Project Summary (OVERALL) Alpha-1 antitrypsin deficiency (AATD) is caused by mutations in the SERPINA1 gene. The E342K (PI*Z) mutant allele is very common among those of European ancestry, and E342K homozygotes encode a protein with impaired secretion, resulting in deficient AAT serum levels. Since AAT normally protects elastin in the lung from degradation, loss of effective AAT triggers lung inflammation, airways obstruction and emphysema, which is the primary life-limiting manifestation of AATD. The projects within this proposal seek to pursue numerous parallel strategies to develop a gene therapy for AATD. Most of these strategies revolve around the use of recombinant adeno-associated virus (rAAV)-based vectors, a platform technology that has been very successful for other genetic diseases. In Project 1, optimized rAAV vectors will be studied in genetically defined animal models (including mice and ferrets) in comparison with transgenic reconstitution studies using a regulated conditional transgenic system to compare two relevant potential target replacement levels (11µM and 25µM) and clinically relevant endpoints will be studied. In Project 2, novel CRISPR variants will be used for gene editing, base editing and prime editing strategies to treat AATD. In Project 3, we will screen naturally occurring AAV capsid libraries obtained from remote populations in Western China to identify capsids with enhanced efficacy and safety for AATD gene therapy. Finally, in project 4, we will use novel Treg and CAR-Treg strategies to selectively modulate anti-vector immune responses. There will also be two cores. Core A will provide each project with important Vector Immunology assays, which can identify limitations due to host immune responses to AAV capsids, the AAT transgene or to Cas9-derived proteins. Core B will provide animal models and physiologic measurements in the animal models for testing of optimized rAAV vectors, gene editing tools and immune modulation approaches. Program investigators have a track record of interactions and collaborations that we anticipate will continue in future years.

OVERALL - Project Narrative In this project, we will develop new gene therapy drugs to treat genetic emphysema due to alpha-1 antitrypsin deficiency (AATD), a common genetic disease and form of chronic obstructive pulmonary disease (COPD). In the proposal, we will use advanced gene editing tools to create genetically defined animal models of AATD (known as transgenic animals), both in mice and in ferrets, which are a good model to study lung diseases. In the course of the study, we will use the most cutting-edge tools in gene therapy, gene editing, and immune modulation to treat AATD in both animal models, and determine the best ways to measure the success of gene therapy over time.