Abstract 2782: Targeted Delivery of Plasmid DNA by Ultrasound Following Acute Angioplasty
Introduction: Although stents have been shown to be able to deliver therapeutic drugs to the blood vessel wall, safety issues are still a concern. FDA-approved microbubble contrast reagents can be modified to carry therapeutic reagents, as well as DNA, and visualized in vivo by non-invasive ultrasound. Moreover, acoustic parameters can be altered to ``burst” microbubbles and increase cell permeability and reagent penetration into the target tissue. We tested the hypothesis that a reporter gene can be delivered to the vascular wall following balloon angioplasty via ultrasound-triggered microbubble delivery.
Methods: Lipid monolayer microbubbles formed by self-assembly during ultrasonic dispersion of decafluorobutane gas in an aqueous micellar mixture of phosphatidylcholine and PEG stearate with rapamycin (0.2 mg/ml). Positively charged microbubbles were coated with plasmid DNA expressing red fluorescent protein (CMV-RFP, 4 ug/ml). Acoustic parameters were optimized in vitro using rat aortic SMCs and applied in vivo in the rat carotid artery balloon injury model.
Results: Ultrasound-triggered (8 MHz, Mechanical Index = 1.9) release of CMV-RFP resulted in effective RFP expression. Release of rapamycin (final conc. 10 ng/ml) decreased proliferation by 65% in SMCs maintained in growth media. Microbubbles and/or ultrasound alone did not result in toxicity. CMV-RFP microbubbles were infused (2x107 bubbles/min) into the right jugular vein of a rat that had undergone balloon injury. Microbubbles were visualized in the injured left carotid artery by non-invasive ultrasound (Sequoia, Siemens). Using the same acoustic parameters to release reagents in vitro, focused ultrasound was delivered for 5 minutes to the injured blood vessel. 3 days after CMV-RFP delivery, the animal was euthanized and vessels were processed for frozen histological cross sectioning. RFP expression was visible by fluorescence microscopy in the inner-most SMC layer of the injured vessel and not in the contralateral vessel.
Conclusion: We demonstrate that CMV-RFP can be delivered to a blood vessel following angioplasty and that this method may be useful for clinical gene therapy as well as potential drug delivery to prevent in-stent restenosis.