Differential Effects of Halofuginone Enantiomers on Muscle Fibrosis and Histopathology in Duchenne Muscular Dystrophy.

Sharon Mordechay,Orna Halevy,Shaun Smullen,Mark Pines,Paul Evans,Olga Genin

doi:10.3390/ijms22137063

Abstract

Progressive loss of muscle and muscle function is associated with significant fibrosis in Duchenne muscular dystrophy (DMD) patients. Halofuginone, an analog of febrifugine, prevents fibrosis in various animal models, including those of muscular dystrophies. Effects of (+)/(−)-halofuginone enantiomers on motor coordination and diaphragm histopathology in mdx mice, the mouse model for DMD, were examined. Four-week-old male mice were treated with racemic halofuginone, or its separate enantiomers, for 10 weeks. Controls were treated with saline. Racemic halofuginone-treated mice demonstrated better motor coordination and balance than controls. However, (+)-halofuginone surpassed the racemic form’s effect. No effect was observed for (−)-halofuginone, which behaved like the control. A significant reduction in collagen content and degenerative areas, and an increase in utrophin levels were observed in diaphragms of mice treated with racemic halofuginone. Again, (+)-halofuginone was more effective than the racemic form, whereas (−)-halofuginone had no effect. Both racemic and (+)-halofuginone increased diaphragm myofiber diameters, with no effect for (−)-halofuginone. No effects were observed for any of the compounds tested in an in-vitro cell viability assay. These results, demonstrating a differential effect of the halofuginone enantiomers and superiority of (+)-halofuginone, are of great importance for future use of (+)-halofuginone as a DMD antifibrotic therapy.

Highlights

Muscular dystrophies (MDs) are a heterogeneous group of genetic disorders characterized by progressive loss of muscle strength and integrity
Recent studies have reported that utrophin is expressed in an inverse relationship with fibrosis in muscles of Duchenne MD (DMD) patients [8] and in the diaphragm of mdx mice, a mouse model of DMD [8,9]
We previously reported the de-novo synthesis of the two optically active enantiomers of halofuginone (Figure 1) [37], and our hypothesis is that the single enantiomers of halofuginone affect muscle function and histopathology of mdx mice differently

Summary

Introduction

Muscular dystrophies (MDs) are a heterogeneous group of genetic disorders characterized by progressive loss of muscle strength and integrity. Disease onset is in early childhood, and it is characterized by the presence of high levels of muscle creatine kinase, muscle hypertrophy and necrotic myofibers. This is accompanied by repeated muscle degeneration–regeneration cycles and progressive replacement of the contractile muscle tissue with collagenous (mainly collagen type I) fibrotic tissue [1,2]. Fibrosis is evident mainly in the diaphragm, the most affected muscle at early stages of DMD, and its prevention enhances muscle repair and function [4]. Recent studies have reported that utrophin is expressed in an inverse relationship with fibrosis in muscles of DMD patients [8] and in the diaphragm of mdx mice, a mouse model of DMD [8,9]

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Discussion

Conclusion