Abstract 18751: Inhalable Delivery of AAV-based Mrp4/abcc4 Silencing Rna Attenuates Monocrotaline-induced Pulmonary Hypertension

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The ATP-binding cassette transporter MRP4 (encoded by ABCC4) functions as a plasma membrane efflux pump to extrude cyclic nucleotides from various cell types. MRP4 regulates membrane cyclic nucleotides concentrations in arterial cells including smooth muscle cells (SMCs). In previous experiments, we have shown that MRP4 silencing inhibits SMCs proliferation in vitro. Concordantly, MRP4/ABCC4 deficient mice display a reduction in SMCs proliferation and a prevention of pulmonary hypertension in response to hypoxia. We here aimed to study gene transfer of a MRP4/ABCC4 silencing RNA via intratracheal delivery (using a MicroSprayer® Aerosolizer) of aerosolized adeno-associated virus 1 (AAV1.shMRP4 or AAV1.control) in a monocrotaline-induced model of pulmonary hypertension in rats. Gene transfer was performed at the time of monocrotaline administration and the effect on the development of pulmonary vascular remodeling was assessed thirty-five days later. The efficiency of this approach to transduce the lung vasculature was firstly assessed using inhalable AAV1 carrying LacZ encoding the β-galactosidase protein (AAV1.βGal, 1x10e11 DRP/animal). X-gal staining was clearly observed in bronchial smooth muscle cells and in the intima and media of large and small vessels. We then aimed to evaluate the effect of targeted vascular gene transfer of AAV1.shMRP4 on pulmonary hemodynamics and vascular remodeling. AAV1.shMRP4 dose-dependently reduced right ventricular systolic pressure and hypertrophy with a significant reduction with the higher doses (i.e., >10e11 DRP/animal) as compared to AAV1.control. The higher dose of AAV1.shMRP4 was also associated with a significant reduction in distal pulmonary arteries remodeling. AAV1.shMRP4 was finally associated with a reduction in the expression of ANF, a marker of cardiac hypertrophy. In conclusion, we showed that localized viral gene knockout of MRP4 in pulmonary vessels help to limit pulmonary vessels remodeling in a model of monocrotaline-induced PAH in rats. Collectively, these results support a therapeutic potential for downregulation of MRP4 for the treatment of pulmonary artery hypertension.

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