Abstract / Summary
Chronic kidney disease (CKD) lacks effective therapeutic interventions targeting its fundamental pathological mechanisms. TWIST1 is a basic helix-loop-helix transcription factor that is a critical regulator in kidney fibrosis pathogenesis. We used genome-wide CRISPR-Cas9 screening to delineate the role of TWIST1 in ferroptosis and explored the impact of an inflammation-responsive lipid nanoparticle (LTH-TK-LNP, a Kim-1-targeted reactive oxygen species-responsive lipid nanoparticle platform that releases si-RNA targeting TWIST1 specifically in inflamed renal tissue) in mouse models of CKD. CRISPR-Cas9 screening showed that TWIST1 is significantly upregulated under hypoxic conditions and drives ferroptosis by suppressing oxidative phosphorylation, leading to mitochondrial dysfunction, lipid peroxidation, and accumulation of reactive oxygen species. In vitro and in vivo studies demonstrated superior targeting of LTH-TK-LNP to injured kidneys compared to unmodified nanoparticles. In CKD mouse models, LTH-TK-LNP treatment significantly reduced TWIST1 expression, attenuated tubular damage and interstitial fibrosis, suppressed lipid peroxidation and the expression of inflammatory cytokines, and preserved mitochondrial integrity. Safety assessments revealed no evidence of hepatotoxicity, nephrotoxicity over a 14-day observation period. This study identifies TWIST1 as a master regulator connecting metabolic dysfunction to ferroptosis in renal tubular epithelial cells during CKD progression. Furthermore, we reveal the significant therapeutic potential of targeting TWIST1 and lay the preclinical foundation for kidney-targeted delivery strategies. Graphical Abstract. Schematic illustration of the LTH-TK-LNP nanoplatform for targeted si-TWIST1 delivery to treat chronic kidney disease. LTH-TK-LNPs are assembled via co-incubation and extrusion using a ROS-responsive, Kim-1-targeting lipid (DSPE-TK-PEG-LTHVVWL) and standard lipid components. Upon intravenous injection, the nanoparticles specifically accumulate in injured renal tubules. Release of si-TWIST1 silences TWIST1 expression, which restores oxidative phosphorylation and mitochondrial function. This metabolic reprogramming reduces ROS accumulation and lipid peroxidation, ultimately suppressing ferroptosis and attenuating renal fibrosis.