Recombinant Alpha-1 Antitrypsin as a Novel Therapy for Pandemic Influenza

FY21 PRMRP Topic Area: Respiratory Health Rationale: Influenza A virus (IAV) is a major cause of respiratory disease with a spectrum from mild illness caused by seasonally circulating strains to severe disease observed during infection with highly pathogenic avian influenza (HPAI) virus strains with pandemic potential. Pandemic influenza may be associated with a high mortality rate in healthy young adults, including defense force personnel. HPAI virus strains induce a very marked inflammatory response in the lung. This response damages the lung, particularly the epithelium. In this circumstance, patients may develop complications including acute respiratory distress syndrome (ARDS) and secondary bacterial pneumonia. IAV infection activates inflammatory pathways that may contribute to lung damage, and one of these is the formation of extracellular traps. These are made by neutrophils (NETs) and macrophages (METs). The extracellular traps are composed of extracellular chromatin (DNA) and other components such as proteases which are pro-inflammatory. In certain situations, strong activation of NETs/METs may be damaging to the host. Deoxyribonuclease (DNase) 1 dismantles extracellular traps (and is used as a standard inhalational therapy in the treatment in cystic fibrosis). Alpha-1 antitrypsin (AAT) is the main antiprotease produced in the body and has potent anti-inflammatory effects. It is used as a therapy for AAT deficiency but is costly, unstable, and hard to obtain. We have convincing preliminary data demonstrating that AAT reduces the inflammatory response induced by NETs/METs and there is a synergistic effect when AAT is combined with DNase 1. In addition, we have developed a unique recombinant AAT (rAAT) that is very stable and may be given by inhalation. Hypotheses: Recombinant AAT will reduce the inflammatory response and improve outcomes in severe IAV infection. This effect may be facilitated by the addition of DNase 1. Aims: (1) In human models to define the ability of: (i) rAAT and (ii) combined rAAT/ DNase 1 to reduce lung inflammation in IAV infection. (2) In animal models to define the ability of: (i) rAAT and (ii) combined rAAT/DNase to reduce lung inflammation in IAV infection. Study Design: We have established methods to measure extracellular traps, both in human samples and animal models. There are established models of IAV infection and methods to give nebulized AAT and DNase 1. Physical containment (PC) 2 and PC3 facilities will be used as appropriate. For Aim 1, we will assess responses in macrophages and neutrophils and in lung tissue. We will measure: (1) NET/MET expression, (2) inflammatory markers, and (3) IAV infection. Results will be compared between different treatment groups, ages, and IAV strains. For Aim 2, in vivo responses to infection and treatment will be assessed as listed for Aim 1. Clinical parameters of the mice will also be measured. Innovation: We have developed a unique level of expertise to study extracellular traps in the lung and study inflammatory responses to medications. This work will establish a new paradigm to explain how IAV infection may cause severe lung damage. Most importantly, it will provide a rationale for a new therapeutic approach to manage IAV infection by repurposing standard drugs (AAT and DNase 1). To facilitate this, we have developed a unique AAT formulation that circumvents the traditional problems with blood donor-derived products. The concept of a synergistic action between AAT and DNase 1 is also novel. Expected Results: This study will demonstrate that extracellular traps help drive lung inflammation in more virulent IAV infection. This inflammatory response will be reduced by the use of our recombinant AAT. This effect of AAT may be enhanced by the addition of DNase 1. How the Results Will Be Used as a Foundation for Future Research Projects: Both DNase 1 (dornase alfa) and AAT have been used safely for many years in patients. If successful, this study will provide a rationale for the use of AAT and DNase 1 in clinical trials to reduce/prevent severe lung damage arising from extracellular trap expression. Less