Development of a Neutrophil Degranulation Inhibitor to Treat ARDS

Project Summary Acute respiratory distress syndrome (ARDS) is a critical problem in pulmonary medicine, accounting for 10% of intensive care unit (ICU) admissions and totaling over 200,000 patients/year in the U.S. Current treatment cost typically exceeds $70,000 per patient. Decades of clinical trials failed to identify effective pharmacologic therapy for ARDS, while mortality remains above 30%. Thus, there is a critical, unmet clinical need for successful pharmacologic strategies to treat ARDS. Neutrophils play an essential role in the lung injury leading to ARDS, including that due to COVID-19, through extracellular release of reactive oxygen species (ROS), granule constituents, and neutrophil extracellular traps (NETs). The scientific foundation underlying our innovative therapy is that inhibition of neutrophil degranulation also prevents release of ROS and NETs, making degranulation a therapeutic target. We generated a recombinant protein, degranin-23 (degranulation inhibitor of SNAP-23, DGN-23) containing a SNARE motif from SNAP-23 and a cell permeability peptide. DGN-23 rapidly enters human neutrophils in vitro and in vivo, inhibits degranulation by 50% to 80%, prevents priming of ROS release, and reduces NET formation. Importantly, in vitro studies show DGN-23 does not impair neutrophil phagocytosis, granule fusion with phagosomes, or bacterial killing within phagosomes, and in vivo studies show that DGN-23 administration at the initiation of lung injury inhibits acute lung injury in 3 rodent models, without obvious toxicity. Thus, in vitro and in vivo data provide strong support for our novel therapeutic strategy that inhibition of neutrophil exocytosis attenuates lung injury leading to ARDS. The next steps are development of manufacturing capability and quality control to move from a laboratory grade recombinant protein to a pharmaceutical grade drug and validation of therapeutic effect without toxicity under clinically relevant conditions. This transition will be accomplished by two Aims. Aim 1: Characterize production characteristics and off-target/side effects of DGN-23. This aim will determine purity, presence of bacterial contaminants, yield, reproducibility of potency, and stability in multiple production runs of DGN-23. Cell toxicity and off target effects on circulating immune cells will be determined. Organ and cell localization of DGN-23 will be determined in all major organs from animals undergoing experiments described in Aim 2. Aim 2: Determine therapeutic efficacy of DGN-23 under clinically relevant conditions. The ability of DGN-23 to prevent acute lung injury and improve mortality when administered at various times after the initiating injury will be determined in a mouse model of H1N1 influenza virus-induced acute lung injury. Improved survival and improved blood oxygenation will serve as outcomes to proceed with development. At the conclusion of this work, this Phase 1 project will have identified a drug candidate with an optimal combination of efficacy, yield, stability, and potency to advance to advanced pre-clinical studies in a Phase 2 project.