Abstract 13446: Role of the Epigenetic Factor Ezh2 in Pulmonary Arterial Hypertension.

Background: Pulmonary arterial hypertension (PAH) is a vascular remodeling disease characterized by persistent elevation of pulmonary vascular resistance, which results in a striking increase in right ventricular (RV) afterload and subsequent failure. Pulmonary artery (PA) smooth muscle cells (PASMCs) from PAH patients exhibit a cancer-like phenotype with increased glycolysis (Hexokinase 2 activity) and reduced glucose oxidation (inhibition of pyruvate dehydrogenase (PDH)) promoting cell proliferation and apoptosis-resistant contributing to obliteration of the PA lumen. Emerging evidence have demonstrated a critical role for the histone methyltransferase EZH2 in cancer cell survival and several EZH2 inhibitors are now clinically investigated. However, its role in PAH remains unknown. We hypothesized that EZH2 expression is upregulated in PAH-PASMCs leading to PA remodeling. Methods and Results: Using Western blot (WB), we showed that EZH2 is upregulated in lungs, distal PAs and isolated PASMCs from PAH patients (p<0.01, n=10) compared to controls. Consistently, increased EZH2 expression was found in distal PAs from animal models mimicking PAH (monocrotaline and Sugen/Hypoxia). Using proteomic LC-MSMS approach followed by Maxquant/Andromeda analysis in PASMCs isolated from 4 PAH patients, we identified 139 proteins differentially expressed (27 up-regulated ≥ 1.2 or and 112 down-regulated ≤ 0.84; p ≤ 0.05) following EZH2 knockdown. Results of KEGG pathway analysis revealed that the tricarboxylic acid (TCA) cycle, carbon metabolism and metabolic pathways were the top significant pathways (FDR p-value <10-4). This suggests that EZH2 is important for regulating bioenergetic metabolism in PAH-PASMCs. In agreement with this, pharmacological (GSK126 or EPZ-6438) or molecular (siEZH2) inhibition of EZH2 leads to reduced expression (WB) of Hexokinase 2, pPDH and PDH. This was associated with a significant reduction in PAH-PASMC proliferation (Ki67 labeling and EdU assay, p<0.001) and resistance to apoptosis (Annexin V assay p<0.001). Conclusion: We provide evidence that EZH2 is overexpressed in human PAH and implicated in the pro-proliferative and anti-apoptotic phenotype of PAH-PASMCs. [ABSTRACT FROM AUTHOR]