# APE1/Ref‐1 inhibition via APX3330 lowers monocyte/macrophage infiltration without ameliorating the structure and function of dystrophic mdx hindlimb muscles

## TL;DR

A drug that inhibits APE1/Ref-1 reduces inflammation in a mouse model of Duchenne muscular dystrophy but does not improve muscle function or structure.

## Contribution

The study shows that APE1/Ref-1 inhibition reduces monocyte/macrophage infiltration in dystrophic muscles without improving muscle function.

## Key findings

- APE1/Ref-1 protein levels were higher in dystrophic mdx mice compared to wild type.
- APX3330 treatment reduced CD68-positive monocyte/macrophage infiltration by 47% in dystrophic EDL muscles.
- Muscle contractile function and pathology were not improved by APE1/Ref-1 inhibition.

## Abstract

Chronic inflammation and oxidative stress exacerbate muscle wasting and weakness in Duchenne muscular dystrophy (DMD). Apurinic/apyrimidinic endonuclease 1/redox factor‐1 (APE1/Ref‐1) regulates transcription factors involved in inflammatory and oxidative stress pathways. APE1/Ref‐1 is an emerging therapeutic target in inflammatory conditions. This study aimed to investigate the effects of APX3330, a small molecule inhibitor of APE1's Ref‐1 on mdx mouse pathology, a model of DMD. Six‐week‐old mdx mice and wild type (WT) C57Bl/10 mice were treated with APX3330 (25 mg·kg−1) or vehicle for 6 weeks. Ex vivo contractile function, histological and biochemical analysis were performed in extensor digitorum longus (EDL) and soleus muscles. APE1/Ref‐1 protein was greater in mdx hindlimb muscles compared to WT (p < 0.0001) and APE1/Ref‐1 protein abundance was not altered by treatment with APX3330. In dystrophic EDL muscles, APX3330 treated mice had fewer (47%) infiltrating CD68‐positive monocytes/macrophages (p < 0.05) compared to vehicle‐treated mdx mice. Markers of oxidative stress, NRF2/KEAP‐1, were unchanged, yet phospho‐NF‐κB abundance was higher with treatment (p < 0.01). APX3330 treatment neither improve force output and fatiguability of isolated hindlimb muscles, nor affect muscle pathology. As APE1/Ref‐1 inhibition modestly lowered inflammation, with no improved contractile function, targeting solely inflammation and oxidative stress in 6‐week‐old mdx mice appears insufficient.