Faculty Bibliography

Both animal and cell culture data support a beneficial role of interferon gamma in the amelioration/treatment of pulmonary fibrosis. Despite this, two large randomized clinical trials failed to show a clinical response to subcutaneous interferon gamma. This negative response was associated with low/non-detectable levels of interferon gamma levels in the BAL fluid. Data from normal volunteers supports the inability of interferon gamma delivered subcutaneously to reach the lung epithelial lining fluid or activate BAL cells. We completed a phase I clinical trial with inhaled interferon gamma for 80 weeks in 10 patients with IPF. Prior studies in IPF patients suggest that aerosol interferon gamma is not detectable in BAL fluid if obtained > 24 hours after nebulization. Patients tolerated treatment well with stabilization in FVC and DLCO over 80 weeks. BAL was obtained at baseline and between 1 and 4 hours after treatment with inhaled interferon gamma. BAL fluid showed significant increases in interferon-gamma levels (17.07 +/- 13.67 pg/ml to 1077 +/- 501.97 pg/ml p=0.062) This level was associated with immunomodulation of lung environment. Inhaled interferon gamma may be beneficial in the treatment of IPF

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease with few treatment options. Previous trials of systemic interferon- gamma (IFN-gamma) treatment in IPF were discouraging. Recently, in IPF patients, we demonstrated that inhaled IFN-gamma was safe with no systemic side effects and associated with significant improvement in DLCO. AIM: Using microarray and proteomic techniques, we examined potential mechanisms of local anti-fibrotic effects of inhaled IFN-gamma by testing bronchoalveolar fluid (BALF) for gene and protein expression. Methods: Ten patients with IPF received inhaled IFN- gamma three times a week for 50 weeks. BAL was performed before and during therapy. Gene expression was measured on RNA samples from the BALF cellular component using Affymetrix human U133Av2 chips. Protein expression was studied using a label-free LC-MS based approach with systems biology to identify signaling pathways and regulatory nodes within protein interaction networks that explain the mechanism of anti-fibrotic effects of inhaled IFN-gamma. Zymography, Western blot and Sircol assay were used to validate the differential protein expression data. Results: Microarray and proteomic studies demonstrated that IFN-gamma treatment down regulates multiple signal pathways involved in T cell and B cell immunity, actin cytoskeleton, apoptosis, MMPs, protease, chemokine, and PI3 signaling pathway. BALF zymography and western blot studies showed that IFN-gamma inhibited MMP-9, MMP-1 and Osteopontin. IFN- gamma treatment resulted in amarked decrease in BALF collagen content. Conclusion: Inhaled INF-c has potent anti-fibrotic effects via multiple signaling pathways, immune modulatory effects and MMP inhibition. Inhaled IFN-gamma has potential as a novel therapy for IPF

Background: Therapies targeting the inflammatory process revealed inconsistent results in Idiopathic Pulmonary Fibrosis (IPF). Aerosolized Interferon Gamma (IFN-y) has been proposed as a novel therapy. Additionally, changes in the host airway microbiome have been associated with inflammatory tone and, in IPF, may be associated with disease progression.We evaluate whether treatment with aerosolized IFN-y impacts the lower airway microbiome or host immune phenotype. Methods: Patients with IPF were enrolled in an aerosolized IFNy trial. Patients underwent a bronchoscopy at baseline and after 6 months. 16S rRNA sequencing of BAL samples were used to evaluate the lower airway microbiome. Cytokines were measured by Luminex on plasma, BAL fluid and BAL cell supernatant. Findings: IFN-y treatment did not change alpha or beta diversity of the lung microbiome and few taxonomic changes in low abundant taxa occurred (Figure 1). The measured cytokines were unchanged in plasma, however, there was an increase in IFN-y and a decrease in Fit 3 Ligand, IFN-A2 and IL- 5 in BAL cell supernatant, and TNF-B in BAL. Multiple correlations between taxa and inflammatory cytokines were found. Network analysis showed correlations were more abundant between lung microbial taxa and local inflammatory cytokines (either in BAL or by ex vivo BAL cell production) than between lung microbial taxa and systemic inflammatory cytokines. Importantly, analysis showed persistent of these association post IFN treatment. Interpretation: This data suggests that the lower airway microbiome has an independent effect on host immune tone and thus may provide a novel therapeutic target for treatment of IPF

Parenteral IFN-gamma was unsuccessful as a treatment for pulmonary fibrosis. Inhaled IFN-gamma targeted to the lungs may be more effective. Our patient, a 56-year-old male with biopsy proven usual interstitial pneumonia (UIP) and declining pulmonary function tests (PFTs) was initially diagnosed with idiopathic pulmonary fibrosis (IPF). He enrolled in a 2-year research protocol and was treated with inhaled IFN-gamma (100 mug, Actimmune, Horizon Pharma, Deerfield, IL) 3 times per week. After completion of the protocol, he was able to secure the drug and continued therapy for a total of 7 years. He felt better, returning to work. His only complaint was transient cough during inhalation. PFTs improved (e.g., DLCO, 58% at baseline, 81% at 2 years, 69% currently). Clinical monitoring showed preserved exercise tolerance and stable CT scans. He was ultimately diagnosed (year 5) with scleroderma-like connective tissue disease after he developed sclerodactyly and a positive antinuclear antibody. Inhaled IFN-gamma was well tolerated for 7 years and may stabilize fibrotic lung disease.

BACKGROUND: Using data from a previously reported phase 2 safety trial, testing inhaled interferon gamma (IFN-gamma) for IPF, we analyzed effects on full pulmonary function tests (PFTs) for efficacy before and after therapy and designed a randomized controlled trial of inhaled IFN-gamma to treat IPF. METHODS: Ten patients with IPF had received inhaled IFN-gamma (Actimmune, InterMune) for 80 weeks. Full PFTs were available 20-50 weeks before Rx and monthly during Rx. Eighty-nine observations were used in the analysis. Linear mixed models for modeling longitudinal data were used to test if the PFT change over time was significantly different before and after IFN-gamma. Autoregressive dependence structure with order one was consistently selected as the best one to model the intra-patient correlation over time. Normality assumption was confirmed. Significance level was set at 0.05. Using published literature and our data we performed a sample size calculation based on simulated data. RESULTS: The change over time in DLCO was significantly different before and after IFN-gamma treatment. DLCO decreased over time before treatment but increased after treatment (p-value=0.03). Changes in TLC, FRC, RV and FVC were not statistically significant. With a sample size of 60, a placebo controlled, randomized trial has about 90% power to detect a significant difference in the change rate of DLCO in the groups of patients treated with IFN-gamma vs placebo. CONCLUSIONS: DLCO was significantly improved following inhaled (IFN-gamma) as treatment for IPF. Our data suggest that previous studies utilizing parenteral IFN-gamma may have failed because of the mode of delivery. Future randomized, controlled, phase 3 trials, comparing the difference in PFT behavior (specifically DLCO) longitudinally may be more sensitive to drug effect and serve as a valuable clinical endpoint.