanistic understanding of hemolysis and complement in the pathophysiology of SCD.