In Vitro and In Vivo Assessment of PEGylated PEI for Anti-IL-8/CxCL-1 siRNA Delivery to the Lungs

Abstract

<jats:p>Inhalation offers a means of rapid, local delivery of siRNA to treat a range of autoimmune or inflammatory respiratory conditions. This work investigated the potential of a linear 10 kDa Poly(ethylene glycol) (PEG)-modified 25 kDa branched polyethyleneimine (PEI) (PEI-LPEG) to effectively deliver siRNA to airway epithelial cells. Following optimization with anti- glyceraldehyde 3-phosphate dehydrogenase (GAPDH) siRNA, PEI and PEI-LPEG anti-IL8 siRNA nanoparticles were assessed for efficacy using polarised Calu-3 human airway epithelial cells and a twin stage impinger (TSI) in vitro lung model. Studies were then advanced to an in vivo lipopolysaccharide (LPS)-stimulated rodent model of inflammation. In parallel, the suitability of the siRNA-loaded nanoparticles for nebulization using a vibrating mesh nebuliser was assessed. The siRNA nanoparticles were nebulised using an Aerogen® Pro vibrating mesh nebuliser and characterised for aerosol output, droplet size and fine particle fraction. Only PEI anti-IL8 siRNA nanoparticles were capable of significant levels of IL-8 knockdown in vitro in non-nebulised samples. However, on nebulization through a TSI, only PEI-PEG siRNA nanoparticles demonstrated significant decreases in gene and protein expression in polarised Calu-3 cells. In vivo, both anti-CXCL-1 (rat IL-8 homologue) nanoparticles demonstrated a decreased CXCL-1 gene expression in lung tissue, but this was non-significant. However, PEI anti-CXCL-1 siRNA-treated rats were found to have significantly less infiltrating macrophages in their bronchoalveolar lavage (BAL) fluid. Overall, the in vivo gene and protein inhibition findings indicated a result more reminiscent of the in vitro bolus delivery rather than the in vitro nebulization data. This work demonstrates the potential of nebulised PEI-PEG siRNA nanoparticles in modulating pulmonary inflammation and highlights the need to move towards more relevant in vitro and in vivo models for respiratory drug development.</jats:p>

Aerosol Delivery of Surfactant Liposomes for Management of Pulmonary Fibrosis: An Approach Supporting Pulmonary Mechanics

“…The upper (stage I) and lower (stage II) chambers of TSI were filled with methanol, 7 and 30 mL, respectively [20]. The liposomal naringin (1 mg/mL, 5 mL) was aerosolized using an Aeroneb ® Lab micropump nebulizer fitted at the entrance of TSI [21]. During nebulization, a DFM 2000 flow meter and an HCP5 vacuum pump (Copley Scientific, Nottingham, UK) were utilized to maintain a 60 L/min airflow rate inside the impinger.…”

“…Groups II, III, and IV were given a single dosage of 0.1 mL bleomycin (5 mg/kg, 5 mg/mL in 0.9 percent w/v normal saline, intratracheal) to induce pulmonary fibrosis. [21]. The process was carried out in the same way for the NC group of normal control rats, with the difference that saline was used instead of bleomycin.…”

“…In pulmonary fibrosis, the activation of macrophages and massive influx of neutrophils cause lung alveolitis and epithelial cells deterioration [21]. These cells affect alveolar/capillary barrier permeability that causes edema.…”

“…Inhalation therapy with naringin thrice a week significantly reduced total cell count, neutrophils as wel as lymphocytes compared to the diseased control (DC) group. Pristine naringin could In pulmonary fibrosis, the activation of macrophages and massive influx of neutrophils cause lung alveolitis and epithelial cells deterioration [21]. These cells affect alveolar/capillary barrier permeability that causes edema.…”

Investigation of water‐insoluble hydrophobic polyethylenimines as RNAi vehicles in chronic myeloid leukemia therapy

“…Although the transfection efficiency of PEI enhances with increasing molecular weight (MW), 32 it is accompanied by a proportional increase in the cytotoxicity 33 due to the higher cationic charge density. Attempts to screen the positive charge of high MW PEI by derivatizing it with small hydrophobic moieties, 34,35 peptides, 36,37 and polyethylene glycol (PEG) 38,39 were successful in lowering toxicity and improving siRNA delivery. Low MW PEIs are not effective delivery vehicles but can be transformed into effective carriers by substituting hydrophobic groups onto the polymer 40,41 .…”