Therapeutic administration of inhaled INS1009, a treprostinil prodrug formulation, inhibits bleomycin-induced pulmonary fibrosis in rats

• Published in: Pulmonary pharmacology & therapeutics Vol. 49; pp. 95 - 103
• Main Authors: Corboz, Michel R., Zhang, Jimin, LaSala, Daniel, DiPetrillo, Keith, Li, Zhili, Malinin, Vladimir, Brower, Jeremy, Kuehl, Philip J., Barrett, Ted E., Perkins, Walter R., Chapman, Richard W.
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 01.04.2018
• Subjects: Administration, Inhalation; Animals; Antihypertensive Agents - administration & dosage; Antihypertensive Agents - pharmacokinetics; Antihypertensive Agents - pharmacology; Bleomycin - administration & dosage; Bleomycin - toxicity; Cells, Cultured; Disease Models, Animal; Dose-Response Relationship, Drug; Epoprostenol - administration & dosage; Epoprostenol - analogs & derivatives; Epoprostenol - pharmacokinetics; Epoprostenol - pharmacology; Fibroblasts; Fibroblasts - drug effects; Fibroblasts - metabolism; Hexadecyl-treprostinil; Humans; Hydroxyproline - metabolism; Idiopathic Pulmonary Fibrosis - drug therapy; Idiopathic Pulmonary Fibrosis - physiopathology; Inhalation treatment; INS1009; Lipids - chemistry; Male; Nanoparticles; Prodrugs; Pulmonary fibrosis; Pyridones - pharmacology; Rats; Rats, Inbred F344; Real-Time Polymerase Chain Reaction; Fibroblasts; Rats; Hexadecyl-treprostinil; Pulmonary fibrosis; Inhalation treatment; INS1009
• ISSN: 1094-5539; 1522-9629
• DOI: 10.1016/j.pupt.2018.01.012

Idiopathic pulmonary fibrosis is a progressive and lethal disease and while there are now two approved drugs (Esbriet® and Ofev®) additional effective treatments are still needed. Recently, prostacyclin analogs such as iloprost and treprostinil (TRE) have been shown to exert some protection against bleomycin-induced pulmonary fibrosis in mice when administered in a prophylactic regimen. In this study, we evaluated the effect of the inhaled treprostinil prodrug hexadecyl-treprostinil (C16TR) formulated in a lipid nanoparticle (INS1009) administered therapeutically in a fibrotic rat model. Male Fischer 344 rats challenged with intra-tracheal saline instillation were then treated with daily inhaled phosphate buffered saline (PBS) while rats challenged with bleomycin sulfate (3.5–4.0 mg/kg) instillation were treated with either daily inhaled PBS, daily inhaled INS1009 (10, 30, or 100 μg/kg), or twice-daily orally with the anti-fibrotic compound pirfenidone (100 mg/kg). Dosing started on day 10 post-bleomycin challenge and continued until day 27 after bleomycin. Lungs were harvested 24 h after the last dose of treatment for evaluation of lung hydroxyproline content and pulmonary histology. Lung hydroxyproline content increased from 421 μg/lung lobe in saline challenged and PBS treated animals to 673 μg/lung lobe in bleomycin challenged and PBS treated rats. Treatment of bleomycin challenged rats with 10, 30, or 100 μg/kg INS1009 dose-dependently reduced lung hydroxyproline content to 563, 501, and 451 μg/lung lobe, respectively, and pirfenidone decreased hydroxyproline content to 522 μg/lung lobe. Histologically, both INS1009 (100 μg/kg) and pirfenidone (100 mg/kg) reduced the severity of subepithelial fibrosis. Single dose pharmacokinetic (PK) studies of inhaled INS1009 in bleomycin challenged rats showed dose-dependent increases in lung C16TR concentration and plasma TRE on day 10 post-bleomycin challenge. Multiple dose PK studies of inhaled INS1009 showed dose-dependent increases only in lung C16TR concentration on day 27 post-bleomycin challenge. We also investigated the effects of TRE on the cytokine transforming growth factor-β1 (TGF-β1)-stimulated collagen gene and protein expressions in cultured human lung fibroblasts, assessed by real-time PCR and Sirius Red staining, respectively. In human fibroblasts, TRE (0.001–10 μM) inhibited TGF-β1 (20 ng/mL)-induced expression of collagen mRNA and protein in a concentration-dependent manner. These results demonstrated that inhaled INS1009, administered in a therapeutic dosing paradigm, dose-dependently (10–100 μg/kg) inhibited bleomycin-induced pulmonary fibrosis in rats. This effect may involve direct actions of TRE in suppressing collagen expression in lung fibroblasts.