Chemical Proteomics and Phenotypic Profiling Identifies the Aryl Hydrocarbon Receptor as a Molecular Target of the Utrophin Modulator Ezutromid.

Isabel V.l Wilkinson ,Graham M Wynne ,Stephen G Davies ,Hannah L Dugdale ,Lee Moir ,Kelly Perkins ,Fraydoon Rastinejad ,Alexander Lomow ,Aini Vuorinen ,Maria Chatzopoulou ,Sebastian Monecke ,Jonathon M Tinsley ,Peter M Burch ,Marcus Geese ,Francis X Wilson ,Kay E Davies ,Shabaz Mohammed ,Sarah Squire ,P David Charles ,Angela J Russell

doi:10.1002/anie.201912392

Abstract

Duchenne muscular dystrophy (DMD) is a fatal muscle‐wasting disease arising from mutations in the dystrophin gene. Upregulation of utrophin to compensate for the missing dystrophin offers a potential therapy independent of patient genotype. The first‐in‐class utrophin modulator ezutromid/SMT C1100 was developed from a phenotypic screen through to a Phase 2 clinical trial. Promising efficacy and evidence of target engagement was observed in DMD patients after 24 weeks of treatment, however trial endpoints were not met after 48 weeks. The objective of this study was to understand the mechanism of action of ezutromid which could explain the lack of sustained efficacy and help development of new generations of utrophin modulators. Using chemical proteomics and phenotypic profiling we show that the aryl hydrocarbon receptor (AhR) is a target of ezutromid. Several lines of evidence demonstrate that ezutromid binds AhR with an apparent KD of 50 nm and behaves as an AhR antagonist. Furthermore, other reported AhR antagonists also upregulate utrophin, showing that this pathway, which is currently being explored in other clinical applications including oncology and rheumatoid arthritis, could also be exploited in future DMD therapies.

Highlights

Duchenne muscular dystrophy (DMD) is a fatal, X-linked muscle wasting disease that affects approximately 1 in 3500– 5000 boys.[1,2] DMD is caused by loss-of-function mutations in the dystrophin gene
Using chemical proteomics and phenotypic profiling we show that the aryl hydrocarbon receptor (AhR) is a target of ezutromid
Prototypical AhR agonist TCDD has been shown to decrease activity and abundance of transcriptional coactivator peroxisome proliferator-activated receptor-g coactivator-1a (PGC1a),[55] which stimulates utrophin expression at neuromuscular junctions.[56]. This dysregulation was removed by co-treatment with an AhR antagonist, leading to stabilisation of active PGC1a

Summary

Introduction

Duchenne muscular dystrophy (DMD) is a fatal, X-linked muscle wasting disease that affects approximately 1 in 3500– 5000 boys.[1,2] DMD is caused by loss-of-function mutations in the dystrophin gene. With an estimated one-third caused by spontaneous mutations.[3,4] Loss of dystrophin results in progressive muscle degeneration, with patients wheelchair-dependent by their early teens and average life expectancy reduced to the late 20 s to 30 s, due to heart and respiratory failure.[5,6]. Developments in the clinical standard of care have led to improvements in quality of life and longevity[7,8,9] but there is currently no available cure for DMD. Several therapeutic strategies have reached the clinic, including stop codon readthrough and exon skipping which target particular mutations, and gene therapy which faces multiple challenges, including viral production and immune response.[10,11]

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