Michael J. Bollong, Baiyuan Yang, Naja Vergani, Brittney A. Beyer, Emily N. Chin, Claudio Zambaldo, Danling Wang, Arnab K. Chatterjee, Luke L. Lairson, and Peter G. Schultz.

CELL BIOLOGY

Significance

The treatment of fibrosis remains a critically important unmet medical need, as nearly 45% of all natural deaths in the Western world are attributed to chronic fibroproliferative disease complications. Fibrosis is characterized by the excessive deposition of extracellular matrix proteins by resident fibroblast-derived myofibroblasts. From an imaging-based screen, we identified the antifungal drug itraconazole as an inhibitor of myofibroblast transdifferentiation from multiple resident fibroblast populations. A derivative of this drug was found to inhibit fibrotic disease progression in mouse models of lung, liver, and skin fibrosis, demonstrating that inhibiting differentiation to the myofibroblast cell state is a practical strategy to treat a wide range of fibrosis-related diseases.

 Abstract

Fibrosis, a disease in which excessive amounts of connective tissue accumulate in response to physical damage and/or inflammatory insult, affects nearly every tissue in the body and can progress to a state of organ malfunction and death. A hallmark of fibrotic disease is the excessive accumulation of extracellular matrix-secreting activated myofibroblasts (MFBs) in place of functional parenchymal cells. As such, the identification of agents that selectively inhibit the transdifferentiation process leading to the formation of MFBs represents an attractive approach for the treatment of diverse fibrosis-related diseases. Herein we report the development of a high throughput image-based screen using primary hepatic stellate cells that identified the antifungal drug itraconazole (ITA) as an inhibitor of MFB cell fate in resident fibroblasts derived from multiple murine and human tissues (i.e., lung, liver, heart, and skin). Chemical optimization of ITA led to a molecule (CBR-096-4) devoid of antifungal and human cytochrome P450 inhibitory activity with excellent pharmacokinetics, safety, and efficacy in rodent models of lung, liver, and skin fibrosis. These findings may serve to provide a strategy for the safe and effective treatment of a broad range of fibrosis-related diseases.

See: http://www.pnas.org/content/114/18/4679.full

PNAS May 2, 2017; vol.114; no.18: 4679–4684

Figure 1: ITA inhibits MFB transdifferentiation. (A) Images of rat HSCs treated with ITA (0.5 μM) or SB-431542 (10 μM) in MFB formation conditions immunostained for αSMA (Scale bar, 100 μm). (B) Image analysis quantification of rat HSCs treated with ITA in MFB formation conditions (n = 9, mean and SEM). (C) Western blot analyses of αSMA and GFP from COL1–GFP HSCs subjected to MFB conditions and treated with ITA. (D) qRT-PCR analyses of MFB identity genes from human lung fibroblasts (HLFs) treated for 72 h (ITA, 0.3 μM; n = 3, mean and SD; ***P < 0.0005, t test). (E) Heatmap displaying the log₂ fold change from untreated controls of all active transcripts (Left) or core MFB genes (Right) as measured by RNA-seq from HLFs treated with TGF-β1+DMSO or ITA (300 nM) for 72 h. (F) Enrichment P values from DAVID analyses for selected GO categories of genes up-regulated by TGF-β1 but suppressed by ITA treatment.

PNAS 2017 114 (18) 4679-4684; published ahead of print April 17, 2017, doi:10.1073/pnas.1702750114