Abstract Body (Do not enter title and authors here): Background: Pulmonary arterial hypertension (PAH) is a progressive disease characterized by vasoconstriction and remodeling of small pulmonary arteries (PAs) due to hyper-proliferation and apoptosis resistance of resident PA cells, the mechanisms of which are not completely understood. Specifically, the role of cell division cycle protein 20 (CDC20), a key regulator of cell cycle, canonically acting as an APC/C E3 ubiquitin ligase activator and substrate recruiter, is not known.
Goals: To determine the role of CDC20 in PA remodeling in PAH.
Methods: RNAseq, immunoblot, immunohistochemistry, proliferation, apoptosis assays; SU5416/Hypoxia (SuHx) mouse model of PH.
Results: CDC20 was over-expressed in lungs, small PAs, isolated PA smooth muscle cells (PASMCs) and adventitial fibroblasts (PAAFs), but not endothelial cells from PAH patients compared to non-diseased controls. siRNA CDC20 significantly reduced proliferation and induced apoptosis in PAH PASMCs and PAAFs. This effect was re-capitulated by CDC20 inhibitor apcin (blocks binding of CDC20 with substrates); however, pan-APC inhibitor proTAME had no effect. Over-expression of CDC20 in PAH PASMCs and PAAFs was correlated with over-accumulation of its substrate, pro-proliferative protein securin, and deficiency of pro-apoptotic protein Bcl-2 interacting mediator of cell death (BIM). Depletion of CDC20 with specific siRNA, but not treatment with proTAME, dramatically reduced securin and restored BIM protein levels in PAH PASMCs and PAAFs. Apcin restored BIM protein content in human PAH PASMCs, and siRNA BIM protected PAH PASMCs from apcin-induced apoptosis. Pharmacological targeting of CDC20 with specific PROTAC degrader CP5V down-regulated CDC20 and securin, restored BIM, inhibited hyper-proliferation, and induced apoptosis in PAH, but not in control PASMCs, and reversed SuHx-induced pulmonary vascular remodeling, PH, and right ventricular hypertrophy in male and female mice.
Conclusions: Collectively, our data demonstrate that CDC20 is overexpressed and supports proliferative, apoptosis resistant PASMC/PAAF phenotype in PAH via non-canonical APC/C independent upregulation of pro-proliferative securin and downregulation of pro-apoptotic BIM. Our data also suggest that targeting CDC20 could represent a novel potentially attractive strategy to suppress PA resident cells proliferation and reverse established PAH.