ATP-Citrate Lyase Inhibition Improves Chronic Kidney Disease Through Multiple Mechanisms

Abstract

ATP-citrate lyase (ACLY), upregulated in chronic kidney disease (CKD), catalyzes the synthesis of acetyl-coA from citrate. Acetyl-CoA is a vital precursor for lipid/cholesterol synthesis and histone acetylation that regulates gene expression. In renal cells, ACLY regulates fibrogenic, lipogenic and inflammatory gene expression; its inhibition reduced fibrosis in the unilateral ureteral obstruction (UUO) model. The ACLY metabolic by-product malonyl-coA is also an important inhibitor of fatty acid oxidation (FAO), and defective FAO in proximal tubular epithelial cells (PTEC) is now established as a major contributor to fibrosis. Here we tested the efficacy of a novel ACLY inhibitor on reducing fibrosis and its potential role in improving FAO in UUO. 8-week-old male C57BL/6J mice underwent UUO surgery and were treated orally with an ACLY inhibitor (EVT0185, Espervita Therapeutics) for 10 days. Kidneys were assessed by immunohistochemistry, immunoblotting, and RNAseq. Effects of ACLY inhibition were tested on the HK2 PTEC cell line and primary renal fibroblast responses to TGFβ1 (5ng/ml, 48h), a cytokine known to promote fibrosis and reduce FAO. Lipid accumulation was assessed by Oil Red O staining and LC/MS analysis. ACLY inhibition significantly and dose-dependently decreased fibrosis in the UUO model determined by trichrome, PSR, fibronectin, and α-smooth muscle actin (SMA) expression. ACLY inhibition decreased macrophage (F4/80) infiltration including that of the profibrotic M2 phenotype marked by CD206. RNAseq analysis showed upregulation of FAO-related hallmark pathways and reduction in inflammation pathways with ACLY inhibition. Defective FAO is known to result in PTEC apoptosis and lipid accumulation. ACLY inhibition reduced both apoptosis, as assessed by the presence of cleaved caspase 3, as well as lipid accumulation, with a particular decrease in cholesteryl esters. In HK2 cells and renal fibroblasts, TGFβ1-induced fibrotic protein expression was inhibited by ACLY inhibition, and lipid accumulation was reduced in PTECs. ACLY inhibition reduced renal fibrosis, apoptosis, and lipid accumulation in UUO mice. ACLY inhibition also prevented profibrotic responses to TGFβ1 in PTECs and fibroblasts. Current studies are ongoing to confirm beneficial effects on restoring FAO.