Transduction of Ferret Surface and Basal Cells of Airways, Lung, Liver, and Pancreas via Intratracheal or Intravenous Delivery of Adeno-Associated Virus 1 or 6.

Cystic fibrosis (CF) is potentially treatable by gene therapy. Since the identification of the CF gene, preclinical and clinical trials have concentrated on achieving effective gene therapy targeting the lung. However, the lung has proven to be a formidable barrier to successful gene therapy especially for CF, and many clinical trials failed to achieve efficacy. Recent advances in vector design and adeno-associated virus (AAV) serotypes have increased the chances of success. Given that CF is a multi-organ disease, the goal of this study was to test whether a gene therapy approach involving AAV1 or AAV6 vector delivery via the systemic circulation would at the same time overcome the barrier of lung delivery and transduce organs commonly affected by CF. To accomplish this, we sprayed AAV1 containing green fluorescent protein (GFP) into the trachea or injected it intravenously (IV). We also tested AAV6 injected IV. No adverse events were noted. Ferrets were necropsied 30 days after vector delivery. AAV1 or AAV6 vector genomes, messenger RNA (mRNA) expression, and GFP were detected in all the tracheal and lung samples from the treated animals, whether AAV1 was sprayed into the trachea or injected IV or AAV6 was injected IV. Importantly, both surface epithelial and basal cells of the trachea and lung airways were successfully transduced, regardless of which route of delivery or vector serotype used for transduction. We detected also AAV1 and AAV6 vector genomes, mRNA expression, and GFP in the livers and pancreases, particularly in the acinar cells of the pancreatic duct. These data suggest that gene transfer is attainable in the airways, liver, and pancreas using either serotype, AAV1 or AAV6. Given that these same organs are affected in CF, systemic delivery of AAV may be the preferred route of delivery for a gene therapy for CF.

Ameliorating liver disease in an autosomal recessive polycystic kidney disease mouse model.

Systemic and portal hypertension, liver fibrosis, and hepatomegaly are manifestations associated with autosomal recessive polycystic kidney disease (ARPKD), which is caused by malfunctions of fibrocystin/polyductin (FPC). The goal is to understand how liver pathology occurs and to devise therapeutic strategies to treat it. We injected 5-day-old Pkhd1del3-4/del3-4 mice for 1 mo with the cystic fibrosis transmembrane conductance regulator (CFTR) modulator VX-809 designed to rescue processing and trafficking of CFTR folding mutants. We used immunostaining and immunofluorescence techniques to evaluate liver pathology. We assessed protein expression via Western blotting. We detected abnormal biliary ducts consistent with ductal plate abnormalities, as well as a greatly increased proliferation of cholangiocytes in the Pkhd1del3-4/del3-4 mice. CFTR was present in the apical membrane of cholangiocytes and increased in the Pkhd1del3-4/del3-4 mice, consistent with a role for apically located CFTR in enlarged bile ducts. Interestingly, we also found CFTR in the primary cilium, in association with polycystin (PC2). Localization of CFTR and PC2 and overall length of the cilia were increased in the Pkhd1del3-4/del3-4 mice. In addition, several of the heat shock proteins; 27, 70, and 90 were upregulated, suggesting that global changes in protein processing and trafficking had occurred. We found that a deficit of FPC leads to bile duct abnormalities, enhanced cholangiocyte proliferation, and misregulation of heat shock proteins, which all returned toward wild type (WT) values following VX-809 treatment. These data suggest that CFTR correctors can be useful as therapeutics for ARPKD. Given that these drugs are already approved for use in humans, they can be fast-tracked for clinical use.NEW & NOTEWORTHY ARPKD is a multiorgan genetic disorder resulting in newborn morbidity and mortality. There is a critical need for new therapies to treat this disease. We show that persistent cholangiocytes proliferation occurs in a mouse model of ARPKD along with mislocalized CFTR and misregulated heat shock proteins. We found that VX-809, a CFTR modulator, inhibits proliferation and limits bile duct malformation. The data provide a therapeutic pathway for strategies to treat ADPKD.