Efficient Gene Delivery to Pig Airway Epithelia and Submucosal Glands Using Helper-Dependent Adenoviral Vectors

Abstract

Airway gene delivery is a promising strategy to treat patients with life-threatening lung diseases such as cystic fibrosis (CF). However, this strategy has to be evaluated in large animal preclinical studies in order to translate it to human applications. Because of anatomic and physiological similarities between the human and pig lungs, we utilized pig as a large animal model to examine the safety and efficiency of airway gene delivery with helper-dependent adenoviral vectors. Helper-dependent vectors carrying human CFTR or reporter gene LacZ were aerosolized intratracheally into pigs under bronchoscopic guidance. We found that the LacZ reporter and hCFTR transgene products were efficiently expressed in lung airway epithelial cells. The transgene vectors with this delivery can also reach to submucosal glands. Moreover, the hCFTR transgene protein localized to the apical membrane of both ciliated and nonciliated epithelial cells, mirroring the location of wild-type CF transmembrane conductance regulator (CFTR). Aerosol delivery procedure was well tolerated by pigs without showing systemic toxicity based on the limited number of pigs tested. These results provide important insights into developing clinical strategies for human CF lung gene therapy.

Efficient intratracheal delivery of airway epithelial cells in mice and pigs

“…We will apply whole lung lavage to remove nonadherent cells before harvest in future studies. Although delivery of viruses to pig airways has been reported by several groups (6,50), this is the first time that human lung epithelial cells were delivered and retained in pig lungs. As in mouse models, pretreatment of pig lungs with PDOC might further improve cell attachment.…”

Impact of gene editing on the study of cystic fibrosis

“…Of particular interest for CF, the combination of non-viral and viral delivery, specifically the use of chemically modified mRNA encoding site-specific nucleases delivered with chitosan nanoparticles, and AAV to deliver the donor, resulted in precise editing in the lungs, within a notable phenotypic change in a well-established transgenic mouse model of surfactant protein B deficiency (Mahiny et al 2015). The availability of large animal CF models has also established the feasibility of delivery of aerosolised virus vectors intratracheally into pigs under bronchoscopic guidance (Cao et al 2013;Yan et al 2015). With regard to cell-based therapies for CF, there are less data at present, but a recent proof-of-concept study in mice showed successful lung reconstitution by canicular-stage lung cells (Rosen et al 2016) raising the possibility of testing some of the recently described gene-edited iPSc models in conjunction with CF animal models as an alternative to direct editing of the lung.…”

“…The availability of three geneedited species placed CF researchers in a unique position for a genetic disease to undertake comparative pathophysiological studies which have revealed a number of less wellcharacterised features of the disease, such as abnormalities in alveolar macrophages, bone, and cartilage (reviewed by Wilke et al 2011;Keiser and Engelhardt 2011). These models will also be critical to optimization of delivery of editing machinery (Cao et al 2013(Cao et al , 2016 and/or edited cells (Butler et al 2016).…”

Barriers to inhaled gene therapy of obstructive lung diseases: A review

“…Importantly, Croyle et al showed that gene transduction in mouse lungs with HD-AdV was maintained after a second administration conducted 28 days after the initial dosing [92]. Airway transduction with HD-AdV has also been demonstrated in larger animals, including the ferret [93] and pig [94], further supporting its potential for inhaled gene therapy applications.…”