Cystic Fibrosis Gene Therapy with adeno-associated viral vectors

Abstract: We and others have shown that AAV vectors have great potential as gene therapeutic agents (3-5), in particular for CF. Studies originating from our group led to the first use of rAAV in humans (7). Many pre- clinical and clinical trials have shown that AAV vectors can be used safely (5,6,9). The major challenge, however, is that they have not achieved a reproducible therapeutic effect, making it necessary to take a new approach to AAV gene therapy. In previous work, we have identified three new strategies to alleviate these problems: 1) the use of AAV1, which is more tropic for the lung; 2) use of 27-264, a truncated version of CFTR that corrects F508 by a novel mechanism; and 3) inclusion of a powerful chicken β-actin (CBA) promoter. Recent advances have been made in developing corrector and potentiator compounds to rescue and activate F508-CFTR (8). How- ever, although rescuing F508-CFTR would benefit many CF patients, there are more than 1000 other mutations in the CF gene (http://www.cftr2.org/), many of which create mutant proteins that are not repairable by the same compounds that rescue F508. These limitations in rescuing F508-CFTR and other mutant CFTR molecules make it even more important to develop a gene therapy for CF to treat all patients with CF. To further explore the potential usefulness of gene therapy for CF, we will use Rhesus macaques to explore the toxicology and pharmacology of AAV1-CBΔ27-264. We will use a ferret model bearing the F508 mutation to evaluate rescue of the CF phenotype in a large animal model. This model is ideal for our studies because ferrets show a tropism for AAV1 similar to that of humans and monkeys (9), and the disease phenotype can be switched on and off following application or cessation CFTR corrector/potentiator treatment. Finally, we will conduct a phase I clinical trial in patients bearing the F508 mutation. We propose three overall Specific Aims: Aim I: To evaluate single-dose administration of AAV1-Δ27-264CFTR to Rhesus macaques. Aim 2: To determine the therapeutic effects of AAV1 vectors containing truncated CFTR in F508-del Ferrets. Aim 3: To determine whether dosing with an AAV1 vector containing a truncated CFTR will lead to transduc- tion in ferret and human primary airway cells. The overarching questions for CF gene therapy are whether gene transduction can rescue the CF phenotype and how long the therapeutic effect will last before a repeated administration is necessary. The answers to these practical questions have enormous clinical consequences for the development of effective CF gene therapy. This application will address the extent to which an AAV1 vector containing AAV1-CBΔ27-264 will be effective in rescuing the CF phenotype and how long the rescue will last. Significance: CF is an autosomal disease that leads to significant morbidity and mortality in patients with the disorder (10). The work is significant because it will address the safety and efficacy of CFTR delivery via AAV1 viral vectors to lung and to demonstrate CFTR expression.