KD025 Shifts Pulmonary Endothelial Cell Bioenergetics and Decreases Baseline Lung Permeability

Abstract

KD025 is a ROCK2 inhibitor currently being tested in clinical trials for the treatment of fibrotic lung diseases. The therapeutic effects of KD025 are partly due to its inhibition of profibrotic pathways and fat metabolism. However, whether KD025 affects pulmonary microvascular endothelial cell (PMVEC) function is unknown, despite evidence that alveolar-capillary membrane disruption constitutes major causes of death in fibrotic lung diseases. We hypothesized that KD025 regulates PMVEC metabolism, pH, migration, and survival, a series of interrelated functional characteristics that determine pulmonary barrier integrity. We used PMVECs isolated from Sprague Dawley rats. KD025 dose-dependently decreased lactate production and glucose consumption. The inhibitory effect of KD025 was more potent compared with other metabolic modifiers, including 2-deoxy-glucose, extracellular acidosis, dichloroacetate, and remogliflozin. Interestingly, KD025 increased oxidative phosphorylation, whereas 2-deoxy-glucose did not. KD025 also decreased intracellular pH and induced a compensatory increase in anion exchanger 2. KD025 inhibited PMVEC migration, but fasudil (nonspecific ROCK inhibitor) did not. We tested endothelial permeability in vivo using Evans Blue dye in the bleomycin pulmonary fibrosis model. Baseline permeability was decreased in KD025-treated animals independent of bleomycin treatment. Under hypoxia, KD025 increased PMVEC necrosis as indicated by increased lactate dehydrogenase release and propidium iodide uptake and decreased ATP; it did not affect Annexin V binding. ROCK2 knockdown had no effect on PMVEC metabolism, pH, and migration, but it increased nonapoptotic caspase-3 activity. Together, we report that KD025 promotes oxidative phosphorylation; decreases glycolysis, intracellular pH, and migration; and strengthens pulmonary barrier integrity in a ROCK2-independent manner.

Belumosudil: First Approval

“…In human fibrosis-derived smooth muscle cells, belumosudil reduced connective tissue growth factor mRNA and remodelled the actin cytoskeleton [ 12 ]. In the lung endothelium, belumosudil promoted oxidative phosphorylation, inhibited glycolysis, intracellular pH and migration, and strengthened barrier integrity [ 13 ].

Chemical structure of belumosudil

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Insight Into Rho Kinase Isoforms in Obesity and Energy Homeostasis

“…Recent studies explored therapeutic potential of KD025 in metabolic disease including renal fibrosis in the unilateral ureteral obstruction mice ( 147 ), arteriosclerosis and vascular fibrosis in a diabetic mouse model ( 148 ). Their therapeutic effects are partly due to the inhibition of profibrotic pathways and fat metabolism in both ROCK2-dependent and independent manners ( 149 ). Moreover, KD025 has been shown to inhibit adipogenesis in human adipose-derived stem cells, 3T3-L1 preadipocytes and human orbital fibroblasts through a ROCK2-independent mechanism ( 150 – 152 ), by inhibiting casein kinase 2 ( 153 ).…”

Roles of the RhoA-ROCK Signaling Pathway in the Endothelial H2S Production and Vasodilation in Rat Cerebral Arteries

“…Figure 5 shows that both the RhoA inhibitor CCG-1423 37 and the ROCK inhibitor KD025 38 induced significant vasodilation in KCl - precontracted rat CBA compared to that in the vehicle group, suggesting that inhibition of the RhoA-ROCK signaling pathway could produce vasodilation in rat CBA. Figure 5 also shows that CCG-1423 or KD025 at 1 × 10 –6.5 mol/L neither had an effect on resting rat CBA nor relaxed KCl - precontracted rat CBA.…”