Strategies to Provide Sufficient Anti-Protease Protection at Rest and During Acute Inflammation in ZZ Alpha-1 Antitrypsin Deficiency

Alpha-1 antitrypsin (AAT) is a 52 kDa glycoprotein synthesized predominantly in the liver. AAT is a serine protease inhibitor but has also been shown to possess significant anti-inflammatory properties. AAT deficiency (AATD) is a hereditary disorder. The most common mutation causing AATD is the Z mutation, clinically characterized by the retention of polymerised Z protein in the liver, resulting in a toxic “gain of function.” This intracellular retention of AAT leads to decreased secretion of AAT into the circulation and lung, which leads to unopposed proteolytic action of proteases such as neutrophil elastase (NE) in the lung, resulting in tissue destruction and emphysema, a “loss of function.” The ideal therapeutic treatment for AATD would increase secretion of Z AAT out of the liver (removing the toxic “gain of function”) and into the circulation and lung, where it would protect the lungs (preventing any “loss of function”). In the setting of the worldwide COVID-19 pandemic, the intense acute inflammatory response observed in healthy individuals in response to SARS-CoV-2 infection highlighted that baseline M AAT levels would not be sufficient for ZZ individuals in an acute infection. 

The aim of this study was to investigate the feasibility of a potential treatment option that would simultaneously treat both lung and liver disease, moving AAT out of the liver into the circulation and lung. The potential treatment was gentamicin and this study evaluated not only its ability to get Z AAT out of the cell, but also the amount of extracellular AAT required to protect the lung not only in quiescent times but also in time of severe inflammation such as SARS-CoV-2 infection. This study also evaluated the possibility that therapeutics aimed at increasing Z AAT extracellular secretion may also inadvertently cause increased intracellular Z AAT retention leading to inflammation and ER stress. 

A variety of cell models of AATD were used, aiming to recapitulate the Z and M phenotypes. The results demonstrated that these cell lines did behave like M and Z cells but the secreted Z AAT response to gentamicin was insufficient when compared to the AAT levels observed in SARS-CoV-2 infection, even in its milder forms with the added problem that there was evidence of increased intracellular AAT retention and ER stress. In the next stages of this study various degrees of SARSCoV-2 infection were used to determine what was driving the AAT response to inflammation and how much AAT was required to safeguard the lung and other organ. The data showed that interleukin (IL)-6 was the main driver of AAT production and glycosylation of the AAT protein. The latter point is important as glycosylation materially affects the function of AAT as an anti-inflammatory specifically with regards to modulation of neutrophil migration.

Furthermore it was discovered that even with large amounts of AAT being produce by the liver in response to severe SARS-CoV-2 infection, there was still inadequate anti-protease protection in the airways due to the large NE burden, further evidence of the lack of utility of the commonly accepted putative protective AAT threshold of 11µM. These data also suggested a possible therapeutic option in SARS-CoV-2 ARDS, namely augmentation therapy with plasma purified AAT even in non-AATD MM individuals and led to a therapeutic trial of this medication in Beaumont hospital, the first randomised control trial of its type in the world. The data also suggested added caution is required in SARS-CoV-2-infected individuals who are treated with tocilizumab, an IL-6 receptor blocker as it blocks the AAT response to SARS-CoV-2 infection. Finally, SARS-CoV-2 infection trends in the ZZ AATD population were examined and using Astra Zeneca (AZ) vaccines as a surrogate for SARS-CoV-2 infection it was possible to demonstrate that while the anti-inflammatory response of ZZ AATD individuals is increased in vivo in response to SARS-CoV-2 infection and minimally to AZ vaccination it is inadequate and may be pro-inflammatory due to increased AAT production, retention and ER stress.

In conclusion, this research will contribute significantly to the body of knowledge regarding AATD, AAT’s role in the inflammatory process and its potential therapeutic use outside of AATD. This study has explored the available in-vitro models, the target levels of secreted AAT to aim for, in particular during acute inflammatory periods and the potential downsides of increasing AATD production in ZZ AATD cells. It also suggests that while gentamicin may not be a feasible therapeutic option, the concept of getting Z AAT out of the liver and into the circulation, either by refolding or gene editing offers significant opportunities for effective treatment.