Stent-based delivery of AAV2 vectors encoding oxidation-resistant apoA1

Abstract

In-stent restenosis (ISR) complicates revascularization in the coronary and peripheral arteries. Apolipoprotein A1 (apoA1), the principal protein component of HDL possesses inherent anti-atherosclerotic and anti-restenotic properties. These beneficial traits are lost when wild type apoA1(WT) is subjected to oxidative modifications. We investigated whether local delivery of adeno-associated viral (AAV) vectors expressing oxidation-resistant apoA1(4WF) preserves apoA1 functionality. The efflux of 3H-cholesterol from macrophages to the media conditioned by endogenously produced apoA1(4WF) was 2.1-fold higher than for apoA1(WT) conditioned media in the presence of hypochlorous acid emulating conditions of oxidative stress. The proliferation of apoA1(WT)- and apoA1(4FW)-transduced rat aortic smooth muscle cells (SMC) was inhibited by 66% ± 10% and 65% ± 11%, respectively, in comparison with non-transduced SMC (p < 0.001). Conversely, the proliferation of apoA1(4FW)-transduced, but not apoA1(WT)-transduced rat blood outgrowth endothelial cells (BOEC) was increased 41% ± 5% (p < 0.001). Both apoA1 transduction conditions similarly inhibited basal and TNFα-induced reactive oxygen species in rat aortic endothelial cells (RAEC) and resulted in the reduced rat monocyte attachment to the TNFα-activated endothelium. AAV2-eGFP vectors immobilized reversibly on stainless steel mesh surfaces through the protein G/anti-AAV2 antibody coupling, efficiently transduced cells in culture modeling stent-based delivery. In vivo studies in normal pigs, deploying AAV2 gene delivery stents (GDS) preloaded with AAV2-eGFP in the coronary arteries demonstrated transduction of the stented arteries. However, implantation of GDS formulated with AAV2-apoA1(4WF) failed to prevent in-stent restenosis in the atherosclerotic vasculature of hypercholesterolemic diabetic pigs. It is concluded that stent delivery of AAV2-4WF while feasible, is not effective for mitigation of restenosis in the presence of severe atherosclerotic disease.

Intraprocedural endothelial cell seeding of arterial stents via biotin/avidin targeting mitigates in-stent restenosis

“…The therapeutic value of this approach can be increased when the seeded cells are genetically engineered to overexpress genes with proven anti-restenotic effects, i.e. nitric oxide synthase[48,49], VEGF[16] or apoA1[50,51].…”

Nucleic Acid Delivery to the Vascular Endothelium

“…In some cases, such as surgical interventions, ECs are accessible for local delivery of gene therapy agents directly to the site of injury. Gene therapy treatments for vascular diseases can be applied to vessels locally by embedding nanocarrier-encapsulated nucleic acids in intravascular stents or in hydrogels that can be implanted surrounding a vessel. , For patients receiving tissue allografts, the transplantation procedure provides a window of direct access to deliver gene therapies to vasculature within the organ graft . Both local and systemic delivery strategies have been further enhanced by the addition of cell-surface targeting molecules on delivery vehicles to promote retention and cellular uptake.…”

“…111 It is therefore often desirable to concentrate a gene therapy agent in specific regions of the vasculature. Transfection of specific vascular regions such as areas of stenosis, inflammation, or tumor-associated vascular growth can be achieved by local delivery 112,113 or by cell-specific targeting, 114 as described in the following sections. Examples of viral and nonviral vehicles designed for gene delivery to endothelial targets are summarized in Table 3.…”

“…In a recent study aiming to prevent in-stent restenosis (ISR) in pigs, gene delivery stents were generated by reversible immobilization of AAV vectors to the stent surface. 112 Vectors encoding a reporter gene successfully transduced ECs in stented arteries in vivo; however, in-stent delivery of vectors encoding apolipoprotein A1 (apoA1), a component of highdensity lipoprotein (HDL) that can prevent restenosis, did not reduce ISR in an atherosclerotic model in vivo. 112 Local delivery of AAV vectors to vasculature has also been pursued by suspending the vector in a biocompatible alginate hydrogel, which can be implanted around segments of the aorta.…”

“…112 Vectors encoding a reporter gene successfully transduced ECs in stented arteries in vivo; however, in-stent delivery of vectors encoding apolipoprotein A1 (apoA1), a component of highdensity lipoprotein (HDL) that can prevent restenosis, did not reduce ISR in an atherosclerotic model in vivo. 112 Local delivery of AAV vectors to vasculature has also been pursued by suspending the vector in a biocompatible alginate hydrogel, which can be implanted around segments of the aorta. While systemically delivered AAV vectors only transduced cells in the liver, implantation of an AAV9-containing hydrogel resulted in transduction of ECs and SMCs in the adjacent section of aorta with no transduction in other organs.…”