167. Genome Editing to Generate the First Mouse Model of Alpha-One Antitrypsin Deficiency, the Leading Cause of Genetic COPD

Abstract

Alpha-one antitrypsin (AAT) deficiency is a common autosomal co-dominant genetic disorder. This condition affects 1:2500 individuals of European ancestry, leading to the development of lung and liver disease. Within North American and Northern European populations, an estimated 4% of individuals are carriers of mutant alleles. AAT deficiency presents with an emphysema phenotype in the lungs of older subjects. AAT deficient subjects can also suffer from liver disease of varying severity; however, lung disease is the principle cause of death. AAT is a protease inhibitor predominantly synthesized in the liver that belongs to the serine protease inhibitor (serpin) family. Upon secretion into the blood stream, AAT enters the lungs where it inactivates excess neutrophil elastase, thereby preventing damage to the alveoli. Mutations of the Serpina1 gene can lead to reduced serum levels of AAT and decreased protein functionality, allowing for unrestricted elastin breakdown, pulmonary inflammation and eventual emphysema. Currently, an animal model simulating the lung condition does not exist, which severely limits the development of therapeutics. This is due to the higher genomic complexity of mice compared to humans. Indeed due to amplification events, C57BL/6 mice have five genes that are homologous to human SERPINA1. To address this we generated a quintuple gene knockout using CRISPR/Cas9 system via zygote microinjection. We generated three founding lines in which all 5 copies of the gene were disrupted. Mice from all three lines demonstrate absence of hepatic and circulatory AAT protein as well as a reduced capability to inactivate neutrophil elastase. We also characterized the lung phenotype in response to a lipopolysaccharide challenge, where the model recapitulated many characteristics of the human lung disease including decreased elastance and increased compliance, and lung morphometry was also affected. Genomic and transcriptomic characterization will be presented. Future work will include challenges with cigarette smoke, a well-known disease accelerator in patients. Further, the ongoing generation of a new transgenic model carrying both the quintuple disruption of the murine Serpina1 genes and a single copy of the Z variant of human SERPINA1 will bring to the field the ultimate disease model that will finally allow researchers to evaluate the effects of liver-directed gene augmentation in the presence of Z-AAT polymers.