Pre-clinical evaluation of three non-viral gene transfer agents for cystic fibrosis after aerosol delivery to the ovine lung

Abstract

We use both large and small animal models in our pre-clinical evaluation of gene transfer agents (GTAs) for cystic fibrosis (CF) gene therapy. Here, we report the use of a large animal model to assess three non-viral GTAs: 25 kDa-branched polyethyleneimine (PEI), the cationic liposome (GL67A) and compacted DNA nanoparticle formulated with polyethylene glycol-substituted lysine 30-mer. GTAs complexed with plasmids expressing human cystic fibrosis transmembrane conductance regulator (CFTR) complementary DNA were administered to the sheep lung (n¼8 per group) by aerosol. All GTAs gave evidence of gene transfer and expression 1 day after treatment. Vector-derived mRNA was expressed in lung tissues, including epithelial cell-enriched bronchial brushing samples, with median group values reaching 1-10% of endogenous CFTR mRNA levels. GL67A gave the highest levels of expression. Human CFTR protein was detected in small airway epithelial cells in some animals treated with GL67A (two out of eight) and PEI (one out of eight). Bronchoalveolar lavage neutrophilia, lung histology and elevated serum haptoglobin levels indicated that gene delivery was associated with mild local and systemic inflammation. Our conclusion was that GL67A was the best non-viral GTA currently available for aerosol delivery to the sheep lung, led to the selection of GL67A as our lead GTA for clinical trials in CF patients.

Synthetic vectors for gene delivery: An overview of their evolution depending on routes of administration.

“…Low MW PEI does not induce inhibition of luciferase expression compared to PEI polymers with a high MW which can been explained by the cytotoxicity associated with high MW PEI [54]. Significant gene expression in lungs was detected in rodents [55] and in sheep [56], specifically in ciliated epithelial cells of the whole murine respiratory system [39], and alveolar type I pneumocytes [39,57]. In parallel, the 22 kDa linear PEI used for systemic administration can effectively transfect several organs in neonatal mice [58].…”

“…To overcome all obstacles mentioned previously, progress is needed. Recently extensive studies have been conducted on the aerosolisation of complexes in order to circumvent these difficulties [54,56]. Nonviral gene delivery was assessed in large animals using pDNA complexed with either 25 kDa-branched PEI, cationic liposome (GL67A) or polyethylene glycol-substituted lysine 30-mer.…”

Highly compacted biodegradable DNA nanoparticles capable of overcoming the mucus barrier for inhaled lung gene therapy

“…4B). Among the conventional DNA-NPs, PLL-CPs have demonstrated a gene transfer level on par with AAV serotype 2 in a clinical trial (39,57), and PEI-CPs have provided in vivo transgene expression comparable to the only gene delivery system being tested currently in a clinical trial for CF gene therapy (GL67A) (58,59). PBAE-MPPs mediated a statistically significant ∼25-fold higher overall transgene expression level compared with these gold standard nonviral gene vectors (Fig.…”

Airway Deposition of Nebulized Gene Delivery Nanocomplexes Monitored by Radioimaging Agents

“…Although this study was designed to focus mainly on methods of administration and the biodistribution of the RTN radiovector, rather than on optimizing transfection efficiency, we were encouraged to find significant levels of transgene expression in the conducting airway ciliated epithelium at the relatively low dose of plasmid DNA administered. The pigs were nebulized with a 6-ml suspension containing only 1 mg of pCpG-free lacZ DNA, in contrast with a recent report where gene transfer to the lungs of sheep was mediated by a liposome containing polyethylene glycol (GL67), administering 20 ml of the formulation containing 52.8 mg of human CFTR plasmid DNA (48).…”