The Development of Novel Lung-Tropic AAVs to Support a Gene Therapy for Cystic Fibrosis

Abstract

Cystic fibrosis (CF) is a life-shortening disease that affects all mucosal surfaces of the body, with the more severe symptoms manifesting in the respiratory system. This monogenic disease is caused by a variety of mutations in the CFTR gene. This project aimed to identify/develop multiple lung targeted AAV capsids in support of a gene therapy for CF. Novel capsids were specifically evaluated for their ability to target human airway basal (AB) cells, which are the progenitors of the CFTR producing airway epithelial cells and ionocytes. To bypass the airway's innate defences, including the thick mucus layer, which is especially prominent in people with CF, whilst also addressing peripheral symptoms, capsids were engineered for effective transduction following systemic administration.

This thesis employed three approaches to develop novel lung targeted AAV capsids. Firstly, a high-throughput approach to identify currently available capsids with previously unrealised lung tropism. Secondly, directed evolution to develop novel, bioengineered capsids with improved lung transduction efficiency. Finally, the isolation of novel, natural AAV capsid variants from human lung tissue sample.

To address challenges relating to clinical translation of AAV transduction efficiency, a ‘multi-model’ approach was employed for capsid evaluation. A mouse model was used to evaluate capsids based on their ability to travel to the lungs following systemic administration, a precision cut human lung slice model was used to assess the capsids’ ability to transduce human lung tissue, and finally primary human airway basal cells were used to evaluate the capsids’ ability to transduce the target cells.

This study identified multiple AAV capsids with improved transduction efficiency in pre-clinical lung models over current benchmarks such as AAV2 and AAV6. Whilst these capsids require further evaluation, they could be prime candidates for use in pre-clinical studies or even clinical translation.