Macrophage-specific targeting of histone demethylases with small-molecule inhibitors suppresses inflammatory response...

Macrophages are versatile immune cells, with the ability to respond to varied intrinsic and extrinsic cues and transition between inflammatory and proreparative phenotypes. A complex network of epigenetic processes, such as DNA methylation, histone methylation, and acetylation, plays key roles in modulating macrophage polarization and inflammatory gene expression. Transcriptional analysis in patients with respiratory failure, long COVID-19, and influenza revealed an augmented expression of chromatin modifiers, including histone demethylases, broadly defined as lysine demethylases (KDMs), in lung macrophages. To investigate the role of KDMs in inflammation, we screened a panel of small-molecule inhibitors of chromatin modifiers for their efficacy in inducing anti-inflammatory macrophage polarization in vitro. We demonstrate that pretreatment with the broad-spectrum KDM inhibitor n-octyl-IOX1 and KDM5-specific inhibitor PB-IT resulted in a significant decrease of lipopolysaccharide (LPS)-induced expression of the inflammatory genes Il1b, Il6, Tnfa, and iNos in bone marrow-derived macrophages. Subsequent RNA sequencing and CUT&RUN analyses revealed that LPS activation led to distinct transcriptomic and epigenomic alterations, including expression of master transcription factors BLIMP-1 and GFI1, whereas n-octyl-IOX1 and PB-IT treatments rewired these regulatory networks, thereby impeding inflammatory gene expression. To further probe the merit of KDM inhibition in perturbing macrophage-mediated inflammation in vivo, we delivered n-octyl-IOX1 selectively to macrophages in mice using cell-specific, targeted lipidoid nanoparticles. n-octyl-IOX1 encapsulated nanoparticles significantly diminished LPS-mediated peritoneal macrophage expansion and inflammatory gene expression in this cell population, underscoring the importance of macrophage-specific targeting of KDMs with small-molecule inhibitors in inflammatory disease.